IDES variant proteins and methods of using the same

Abstract

This disclosure relates generally to variant IdeS polypeptides having lower immunogenicity and preserved or improved IdeS stability and IgG cleavage activity, and methods of using the same.

Description

Attorney Docket No. : CYR-009WOIDES VARIANT PROTEINS AND METHODS OF USING THE SAMECROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to and the benefit of U.S. Provisional Patent Application No. 63 / 699,603 filed September 26, 2024, the contents of which are incorporated by reference herein in their entirety.STATEMENT REGARDING FEDERAL FUNDING

[0002] This invention was made with government support under 1R43 All 8898 awarded by National Institute of Allergy and Infectious Diseases (NIAID) of the National Institutes of Health (NIH). The government has certain rights in the invention.FIELD OF THE DISCLOSURE

[0003] This disclosure relates to variant IdeS polypeptides having lower immunogenicity and preserved or improved IdeS stability and IgG cleavage activity, and methods of using the same.BACKGROUND

[0004] IdeS (Immunoglobulin Degrading Enzyme from Streptococcus pyogenes) is a cysteine protease that catalyzes the rapid cleavage of all human IgG subclasses within the hinge region, severing the antigen binding domains of IgG from the Fc region that mediates immune effector functions. The decoupling of the antigen-binding Fab regions from the Fc moiety prevents Fc-mediated recruitment of immune effector functions to the antigen, including the recruitment of immune cells with killing capabilities (i.e. antibody-dependent cellular cytotoxicity or ADCC) and the deposition of activated complement (i.e. complementdependent cytotoxicity or CDC).

[0005] Due to the ability of IdeS to specifically target and cleave IgG, IdeS can significantly dampen antibody-mediated responses. This has been most comprehensively documented in the context of human kidney transplantation, where donor-specific antibodies (DSAs) in the transplant recipient recognize epitopes on the donor graft to drive transplant rejection. Patients with high levels of DSAs may be desensitized by administration of IdeS prior to transplantation, increasing success. Wild-type IdeS is used clinically for desensitization of kidney transplant recipients with high titers of anti-HLA donor-specificAttorney Docket No. : CYR-009WO antibodies, and may have broader applications in autoimmunity, gene therapy with viral vectors where antibodies are prevalent, and to reduce antibody mediated rejection of xenografts.

[0006] However, IdeS is immunogenic and its administration elicits anti-IdeS antibodies, preventing possible uses where repeated long-term dosing is required. Furthermore, S. pyogenes is a ubiquitous pathogen and is the cause of strep throat infections. Many people have mounted antibody defenses to S. pyogenes and IdeS through prior exposure. The presence and elicitation of anti-drug antibodies (AD As) is of great concern when developing therapeutics, as AD As have the potential to alter pharmacokinetics, drug activity, and bioavailability, or cause hypersensitivity reactions such as severe anaphylaxis.

[0007] Described herein are IdeS variant peptides that have lower immunogenicity, including reduced presentation of peptides on HLA-II and masking of surface epitopes that would otherwise be available for B cell recognition, and preserved or improved IdeS stability and IgG cleavage activity. Such reduced immunogenic IdeS variants may be used to expand the reach of an IdeS-like drug into indications where repeated dosing is necessary.SUMMARY OF THE DISCLOSURE

[0008] Provided herein are IdeS variant proteins comprising one or more than one amino acid modifications resulting in lower immunogenicity, measured as reduced presentation of peptides on HLA-II or masking of surface epitopes that are recognized by antibodies, and preserved or improved IdeS stability and IgG cleavage activity as compared to wild-type protein.

[0009] Described herein, in certain embodiments, is an IdeS variant protein comprising a modification at a position corresponding to position 189 of SEQ ID NO: 1. In some embodiments, the modification is a substitution of a threonine to a glycine. In some embodiments, the IdeS variant protein comprises at least 90% sequence identity to SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 95% sequence identity to SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 98% sequence identity to SEQ ID NO:2. In some embodiments, the IdeS variant protein further comprises a modification at a position corresponding to position 132 of SEQ ID NO: 1. In some embodiments, the modification is a substitution of glutamate to glutamine.Attorney Docket No. : CYR-009WOThe IdeS variant may be conjugated to a human serum albumin or a variant of human serum albumin as disclosed herein.

[0010] In some embodiments, in addition to the modifications corresponding to positions 189 or positions 189 and 132 of SEQ ID NO: 1, the IdeS variant protein further comprises one or more modifications at a position or positions corresponding to a position or position in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 166, 187, 213, 236, 277, 302, 303, 306, and 318. In some embodiments, the variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1. The IdeS variant may be conjugated to a human serum albumin or a variant of human serum albumin as disclosed herein.

[0011] In some embodiments, in addition to the modifications corresponding to positions 189 or positions 189 and 132 of SEQ ID NO: 1, the variant protein further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the variant protein comprises a modification at positions corresponding to positions 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. For example, in some embodiments, the variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1; a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1; a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1; a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1; a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1; a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1; a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1; a substitution of threonine to glycine at a position corresponding to position 189 of SEQ ID NO: 1; a substitution of glutamate to asparagine at aAttorney Docket No. : CYR-009WO position corresponding to position 198 of SEQ ID NO: 1; a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1; a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1. The IdeS variant may be conjugated to a human serum albumin or a variant of human serum albumin as disclosed herein.

[0012] In some embodiments, in addition to the modifications corresponding to positions 189 or positions 189 and 132 of SEQ ID NO: 1, the variant protein further comprises a substitution of the amino acid residues DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with an amino acid sequence selected from the group consisting of:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).

[0013] In some embodiments, the substitution of the amino acid residues DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 is the amino acid sequence QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85). In some embodiments, the IdeS variant protein comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2. The IdeS variant may be conjugated to a human serum albumin or a variant of human serum albumin as disclosed herein.

[0014] Described herein, in certain embodiments, is an IdeS variant protein comprising the amino acid sequence according to SEQ ID NO: 172 or SEQ ID NO: 173. For example, theAttorney Docket No. : CYR-009WOIdeS variant may comprise the amino acid sequence of SEQ ID NO: 172. In other embodiments, the IdeS variant may comprise the amino acid sequence of SEQ ID NO: 173. In some embodiments, the IdeS variant may comprise an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 172 or 173. In some embodiments, the IdeS variant may comprise an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 172 or 173. In some embodiments, the IdeS variant may comprise an amino acid sequence having at least 99% sequence identity with SEQ ID NO: 172 or 173. The IdeS variant may be conjugated to a human serum albumin or a variant of human serum albumin as disclosed herein.

[0015] Described herein, in certain embodiments, is an IdeS polypeptide conjugate comprising an IdeS variant according to any one of the embodiments described herein conjugated to a human serum albumin or a variant of human serum albumin.

[0016] In one embodiment, the IdeS polypeptide conjugate comprises a human serum albumin or variant human serum albumin and an IdeS variant protein comprising a modification at a position corresponding to position 189 of SEQ ID NO: 1. In some embodiments, the modification is a substitution of a threonine to a glycine. In some embodiments, the IdeS variant protein comprises at least 90% sequence identity to SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 95% sequence identity to SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 98% sequence identity to SEQ ID NO:2. In some embodiments, the IdeS variant protein further comprises a modification at a position corresponding to position 132 of SEQ ID NO: 1. In some embodiments, the modification is a substitution of glutamate to glutamine. The IdeS variant is conjugated to a human serum albumin or a variant of human serum albumin.

[0017] In one embodiment, the IdeS polypeptide conjugate comprises a human serum albumin or variant human serum albumin and an IdeS variant protein that, in addition to the modifications corresponding to positions 189 or positions 189 and 132 of SEQ ID NO: 1, further comprises one or more modifications at a position or positions corresponding to a position or position in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 166, 187, 213, 236, 277, 302, 303, 306, and 318. In some embodiments, the variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ IDAttorney Docket No. : CYR-009WONO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1. The IdeS variant is conjugated to a human serum albumin or a variant of human serum albumin.

[0018] In one embodiment, the IdeS polypeptide conjugate comprises a human serum albumin or variant human serum albumin and an IdeS variant protein that, in addition to the modifications corresponding to positions 189 or positions 189 and 132 of SEQ ID NO: 1, further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the variant protein comprises a modification at positions corresponding to positions 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. For example, in some embodiments, the variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1; a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1; a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1; a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1; a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1; a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1; a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1; a substitution of threonine to glycine at a position corresponding to position 189 of SEQ ID NO: 1; a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1; a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1; a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1. The IdeS variant is conjugated to a human serum albumin or a variant of human serum albumin.

[0019] In one embodiment, the IdeS polypeptide conjugate comprises a human serum albumin or variant human serum albumin and an IdeS variant protein that, in addition to theAttorney Docket No. : CYR-009WO modifications corresponding to positions 189 or positions 189 and 132 of SEQ ID NO: 1, the IdeS variant further comprises a substitution of the amino acid residues DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with an amino acid sequence selected from the group consisting of(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).

[0020] In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant where the substitution of the amino acid residues DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 is the amino acid sequence QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85). In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant comprising at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant comprising at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant comprising at least 98% sequence identity with residues 21-310 of SEQ ID NO:2. The IdeS variant is conjugated to a human serum albumin or a variant of human serum albumin.

[0021] In one embodiment, the IdeS polypeptide conjugate comprises a human serum albumin or variant human serum albumin and an IdeS variant protein comprising the amino acid sequence according to SEQ ID NO: 172 or SEQ ID NO: 173. For example, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 172. In other embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 173. In other embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprisingAttorney Docket No. : CYR-009WO at least 90% sequence identity to SEQ ID NO: 172 or SEQ ID NO: 173. In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 172 or 173. In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising an amino acid sequence having at least 99% sequence identity with SEQ ID NO: 172 or 173. The IdeS variant is conjugated to a human serum albumin or a variant of human serum albumin.

[0022] For example, in some embodiments, the human serum albumin or variant of human serum albumin of the IdeS polypeptide conjugate is conjugated to an N-terminal of the IdeS variant protein. In some embodiments, the human serum albumin or variant of human serum albumin is conjugated to a C-terminal of the IdeS variant protein. In some embodiments, the IdeS variant protein is conjugated to the human serum albumin or variant of human serum albumin using a linker. In some embodiments, the linker comprises GS, GSGGGSG (SEQ ID NO: 113), GSGSGSGS (SEQ ID NO: 114), GSGGGSGGGSG (SEQ ID NO: 115), or GSGSGSGSGSGS (SEQ ID NO: 116).

[0023] In some embodiments, the IdeS variant of the IdeS polypeptide conjugate protein is fused to the human serum albumin or variant of human serum albumin. In some embodiments, the human serum albumin or variant human serum albumin is selected from i) the amino acid sequence of SEQ ID NO: 188, ii) the amino acid sequence of SEQ ID NO: 189; iii) the amino acid sequence of SEQ ID NO: 190. For example, the human serum albumin may have the amino acid sequence of SEQ ID NO: 188. For example, the variant human serum albumin may have the amino acid sequence of SEQ ID NO: 189. For example, the variant human serum albumin may have the amino acid sequence of SEQ ID NO: 190. In some embodiments, the variant human serum albumin comprises an amino acid sequence having at least 99% sequence identity with SEQ ID NO: 188. In some embodiments, the IdeS variant protein comprises at least 95% sequence identity to SEQ ID NO: 172 or SEQ ID NO: 173.

[0024] In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 177. In other embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 186. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 187. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ IDAttorney Docket No. : CYR-009WONO: 154. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 155. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 156. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 157. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 158. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 159. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 160. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 161.

[0025] Described herein, in certain embodiments, is an IdeS polypeptide conjugate comprising an IdeS variant according to any one of the embodiments described herein conjugated to an albumin binding domain.

[0026] In one embodiment, the IdeS polypeptide conjugate comprises an albumin binding domain and IdeS variant protein comprising a modification at a position corresponding to position 189 of SEQ ID NO: 1. In some embodiments, the modification is a substitution of a threonine to a glycine. In some embodiments, the IdeS variant protein comprises at least 90% sequence identity to SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 95% sequence identity to SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 98% sequence identity to SEQ ID NO:2. In some embodiments, the IdeS variant protein further comprises a modification at a position corresponding to position 132 of SEQ ID NO: 1. In some embodiments, the modification is a substitution of glutamate to glutamine. The IdeS variant protein is conjugated to an albumin binding domain

[0027] In some embodiments, the IdeS polypeptide conjugate comprises an albumin binding domain and an IdeS variant protein that, in addition to the modifications corresponding to positions 189 or positions 189 and 132 of SEQ ID NO: 1, further comprises one or more modifications at a position or positions corresponding to a position or position in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 166, 187, 213, 236, 277, 302, 303, 306, and 318. In some embodiments, the variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQAttorney Docket No. : CYR-009WOID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1. The IdeS variant protein is conjugated to an albumin binding domain.

[0028] In one embodiment, the IdeS polypeptide conjugate comprises an albumin binding domain and an IdeS variant protein that, in addition to the modifications corresponding to positions 189 or positions 189 and 132 of SEQ ID NO: 1, further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the variant protein comprises a modification at positions corresponding to positions 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. For example, in some embodiments, the variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1; a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1; a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1; a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1; a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1; a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1; a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1; a substitution of threonine to glycine at a position corresponding to position 189 of SEQ ID NO: 1; a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1; a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1; a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1. The IdeS variant protein is conjugated to an albumin binding domain.

[0029] In one embodiment, the IdeS polypeptide conjugate comprises an albumin binding domain and an IdeS variant protein that, in addition to the modifications corresponding to positions 189 or positions 189 and 132 of SEQ ID NO: 1, the IdeS variant protein further comprises a substitution of the amino acid residues DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with an amino acid sequenceAttorney Docket No. : CYR-009WO selected from the group consisting of:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).

[0030] In some embodiments, the IdeS polypeptide conjugate comprises an albumin binding domain and an IdeS variant protein where the substitution of the amino acid residues DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 is the amino acid sequence QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85). In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising at least 98% sequence identity with residues 21-310 of SEQ ID NO:2. The IdeS variant protein is conjugated to an albumin binding domain.

[0031] In certain embodiments, the IdeS polypeptide conjugate comprises an albumin binding domain and an IdeS variant protein comprising the amino acid sequence according to SEQ ID NO: 172 or SEQ ID NO: 173. For example, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 172. In other embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 173. In other embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising at least 90% sequence identity to SEQ ID NO: 172 or SEQ ID NO: 173. In some embodiments, the IdeS polypeptide conjugate comprises an IdeS variant protein comprising an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 172 or 173. In some embodiments, the IdeSAttorney Docket No. : CYR-009WO polypeptide conjugate comprises an IdeS variant protein comprising an amino acid sequence having at least 99% sequence identity with SEQ ID NO: 172 or 173. The IdeS variant protein is conjugated to an albumin binding domain.

[0032] For example, in some embodiments, the albumin binding domain of the IdeS polypeptide conjugate is conjugated to an N-terminal of the IdeS variant protein. In some embodiments, the albumin binding domain is conjugated to a C-terminal of the IdeS variant protein. In some embodiments, the IdeS variant protein is conjugated to the albumin binding domain using a linker. In some embodiments, the linker comprises GS, GSGGGSG (SEQ ID NO: 113), GSGSGSGS (SEQ ID NO: 114), GSGGGSGGGSG (SEQ ID NO: 115), or GSGSGSGSGSGS (SEQ ID NO: 116).

[0033] In some embodiments, the IdeS variant protein of the IdeS polypeptide conjugate protein is fused to the albumin binding domain. In some embodiments, the albumin binding domain comprises the amino acid sequence of SEQ ID NO: 175. In some embodiments, the albumin binding domain comprises the amino acid sequence of SEQ ID NO: 176. In some embodiments, the albumin binding domain comprises an amino acid sequence having at least 90% sequence identity with amin acid sequence of SEQ ID NO: 175. In some embodiments, the albumin binding domain comprises an amino acid sequence having at least 90% sequence identity with amin acid sequence of SEQ ID NO: 176.

[0034] In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 178. In other embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 179. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 180. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 181. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 182. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 183. In some embodiments, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 184. In some embodiments, the IdeS polypeptide conjugate comprises at least 90% sequence identity with the amino acid sequence of any one of SEQ ID NOS: 178-184.

[0035] Also disclosed are polypeptides comprising an IdeS variant protein or wild-type IdeS and an albumin binding domain conjugated to the IdeS variant protein or the wild-typeAttorney Docket No. : CYR-009WOIdeS, wherein the albumin binding domain comprises an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 175 or SEQ ID NO: 176 or a human serum albumin or a human serum albumin variant.

[0036] Similarly, polypeptides are disclosed that comprise an IdeS variant protein and a human serum albumin or a human serum albumin variant.

[0037] The Ides variant protein of these polypeptides may comprise one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 113, 115, 116, 127, 128, 129, 130, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187, 188, 195, 197, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258, 273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.

[0038] In some embodiments, the Ides variant protein of these polypeptides may comprise one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111, 113, 115, 116, 127, 128, 129, 130, 132, 133, 148, 153,154, 159, 166, 167, 168, 175, 187, 188, 189, 195, 197, 198, 210, 213, 218, 219, 220, 228,233, 236, 241, 244, 245, 247, 258, 273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303,306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.

[0039] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303.

[0040] In some embodiments, the polypeptides may comprise an IdeS variant protein where the one or more modifications is as follows: i) the modification corresponding to position 68 of SEQ ID NO: l is a substitution of valine to threonine; ii) the modification corresponding to position 75 of SEQ ID NO: l is a substitution of alanine to proline; iii) the modification corresponding to position 166 of SEQ ID NO: l is a substitution of threonine to arginine or a substitution of threonine to glycine; iv) the modification corresponding to position 187 of SEQ ID NO: 1 is a substitution of serine to aspartate or a substitution of serine to glutamate; v) the modification corresponding to position 213 of SEQ ID NO: l is aAttorney Docket No. : CYR-009WO substitution of threonine to glutamate; vi) the modification corresponding to position 236 of SEQ ID NO:1 is a substitution of serine to cysteine; vii) the modification corresponding to position 277 of SEQ ID NO:1 is a substitution of leucine to cysteine; viii) the modification corresponding to position 302 of SEQ ID NO: l is a substitution of serine to aspartate, a substitution of serine to lysine, or a substitution of serine to glutamate; ix) the modification corresponding to position 303 of SEQ ID NO: l is a substitution of alanine to aspartate, a substitution of alanine to asparagine, or a substitution of alanine to glutamine; x) the modification corresponding to position 306 of SEQ ID NO: l is a substitution of valine to threonine, xi) the modification corresponding to position 318 of SEQ ID NO: 1 is a substitution of isoleucine to lysine, a substitution of isoleucine to aspartate, or a substitution of isoleucine to glycine; xii) the modification corresponding to position 308 of SEQ ID NO: 1 is a substitution of isoleucine to leucine; xiii) the modification corresponding to position 31 of SEQ ID NO: l is a substitution of serine to aspartate or asparagine; xiv) the modification corresponding to position 32 of SEQ ID NO: l is a substitution of phenylalanine to lysine; xv) the modification corresponding to position 33 of SEQ ID NO: l is a substitution of serine to glutamate; xvi) the modification corresponding to position 38 of SEQ ID NO: l is a substitution of isoleucine to valine; xvii) the modification corresponding to position 39 of SEQ ID NO:1 is a substitution of arginine to asparagine or threonine; xviii) the modification corresponding to position 43 of SEQ ID NO: l is a substitution of valine to glutamate; xix) the modification corresponding to position 44 of SEQ ID NO: l is a substitution of threonine to glutamate; xx) the modification corresponding to position 45 of SEQ ID NO: l is a substitution of proline to glutamate; xxi) the modification corresponding to position 47 of SEQ ID NO:1 is a substitution of histidine to lysine; xxii) the modification corresponding to position 54 SEQ ID NO:1 is a substitution of lysine to aspartate; xxiii) the modification corresponding to position 57 of SEQ ID NO: l is a substitution of threonine to lysine or glutamine; xxiv) the modification corresponding to position 60 of SEQ ID NO: l is a substitution of alanine to aspartate or glutamine; xxv) the modification corresponding to position 74 of SEQ ID NO: l is a substitution of valine to lysine; xxvi) the modification corresponding to position 75 of SEQ ID NO: l is a substitution of alanine to asparagine or proline; xxvii) the modification corresponding to position 77 of SEQ ID NO: l is a substitution of glutamine to glycine; xxviii) the modification corresponding to position 82 of SEQ ID NO: l is a substitution of isoleucine to methionine; xxix) the modification corresponding to position 85 of SEQ ID NO: l is a substitution of threonine to glutamine; xxx) the modification corresponding to position 111 of SEQ ID NO: l is a substitution ofAttorney Docket No. : CYR-009WO lysine to asparagine; xxxi) the modification corresponding to position 113 of SEQ ID NO: 1 is a substitution of glutamine to aspartate; xxxii) the modification corresponding to position 115 of SEQ ID NO: 1 is a substitution of lysine to histidine; xxxiii) the modification corresponding to position 116 of SEQ ID NO: l is a substitution of arginine to aspartate; xxxiv) the modification corresponding to position 127 of SEQ ID NO: 1 is a substitution of isoleucine to methionine; xxxiv) the modification corresponding to position 128 of SEQ ID NO: 1 is a substitution of asparagine to glycine; xxxvi) the modification corresponding to position 129 of SEQ ID NO: l is a substitution of phenylalanine to glutamate; xxxvii) the modification corresponding to position 130 of SEQ ID NO: l is a substitution of asparagine to glutamate; xxxviii) the modification corresponding to position 132 of SEQ ID NO: 1 is a substitution of glutamate for glutamine; xxxix) the modification corresponding to position 133 of SEQ ID NO: 1 is a substitution of glutamine at position 133 to aspartate; xl) the modification corresponding to position 148 of SEQ ID NO: 1 is a substitution of leucine to asparagine or arginine; xli) the modification corresponding to position 153 of SEQ ID NO: 1 is a substitution of phenylalanine to methionine or tyrosine; xlii) the modification corresponding to position 154 of SEQ ID NO: 1 is a substitution of glutamate to aspartate; xliii) the modification corresponding to position 159 of SEQ ID NO: 1 is a substitution of lysine to asparagine; xliv) the modification corresponding to position 167 of SEQ ID NO: 1 is a substitution of lysine to proline; xlv) the modification corresponding to position 168 of SEQ ID NO: 1 is a substitution of histidine to aspartate or proline; xlvi) the modification corresponding to position 175 of SEQ ID NO: 1 is a substitution of histidine to glutamine; xlvii) the modification corresponding to position 188 of SEQ ID NO: 1 is a substitution of leucine to glycine or methionine; xlviii) the modification corresponding to position 189 of SEQ ID NO: l is a substitution of threonine for glycine; xlix) the modification corresponding to position 195 of SEQ ID NO: 1 is a substitution of proline to aspartate; 1) modification corresponding to position 198 of SEQ ID NO: l is a substitution of glutamate to asparagine; li) the modification corresponding to position 197 of SEQ ID NO: 1 is a substitution of lysine to glutamate; lii) the modification corresponding to position 210 of SEQ ID NO: l is a substitution of alanine to glycine; liii) the modification corresponding to position 218 of SEQ ID NO: l is a substitution of serine to aspartate; liv) the modification corresponding to position 219 of SEQ ID NO: 1 is a substitution of lysine to glycine; Iv) the modification corresponding to position 220 of SEQ ID NO: 1 is a substitution of leucine to glutamine; Ivi) the modification corresponding to position 228 of SEQ ID NO: 1 is a substitution of lysine to glutamine; Ivii) the modification corresponding to position 233 of SEQ ID NO: l is aAttorney Docket No. : CYR-009WO substitution of lysine to serine; Iviii) the modification corresponding to position 241 of SEQ ID NO: 1 is a substitution of lysine to aspartate; lix) the modification corresponding to position 244 of SEQ ID NO: l is a substitution of threonine at position 244 to lysine; lx) the modification corresponding to position 245 of SEQ ID NO: l is a substitution of glutamate to aspartate; Ixi) the modification corresponding to position 247 of SEQ ID NO: l is a substitution of lysine to asparagine; Ixii) the modification corresponding to position 258 of SEQ ID NO: 1 is a substitution of valine to aspartate; Ixiii) the modification corresponding to position 273 of SEQ ID NO: l is a substitution of serine to aspartate; Ixiv) the modification corresponding to position 274 of SEQ ID NO: l is a substitution of asparagine to glutamate; Ixv) the modification corresponding to position 278 of SEQ ID NO: l is a substitution of lysine to aspartate or threonine; Ixvi) the modification corresponding to position 288 of SEQ ID NO: 1 is a substitution of asparagine to aspartate, glycine, or glutamine; Ixvii) the modification corresponding to position 289 of SEQ ID NO: l is a substitution of alanine to proline; Ixviii) the modification corresponding to position 297 of SEQ ID NO: l is a substitution of phenylalanine to aspartate; Ixix) the modification corresponding to position 299 of SEQ ID NO: l is a substitution of glycine to aspartate; Ixx) the modification corresponding to position 300 of SEQ ID NO: l is a substitution of valine to glutamate; Ixxi) the modification corresponding to position 307 of SEQ ID NO: l is a substitution of alanine to glycine or asparagine; Ixxii) the modification corresponding to position 313 of SEQ ID NO: 1 is a substitution of isoleucine to proline; Ixiii) the modification corresponding to position 314 of SEQ ID NO: 1 is a substitution of lysine to glycine; Ixxiv) the modification corresponding to position 315 of SEQ ID NO: 1 is a substitution of glutamate to proline; Ixxv) the modification corresponding to position 316 of SEQ ID NO: l is a substitution of aspartate to proline; Ixxvi) the modification corresponding to position 322 of SEQ ID NO: l is a substitution of valine to threonine; and / or; Ixxvii) the modification corresponding to position 330 of SEQ ID NO: l is a substitution of threonine to aspartate.

[0041] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a positionAttorney Docket No. : CYR-009WO corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0042] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0043] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1, and a substitution of valine to threonine at a position corresponding to position 306 of SEQ ID NO: 1.

[0044] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, a substitution of valine to threonine at a position corresponding to position 306 of SEQ ID NO: 1, and a substitution of isoleucine to lysine at a position corresponding to position 318 of SEQ ID NO: 1.Attorney Docket No. : CYR-009WO

[0045] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to cysteine at a position corresponding to position 236 of SEQ ID NO: 1, a substitution of leucine to cysteine at a position corresponding to position 277 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1, and a substitution of valine to threonine at a position corresponding to position 306 of SEQ ID NO: 1.

[0046] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to cysteine at a position corresponding to position 236 of SEQ ID NO: 1, a substitution of leucine to cysteine at a position corresponding to position 277 of SEQ ID NO: 1, and a substitution of isoleucine to leucine at a position corresponding to position 308 of SEQ ID NO: 1.

[0047] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1; a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1; a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1; a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1; a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1; a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1; a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1; a substitution of threonine toAttorney Docket No. : CYR-009WO glycine at a position corresponding to position 189 of SEQ ID NO: 1; a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1; a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1; a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0048] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising one or more glycosylation modifications as compared to wild-type. In some embodiments, the glycosylation modification is at a position corresponding to a position in SEQ ID NO: 1 selected from the group consisting of: 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, and 319. In some embodiments, i) the glycosylation modification occurs at a position corresponding to position 31 of SEQ ID NO: 1 where there is a substitution of serine to asparagine; ii) the glycosylation modification occurs at a position corresponding to position 37 of SEQ ID NO: 1 where there is a substitution of glutamate to asparagine; iii) the glycosylation modification occurs at a position corresponding to position 39 of SEQ ID NO: 1 where there is a substitution of arginine to threonine or asparagine; iv) the glycosylation modification occurs at a position corresponding to position 42 of SEQ ID NO: 1 where there is a substitution of glutamate to asparagine; v) the glycosylation modification occurs at a position corresponding to position 74 of SEQ ID NO: 1 where there is a substitution of valine to asparagine; vi) the glycosylation modification occurs at a position corresponding to position 76 of SEQ ID NO: 1 where there is a substitution of asparagine to serine; vii) the glycosylation modification occurs at a position corresponding to position 111 of SEQ ID NO: 1 where there is a substitution of lysine to asparagine; viii) the glycosylation modification occurs at a position corresponding to position 113 of SEQ ID NO: 1 where there is a substitution of glutamine to threonine; ix) the glycosylation modification occurs at a position corresponding to position 119 of SEQ ID NO: 1 where there is a substitution of glutamate to asparagine; x) the glycosylation modification occurs at a position corresponding to position 121 of SEQ ID NO: 1 where there is a substitution of histidine to threonine; xi) the glycosylation modification occurs at a position corresponding to position 130 of SEQ ID NO: 1 where there is a substitution of asparagine to serine; xii) the glycosylation modification occurs at a position corresponding to position 142 of SEQ ID NO: 1 where there is a substitution of aspartate to asparagine; xiii) the glycosylation modification occurs at a position corresponding to position 144 of SEQ IDAttorney Docket No. : CYR-009WONO: 1 where there is a substitution of lysine to threonine; xiv) the glycosylation modification occurs at a position corresponding to position 147 of SEQ ID NO: 1 where there is a substitution of glutamine to serine; xv) the glycosylation modification occurs at a position corresponding to position 148 of SEQ ID NO: 1 where there is a substitution of leucine to asparagine; xvi) the glycosylation modification occurs at a position corresponding to position 198 of SEQ ID NO: 1 where there is a substitution of glutamate to asparagine; xvii) the glycosylation modification occurs at a position corresponding to position 233 of SEQ ID NO: 1 where there is a substitution of lysine to serine; xviii) the glycosylation modification occurs at a position corresponding to position 244 of SEQ ID NO: 1 where there is a substitution of threonine to asparagine; xix) the glycosylation modification occurs at a position corresponding to position 246 of SEQ ID NO: 1 where there is a substitution of glycine to threonine; xx) the glycosylation modification occurs at a position corresponding to position 311 of SEQ ID NO: 1 where there is a substitution of lysine to asparagine; xxi) the glycosylation modification occurs at a position corresponding to position 313 of SEQ ID NO: 1 where there is a substitution of isoleucine to threonine; and / or xxii) the glycosylation modification occurs at a position corresponding to position 319 of SEQ ID NO: 1 where there is a substitution of glycine to serine.

[0049] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising: i) a substitution of glutamate to asparagine at a position corresponding to position 37 of SEQ ID NO: 1 and a substitution of arginine to threonine at a position corresponding to at position 39 of SEQ ID NO: 1; ii) a substitution of valine to asparagine at a position corresponding to at position 74 of SEQ ID NO: 1 and a substitution of asparagine to serine at a position corresponding to at position 76 of SEQ ID NO: 1; iii) a substitution of lysine to asparagine at a position corresponding to at position 111 of SEQ ID NO: 1 and a substitution of glutamine to threonine at a position corresponding to at position 113 of SEQ ID NO: 1; iv) a substitution of glutamate to asparagine at a position corresponding to at position 119 of SEQ ID NO: 1 and a substitution of histidine to threonine at a position corresponding to at position 121 of SEQ ID NO: 1; v) a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 1 and a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 1; vi) a substitution of threonine to asparagine at a position corresponding to position 244 of SEQ ID NO: 1 and a substitution of glycine to threonine at a position corresponding to position 246 of SEQ ID NO: 1; and / or vii) a substitution of lysine to asparagine at a position corresponding to positionAttorney Docket No. : CYR-009WO311 of SEQ ID NO: 1 and a substitution of isoleucine to threonine at a position corresponding to position 313 of SEQ ID NO: 1.

[0050] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising at least one modification at a position corresponding to a position in SEQ ID NO: 12 selected from the group consisting of: 47, 49, 51, 78, 111, 113, 123, 125, 126, 128, 142, 144, 148, 198, 273, 275, 278, 312, and 314 to introduce a glycosylation site. In some embodiments, i) the modification at a position corresponding to position 47 of SEQ ID NO: 12 is a substitution of threonine to asparagine; ii) the modification at a position corresponding to position 49 of SEQ ID NO: 12 is a substitution of glutamine to threonine or asparagine; iii) the modification at a position corresponding to position 51 of SEQ ID NO: 12 is a substitution of valine to threonine; iv) the modification at a position corresponding to position 78 of SEQ ID NO: 12 is a substitution of glycine to serine; v) the modification at a position corresponding to position 111 of SEQ ID NO: 12 is a substitution of lysine to asparagine; vi) the modification at a position corresponding to position 113 of SEQ ID NO: 12 is a substitution of glutamine to threonine; vii) the modification at a position corresponding to position 123 of SEQ ID NO: 12 is a substitution of glutamate to asparagine; viii) the modification at a position corresponding to position 125 of SEQ ID NO: 12 is a substitution of glutamine to threonine; ix) the modification at a position corresponding to position 126 of SEQ ID NO: 12 is a substitution of lysine to asparagine; x) the modification at a position corresponding to position 128 of SEQ ID NO: 12 is a substitution of asparagine to serine; xi) the modification at a position corresponding to position 142 of SEQ ID NO: 12 is a substitution of aspartate to asparagine; xii) the modification at a position corresponding to position 144 of SEQ ID NO: 12 is a substitution of lysine to threonine; xiii) the modification at a position corresponding to position 148 of SEQ ID NO: 12 is a substitution of leucine to asparagine; xiv) the modification at a position corresponding to position 198 of SEQ ID NO: 12 is a substitution of glutamate to asparagine; xv) the modification at a position corresponding to position 273 of SEQ ID NO: 12 is a substitution of serine to asparagine; xvi) the modification at a position corresponding to position 275 of SEQ ID NO: 12 is a substitution of glycine to serine; xvii) the modification at a position corresponding to position 278 of SEQ ID NO: 12 is a substitution of lysine to serine; xviii) the modification at a position corresponding to position 312 of SEQ ID NO: 12 is a substitution of glutamate to asparagine; or xix) the modification at a position corresponding to position 314 of SEQ ID NO: 12 is a substitution of lysine to serine.Attorney Docket No. : CYR-009WO

[0051] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising: i) a substitution of threonine to asparagine at a position corresponding to position 47 of SEQ ID NO: 12 and a substitution of glutamine to threonine at a position corresponding to position 49 of SEQ ID NO: 12.

[0052] I In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12 and a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12.

[0053] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of glutamate to asparagine at a position corresponding to position 123 of SEQ ID NO: 12 and a substitution of glutamine to threonine at a position corresponding to position 125 of SEQ ID NO: 12.

[0054] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12 and a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12.

[0055] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of serine to asparagine at a position corresponding to position 273 of SEQ ID NO: 12 and a substitution of glycine to serine at a position corresponding to position 275 of SEQ ID NO: 12.

[0056] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of glutamate to asparagine at a position corresponding to position 312 of SEQ ID NO: 12 and a substitution of lysine to serine at a position corresponding to position 314 of SEQ ID NO: 12

[0057] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 12, and a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 12.Attorney Docket No. : CYR-009WO

[0058] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0059] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 12, a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0060] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 12, a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0061] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0062] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of glutamine to asparagine at position corresponding to position 49 of SEQ ID NO: 12, a substitution of valine to threonine at a position corresponding to positionAttorney Docket No. : CYR-009WO51 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0063] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198.

[0064] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12, a substitution of asparagine to serine at a position corresponding to position 128, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0065] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of threonine to asparagine at a position corresponding to position 47 of SEQ ID NO: 12, a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0066] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of glutamine to asparagine at a position corresponding to positionAttorney Docket No. : CYR-009WO49 of SEQ ID NO: 12, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12; xvii) a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of glutamate to asparagine at a position corresponding to position 123 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 125 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0067] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0068] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of lysine to asparagine at position a position corresponding to 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0069] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a modification at a position corresponding to position 189 of SEQ ID NO: 1. In some embodiments, the modification is a substitution of threonine to a glycine at a position corresponding to position 189 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein comprises a modification at a position corresponding to position 132 of SEQ ID NO: 1. In some embodiments, the modification is a substitution of glutamate to glutamine at a position corresponding to a position 132 of SEQ ID NO: 1.Attorney Docket No. : CYR-009WO

[0070] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising further comprises a substitution of the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with a sequence selected from the group consisting of:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).

[0071] In some embodiments, the polypeptides may comprise an IdeS variant protein comprising a substitution of the amino acid residues DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with the amino acid sequence QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85). In some embodiments, the IdeS variant protein comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant protein comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0072] In some embodiments, the polypeptides may comprise a wild-type IdeS comprising an amino acid sequence according to SEQ ID NO: 1. In some embodiments, the polypeptides may comprise an IdeS variant protein comprising an amino acid sequence having at least 90% sequence identity with any one of SEQ ID NOs: 3-75, 86-94, 117-153, and 172-173. In some embodiments, the polypeptides may comprise an IdeS variant protein comprising an amino acid sequence having at least 95% sequence identity with any one of SEQ ID NOs: 3- 75, 86-94, 117-153, and 172-173.

[0073] In some embodiments, the polypeptides comprising a IdeS variant that also comprise an albumin binding domain comprise an albumin binding domain comprising the amino acid sequence of SEQ ID NO: 176 or 176.Attorney Docket No. : CYR-009WO

[0074] In some embodiments, the polypeptides comprising a IdeS variant that also comprise a human serum albumin comprise a human serum albumin having the amino acid sequence of SEQ ID NO: 188. In other embodiments, the variant human serum albumin comprises the amino acid sequence of SEQ ID NO: 189. In still other embodiments, the variant human serum albumin comprises the amino acid sequence of SEQ ID NO: 190. In still further embodiments, the variant human serum albumin comprises an amino acid sequence having at least 99% sequence identity with SEQ ID NO: 188.

[0075] Described herein, in certain embodiments, is an IdeS variant comprising the amino acid sequence of any one of SEQ ID NOs:3-75, 86-94, 117-153, 172, or 173.

[0076] The IdeS variant protein, IdeS polypeptide conjugate, or polypeptide according to any of the preceding embodiments may have decreased presentation of epitopes on human leukocyte antigen (HLA) as compared to a wild-type IdeS protein. The IdeS variant protein, IdeS polypeptide conjugate, or polypeptide according to any of the preceding embodiments may have increased cleavage activity of immunoglobulins as compared to a wild-type IdeS protein. The IdeS variant protein, IdeS polypeptide conjugate, or polypeptide according to any of the preceding embodiments may have a decrease in immunogenicity score more than 0 relative to SEQ ID NO: 1 or SEQ ID NO:2.

[0077] Described herein are polynucleotides encoding the IdeS variant protein, IdeS polypeptide conjugate, or polypeptide according to any of the preceding embodiments. Also described herein is an expression plasmid comprising such polynucleotides and a promoter according to the preceding embodiments. Also, described herein, is a cell comprising such polynucleotides according to the preceding embodiments.

[0078] Described herein is pharmaceutical composition comprising the IdeS variant protein, IdeS polypeptide conjugate, or polypeptide according to any of the preceding embodiments and a pharmaceutically acceptable carrier.

[0079] Described herein, in certain embodiments, is a method of treating an autoantibody- mediated autoimmune disease or disorder comprising administering an effective amount of the IdeS variant protein, IdeS polypeptide conjugate, or polypeptide according to any one of the preceding embodiments, or the pharmaceutical composition of any one of the preceding embodiments to a subject in need thereof. In some embodiments, the autoantibody-mediated autoimmune disease is selected from Addison’s disease, Anti-glomerular basementAttorney Docket No. : CYR-009WO membrane antibody disease (ABM, also known as Goodpasture’s disease), Axial Spondylarthritis, Bullous Pemphigoid, celiac disease, chronic inflammatory demyelinating polyneuropathy (CIDP), dermatomyositis, Fetal and Neonatal Alloimmune Thrombocytopenia (FNAIT), generalized myasthenia gravis (gMG) and myasthenic crisis, Grave’s Disease, Guillain-Barre syndrome (GBS), Hashimoto’s thyroiditis, hemolytic disease of the fetus and newborn (HDFN), heparin induced thrombocytopenia (HIT), immune thrombocytopenia (ITP), inflammatory bowel disease (IBD) including Crohn’s disease and ulcerative colitis, Lupus Nephritis, Membranous Nephritis, multiple sclerosis, Myasthenia Gravis, myositis, MOG antibody disease (MOGAD), Obstetric Antiphospholipid Syndrome (OAPS), Pemphigus, Relapsing Polychondritis, psoriatic arthritis, psoriasis, rheumatoid Arthritis, scleroderma, Sjogren’s syndrome, stiff person syndrome, systemic lupus erythematosus (SLE), systemic sclerosis, Thrombotic Thrombocytopenic Purpura (TTP), thyroid eye disease (TED), vitiligo, vasculitis, warm autoimmune hemolytic anemia (wAIHA). In some embodiments, the autoantibody-mediated autoimmune disease is heparin induced thrombocytopenia (HIT). In some embodiments, the autoantibody-mediated autoimmune disease is immune thrombocytopenia (ITP). In some embodiments, the IdeS variant protein, IdeS polypeptide conjugate, polypeptide, or pharmaceutical composition is co-administered with a gene therapy.

[0080] Described herein, in certain embodiments, is a method for reducing presentation on HLA-II of IdeS comprising administering an effective amount of the IdeS variant protein, IdeS polypeptide conjugate, or polypeptide according to any one of the preceding embodiments, or the pharmaceutical composition of any one of the preceding embodiments

[0081] Described herein, in certain embodiments, is a method for prolonging serum halflife of IdeS comprising administering an effective amount the IdeS variant protein, IdeS polypeptide conjugate, or polypeptide according to any one of the preceding embodiments, or the pharmaceutical composition of any one of the preceding embodiments

[0082] Described herein, in certain embodiments, is a method for prolonging IgG depletion of IdeS comprising administering an effective amount of the IdeS variant protein, IdeS polypeptide conjugate, or polypeptide according to any one of the preceding embodiments, or the pharmaceutical composition of any one of the preceding embodimentsAttorney Docket No. : CYR-009WOBRIEF DESCRIPTION OF THE DRAWINGS

[0083] FIGs. 1A-1B depict the identification of HLA-II epitopes in IdeS by MAPPs. IdeS expressed and purified (FIG. 1A) as a secreted protein in Expi293F culture or (FIG. IB) as an intracellular protein in E. coli culture was added to dendritic cells (DCs) from 10 healthy donors and processed IdeS peptides bound to immunoprecipitated HLA-DR complexes were identified by mass spectrometry. The frequency with which IdeS residues are found in HLA- DR-bound peptides is indicated by light (low) to dark (high frequency) colors. The summed total across the 10 donors is shown in the bottom row of each panel.

[0084] FIGs. 2A-2B depict the HLA-DR epitope landscape of IdeS determined by yeast display resembles MAPPs analysis. FIG. 2A (left panel) depicts an experimental schema. Walking along the IdeS sequence by 5 residues per step, 15-mer IdeS peptides (dark green) were expressed as fusions to Aga2p for yeast surface display. Induced yeast were incubated at acidic pH with soluble CLIP -bound HLA-II complexes. In the presence of HLA-DM, CLIP is exchanged for a high affinity peptide epitope displayed on the yeast surface. HLA-II bound to the yeast and peptide expression tags are stained with fluorescent reagents. Using flow cytometry (plot at right of FIG. 2A), yeast are gated for peptide expression (green; this partially controls for differences in expression levels across different peptide sequences) and bound HLA-II is measured. FIG. 2B depicts data of binding signals of different HLA-DR alleles to yeast displaying peptides rastered across the IdeS sequence that are normalized to positive control peptides and plotted. Shown at bottom are MAPPs peptide counts summed across Donors 1-10 (Expi293F produced IdeS) and Donors 11-20 (A. coli produced IdeS).

[0085] FIG. 3 depicts reduction of HLA-II epitopes in engineered variants of IdeS. Peptide walks across the sequences of Wt IdeS (grey), Variant 10.9 (teal), and Variant 116 (green) were combined with yeast display to assess the binding of HLA-II alleles. Binding signals (AMean Fluorescence Intensity, AMFI) from flow cytometry are normalized to control peptides. Notable epitopes in Wt IdeS that were reduced in Variant 116 are indicated by red asterisks.

[0086] FIG. 4 depicts an ML-based algorithm overpredicts reduction of T cell epitopes. Merged yeast display data for the binding of HLA-DR alleles (DRB1, DRB3, DRB4, DRB7, and DRB15) to peptides rastered across the sequences of Wt IdeS (grey) and Variant 10.9 (teal) are compared to predictions from NetMHCIIpan-4.1.Attorney Docket No. : CYR-009WO

[0087] FIG. 5 depicts screening mutations in glycosylated IdeS variants for catalytic activity. Single amino acid substitutions were introduced into Variant 10.9 (left panel of FIG. 5) and Variant 10.2-HSA (right panel of FIG. 5), the proteins were expressed in Expi293F culture, and the expression medium was screened for cleavage activity against a monoclonal human IgGl substrate. Cleavage products are resolved on Coomassie-stained non-reducing SDS gels. Mutations were rationally designed based on inspection of structural models and sequence logos of HLA-II peptide preferences.

[0088] FIG. 6 depicts screening mutations in glycosylated IdeS variants for HLA-II epitope reduction. For each mutation under consideration, a 17-mer peptide from Variant 10.9 with the mutation in the center was expressed on yeast and binding to the indicated HLA-II alleles was measured by flow cytometry.

[0089] FIGs. 7A-7B demonstrate that IdeS variants from a final round of T cell deimmunization have acceptable thermostability and catalytic activity. FIG. 7A depicts data of melting temperatures of purified Variant 10.9 with the indicated mutations were measured by DSF. Mutations E132Q and T189G had minimal effect on thermostability and were considered further. FIG. 7B depicts data from single and double mutants of Variant 10.9 (at left) and Variant 10.2-HSA (at right) that were incubated with a monoclonal human IgGl substrate. Cleavage products were separated by non-reducing gel electrophoresis.

[0090] FIGs. 8A-8H demonstrate that engineered IdeS variants cleave all human IgG subclasses. Variant 116 and Variant 117-HSA were purified by NiNTA affinity chromatography and 4 pg of each protein was loaded on a Coomassie-stained SDS gel (FIG. 8A). Variant 116 eluted from NiNTA resin was separated by size exclusion chromatography on a Superdex 75 increase 10 / 300 column (FIG. 8B). Variant 117-HSA eluted from NiNTA resin was separated by size exclusion chromatography on a Superdex 200 increase 10 / 300 column (FIG. 8C). Human IgG subclasses (3 pM) were incubated with 3 nM Wt IdeS (FIG. 8D), Variant 116 (FIG. 8E), or Variant 117-HSA (FIG. 8F) at 37 °C and cleavage products were analyzed by non-reducing gel electrophoresis over time. Cleavage assay of IgG from human, cynomolgus monkey, rabbit, rat, and mouse by Variant 116 (FIG. 8G) or Variant 117-HSA (FIG. 8H).

[0091] FIG. 9 demonstrates that human-mouse chimeric IgG antibodies are IdeS substrates. Non-reducing SDS-PAGE of IdeS-catalyzed cleavage products of human IgGlAttorney Docket No. : CYR-009WO(hlgGl), rituximab, and a human-mouse chimera (matches chimeric IgGl antibodies produced by the Tg32-hFc mouse strain56). [Substrates] = 2 pM and [IdeS] = 100 nM.

[0092] FIG. 10 demonstrates that IdeS variants cleave B cell receptors. Human primary CD 19+ B cells were treated with 1 pM wild type or variant IdeS enzymes in culture. IgG (dark grey), IgM (white, dotted), and IgA (light grey) class BCR+ cells were gated and the percent that were positive for the Fab domain were quantified.

[0093] FIGs. 11A-11D demonstrate that IdeS variants with extended PK are effective at low doses. NZW rabbits (N=2 per time point) were IV administered (FIG. 11 A) Variant 116 or (FIG. 11B) Variant 10.2-HSA at 1 mg / kg (filled grey circles), 0.1 mg / kg (filled black squares), or 0.01 mg / kg (open triangles). IgG was quantified in serum by ELISA. Serum concentrations of test articles were measured by ELISA in animals treated with (FIG. 11C) Variant 116 or (FIG. HD) Variant 10.2-HSA at 3 doses. Only measurements above the lower limit of quantification (LLOQ) are plotted. All data are mean ± SD.

[0094] FIGs. 12A-12B demonstrate that IgG rebound is in conflict with persistent degradation catalyzed by IdeS variants. Anti-IgG Fab western blot analysis of serum collected at different time points from rabbits administered Variant 116 (FIG. 12A) or Variant 10.2-HSA (FIG. 12B) at 3 doses: 0.01 mg / kg (upper blots), 0.1 mg / kg (center blots), and 1 mg / kg (lower blots). Individual animals (N=4) in the treatment groups are indicated by patterned circles above each lane.

[0095] FIGs. 13A-13C demonstrate that wild type IdeS elicits AD As in rabbits. Serum samples from NZW rabbits that were IV administered E. ^ / / -produced IdeS (1 mg / kg) were analyzed by ELISA for the presence of anti-IdeS AD As. AD As were detected with anti-IgM (FIG. 13A), anti -F(ab')2 (FIG. 13B), or anti-IgG (FIG. 13C) detection antibodies. There were 2 rabbits in the study, indicated by grey and black bars. Individual cut points were determined for each animal and serum samples were considered ADA positive if there was a 95% confidence they were above the cut point (denoted by asterisks).

[0096] FIGs. 14A-14B demonstrate that a hyperglycosylated IdeS variant has low immunogenicity in rabbits. Variant 116 was IV injected into NZW rabbits at 1 mg / kg (top), 0.1 mg / kg (center), or 0.01 mg / kg (bottom) and anti-Variant 116 AD As in serum samples were detected by ELISA using anti-IgM (FIG. 14A) or anti-F(ab')2 (FIG. 14B) detectionAttorney Docket No. : CYR-009WO antibodies. There were 4 rabbits per treatment group with 2 rabbits alternating per time point, indicated with grey and black bars. Individual cut points were determined for each animal and serum samples were considered ADA positive if there was a 95% confidence they were above the cut point (denoted by asterisks).

[0097] FIGs. 15A-15B demonstrate that ADA responses towards an HSA-fused IdeS variant are dependent on dose. NZW rabbits were IV administered Variant 10.2-HSA at 1 mg / kg (top), 0.1 mg / kg (center), or 0.01 mg / kg (bottom). ELISA plates were coated with Variant 10.2-HSA and incubated with rabbit serum. ADA responses were detected with anti- IgM (FIG. 15A) or anti-F(ab')2 (FIG. 15B). There were 4 rabbits per treatment group with 2 rabbits alternating per time point, indicated with grey and black bars. Individual cut points were determined for each animal and serum samples were considered ADA positive if there was a 95% confidence they were above the cut point (denoted by asterisks).

[0098] FIG. 16 demonstrates that the emergence of AD As is associated with a sudden decline of an HSA-fused IdeS variant in serum. Serum concentrations of Variant 10.2-HSA (filled squares and broken black line, left axis) and ADA responses (open triangles with black line and filled circles with grey line, right axis) are plotted for individual rabbits receiving 1 mg / kg (upper plots) or 0.1 mg / kg doses (lower plots). PK measurements are only plotted if they were above the lower limit of quantification (0.01 pg / ml).

[0099] FIGs. 17A-17D depict rabbit AD As towards an HSA-fused IdeS variant target theHSA moiety. Serum samples collected at the indicated times from rabbits treated with Variant 10.2-HSA (1 mg / kg dose) were pre-incubated with competitor: no competitor control (black), Variant 10.2-HSA (grey), HSA (white), and Variant 10.2 without a fusion partner (crosshatched). The serum samples were then added to ELISA plates coated with Variant 10.2-HSA and bound AD As were detected with anti-IgM (FIG. 17A) or anti-F(ab')2 (FIG. 17B) detection antibodies. Alternatively, ELISA plates were coated with Variant 10.2-HSA (grey), HSA (white), or Variant 10.2 without a fusion partner (crosshatched). Plates were incubated with rabbit serum and bound AD As were detected with anti-IgM (FIG. 17C) or anti-F(ab')2 (FIG. 17D) detection antibodies.

[0100] FIGs. 18A-18D depict the PK of wild type IdeS and variants in inbred mice.Single doses of IdeS proteins were administered at 1 mg / kg and concentrations in serum were measured by ELISA. Data are mean ± SEM from male mice (6-8 weeks old). FIGs. 18A-18BAttorney Docket No. : CYR-009WO demonstrate PK in C57B1 / 6 mice of Variant 10.9 (open symbols and solid black line, FIG. 18A) and Variant 10.2-HSA (open symbols and solid black line, FIG. 18B) versus WT Ides (grey squares and line, E. coli produced FIGs. 18A-18B). Proteins treated with Arthrobacter ureafaciens neuraminidase (NA) are shown with black symbols and broken lines. N=3 per time point, IV route. FIG. 18C demonstrates PK of Variant 10.2-HSA in C57B1 / 6 mice (solid line) versus Tg32 mice (broken line). N=2 per time point, IV route. FIG. 18D demonstrates PK of Variant 117-HSA in C57B1 / 6 mice via IV (grey line and symbols), hFcRn / Alb-KO mice via IV (open symbols and black solid line), or hFcRn-Alb-KO mice via SC (black symbols and broken lines). N=3 per time point. See Table 4.

[0101] FIG. 19 demonstrates that glycosylated IdeS variants have heterogenous, sialylated glycoforms. IEF analysis of IdeS variants (20 pg per lane). Theoretical pl values are in parentheses. IdeS (Wt,EC) was produced from E. coli, all other proteins were purified from Expi293F culture. Variant 10.14 is missing N-glycosylation sites and represents charge heterogeneity before glycans are added.

[0102] FIGs. 20A-20D demonstrate reduced immunogenicity of an IdeS variant following a second dose. IgM class AD As and IgG class AD As directed towards the respective test articles were detected by ELISA in serum from rabbits dosed IV on Day 0 and Day 14 with 1 mg / kg WT IdeS (FIG. 20A), 0.01 mg / kg WT IdeS (FIG. 20B), 1 mg / kg Variant 116 (FIG. 20C), or 0.01 mg / kg Variant 116 (FIG. 20D). OD450 measurements are plotted for individual animals, N=3 per group. Asterisks indicate ADA positive samples with 95% confidence.

[0103] FIGs. 21A-21-D demonstrate low ADA titers after two doses of an engineered IdeS variant in the rabbit model. Reciprocal ADA titers are plotted for individual rabbits dosed IV on Day 0 and Day 14 with 1 mg / kg WT IdeS (FIG. 21 A), 0.01 mg / kg WT IdeS (FIG. 21B), 1 mg / kg Variant 116 (FIG. 21C), or 0.01 mg / kg Variant 116 (FIG. 21D). N=3 per group.

[0104] FIGs. 22A-22B demonstrate an IdeS variant with low immunogenicity is not neutralized after a second dose in the rabbit model. On Days 0 and 14, rabbits were IV administered WT IdeS (black) or Variant 116 (grey) at 1 mg / kg (solid lines, filled symbols) or 0.01 mg / kg (broken lines, open symbols). IgG levels as a percent of the baseline were measured by ELISA and are plotted in FIG. 22A. N=3 per group, data are mean ± SD. Anti-Attorney Docket No. : CYR-009WOIgG western blots of serum samples from single rabbits that are representative of each treatment group are shown in FIG. 22B.

[0105] FIGs. 23A-23E demonstrate a rabbit albumin fused IdeS variant has long PK-PD and low immunogenicity. FIG. 23A shows serum IgG levels in rabbits IV administered Variant 117-RSA at 1 mg / kg (solid line and filled symbols) or 0.1 mg / kg (broken line and open symbols), N=2 per group. FIGs. 23B-23C show ELISA-based detection of IgM class AD As (FIG. 23B) and IgG class AD As (FIG. 23C) in serum from rabbits treated with human albumin (black columns) or rabbit albumin (grey columns) fused Variant 117. Measurements from individual animals are plotted and positive values with 95% confidence are indicated with asterisks. FIG. 23D shows reciprocal ADA titers in rabbits treated with WT IdeS (black columns) versus Variant 117-RSA (grey columns). FIG. 23E shows serum concentrations of human albumin (black) or rabbit albumin (grey) fused Variant 117 after IV administration at 1 mg / kg (solid lines and filled symbols) or 0.1 mg / kg (broken lines and open symbols). Data are plotted for individual animals up until time points at which concentrations are below LLOQ.

[0106] FIGs. 24A-24J demonstrate a half-life extended albumin-fused IdeS variant is effective and has low immunogenicity after three doses. FIG. 24A depicts the study design, N=3 NZW rabbits per group. Triangles indicate doses via IV route. Rabbits were dosed 3 times with 10 pg / kg WT IdeS (FIGs. 24B-24D), 10 pg / kg Variant 117-RSA (FIGs. 24E- 24G), or 0.3 pg / kg Variant 117-RSA (FIGs. 24H-24J). Serum samples at the indicated days were analyzed by western blot to qualitatively visualize changes in IgG levels for a representative individual animal from each group (FIGs. 24B, 24E, and 24H). Serum samples were analyzed by ELISA for IgM class AD As (FIGs. 24C, 24F, and 241) and IgG class AD As (FIGs. 24D, 24G, and 24J). For ADA analysis, ELISA signals are plotted for individual animals.

[0107] FIG. 25 demonstrates ADA titers elicited by repeat doses of an albumin-fused IdeS variant are substantially lower compared to WT IdeS. Reciprocal ADA titers were determined by ELISA using 2-fold serial dilutions from serum samples collected from rabbits treated with 3 biweekly doses of WT IdeS at 10 pg / kg (black circles, black solid line), Variant 117-RSA at 10 pg / kg (grey squares, grey solid line), or Variant 117-RSA at 0.3Attorney Docket No. : CYR-009WO pg / kg (open diamond, broken line). Data are plotted on a log scale in the inset, mean ± SEM, N=3.

[0108] FIGs. 26A-26D demonstrate that an albumin-fused IdeS variant manufactured from stable CHO-K1 pools is highly sialylated and catalytically active. Variant 117-HSA proteins, with cysteine, serine or alanine at position 34 of the HSA moiety, were purified from stable CHO-K1 pools and compared to research material produced from transiently transfected Expi293F cells. Neu5Gc (dark grey) and Neu5Ac (light grey) content are plotted in FIG. 26A as moles per mole of protein. N-Glycan composition was also determined by LC-MS. The ratio of glycan branches ending in terminal sialic acid residues to branches that end in other sugars is shown in FIG. 26B. N-Glycan structures are compared between research grade Variant 117-HSA and Variant 117-HSA(C34S) manufactured from stable CHO-K1 in FIG. 26C. A0 to A4 specifies the number of antennae / branches. Catalytic activity was measured by incubating the IdeS proteins with a monoclonal human IgGl and quantifying the formation of the F(ab')2 fragment with HPLC (FIG. 26D).

[0109] FIGs. 27A-27B depict PK-PD in the rabbit model for representative drug substance of an albumin-fused IdeS variant. PK (FIG. 27A) and IgG levels (FIG. 27B) in rabbits administered 0.1 mg / kg Variant 117-HSA (black lines) or Variant 117-HSA(C34S) (grey line) manufactured from stable CHO-K1 culture. Proteins were administered IV (solid lines and filled symbols) or SC (broken line and open symbols). Data are mean ± SEM, with parallel sampling between 2 cohorts of N=2 (N=4 total per group).

[0110] FIG. 28 depicts the HLA-II epitope landscapes of de novo designed ABDs. Using yeast display with flow cytometry, 15-mer peptides staggered by 3 amino acids covering the full sequences of ABD5 (left) and ABD7 (right) were evaluated for loading onto 7 HLA-II alleles (indicated on right). Delta mean fluorescence intensities were normalized to positive control peptides specific to each HLA-II allele.

[0111] FIGs. 29A-29E demonstrates that IdeS-ABD fusion proteins are stable monomers. His-tagged proteins were purified by NiNTA affinity chromatography and separated by SEC on a Superdex 75 10 / 300 column. Elution traces are shown for ABD5 and ABD7 fusions to Variant 10.9 (" 10.9") (FIG. 29A), Variant 10.2 (" 10.2") (FIG. 29B), and Wt IdeS ("Wt") (FIG. 29C). FIG. 29D depicts DSF analysis of IdeS-ABD fusion proteins showing cooperative unfolding transitions. FIG. 29E depicts data of melting temperatures. N=4.Attorney Docket No. : CYR-009WO

[0112] FIGs. 30A-30C demonstrates that de novo designed ABDs fused to IdeS bind serum albumin. BLI kinetics of immobilized IdeS-ABD proteins binding to human (FIG. 30A) or murine albumin (FIG. 30B) (120 nM). Association was measured from 0-500 s and dissociation from 500-1200 s. FIG. 30C depicts data of the catalytic activity of IdeS-ABD fusion proteins. Polyclonal human IgG (10 pM) was incubated with 200 nM IdeS-ABD for the indicated times and cleavage products were separated by non-reducing SDS-PAGE. In the reactions shown at bottom, 5 pM HSA was added to saturate ABD binding.

[0113] FIGs. 31A-31C demonstrate that Variant 116-ABD7 is a stable monomer. FIG. 31 A depicts reducing SDS-PAGE analysis of Variant 116-ABD7 (4 pg) purified by NiNTA affinity chromatography from Expi293F culture. FIG. 31B depicts SEC separation of the sample in panel A on a Superdex 75 increase 10 / 300 column. FIG. 31C depicts data (mean and standard deviation) of thermal melt of Variant 116 (unfused in grey, ABD7 fused in black) measured by DSF. N = 3.

[0114] FIGs. 32A-32B demonstrate that Variant 116-ABD7 cleaves human IgG subclasses and human-mouse chimeric IgG. Human IgG subclasses (3 pM) were incubated with Variant 116-ABD7 (3 nM) and cleavage products over time were analyzed by nonreducing SDS-PAGE (FIG. 32A). Human IgGl (hlgGl), rituximab, and a human-mouse IgGl chimera (matches the Tg32-hFc mouse strain) (2 pM) were incubated with Variant 116- ABD7 (100 nM) and cleavage products were analyzed by non-reducing SDS-PAGE (FIG.32B)

[0115] FIGs. 33A-33B demonstrate that Variant 116-ABD7 binds albumin. FIG. 33A depicts BLI sensorgrams of immobilized Variant 116-ABD7 dipped in 60 nM solutions of human (black) or murine (grey) serum albumin from 0-300 s and then returned to assay buffer from 300-1200s. FIG. 33B depicts Electrophoretic mobility shift assays of Variant 116-ABD7 (titrated from 19 to 0 pM) incubated with 2.5 pM serum albumin of human (left), mouse (center), and cow (right). Mixtures were analyzed by native PAGE for shift of albumin, which runs around the 100 kD MW marker under these conditions, indicative of complex formation with reduced electrophoretic mobility.

[0116] FIG. 34 demonstrates PK of an ABD fused IdeS variant. Variant 116-ABD7 was administered to C57B1 / 6 mice (IV, 1 mg / kg) and concentration in serum was determined by ELISA. Data are mean ± SEM, N=3 per time point.Attorney Docket No. : CYR-009WO

[0117] FIGs. 35A-35B demonstrate that an engineered IdeS variant efficiently cleaved heparin-induced thrombocytopenia (HIT) immune complexes. FIG. 35A depicts data of cleavage of antigen-free 5B9, a monoclonal antibody that binds PF4 / H to cause HIT. Solutions of 5B9 were untreated (broken grey line with open symbols) or treated overnight at 37 °C with 10 pg / ml Variant 117-HSA (solid black line with filled symbols). The solutions were then incubated on ELISA plates coated with 2.5 pg / ml PF4 plus 0.1 U / ml unfractionated heparin. Intact, bound 5B9 was detected with anti-HsFcy antibody that recognizes the Fc region of human IgG. FIG. 35B depicts data of cleavage of 5B9 / PF4 / H immune complexes. The HIT monoclonal antibody 5B9 was pre-bound to a PF4 / H coated ELISA plate for 1 hour prior to treatment with 10 pg / ml Variant 117-HSA overnight at 37°C. Intact, bound 5B9 was detected with anti-HsFcy antibody.

[0118] FIGs. 36A-36B demonstrate prophylactic and therapeutic efficacy of an engineered IdeS variant in a mouse model of immune thrombocytopenia (ITP). FIG. 36A depicts data of platelet counts in blood after C57BL / 6 mice were injected intraperitoneally (IP) with 0.02 mg rabbit anti-mouse platelet IgG at Time = 0 h (all groups). Group 2 and Group 3 received a second injection of anti-platelet IgG at 48 h and 72 h, respectively. Data are mean ± SEM, N=3. Anti-platelet IgG dosing schedule is indicated at top with arrowheads on a timeline. FIG. 36B depicts data of platelet counts in blood of C57BL / 6 mice that were injected IP with 0.02 mg rabbit anti-mouse platelet IgG at Time = 0 h. Groups of mice were then treated with Variant 117-HSA(C34S) (1 mg / kg, intravenous) at different times. Data are mean ± SEM, N=2 (control group) or N=3 (all other groups).DETAILED DESCRIPTION

[0119] The present application provides for a modified IdeS protein comprising lower immunogenicity, measured as reduced presentation of peptides on HLA-II or shielding of surface epitopes that are recognized by antibodies, and preserved or improved IdeS stability and IgG cleavage activity. To facilitate an understanding of the present disclosure, a number of terms and phrases are defined below.

[0120] The terms “a” and “an” as used herein mean “one or more” and include the plural unless the context is inappropriate.Attorney Docket No. : CYR-009WO

[0121] As used herein, the term “about” means a range of values including the specified value, which a person of ordinary skill in the art would consider reasonably similar to the specified value. In embodiments, “about” means within a standard deviation using measurements generally acceptable in the art. The term “about,” as used herein when referring to a measurable value such as an amount of mass, weight, time, volume, concentration, or percentage, is meant to encompass variations of in certain aspects ±20%, in certain aspects ±10%, in certain aspects ±9%, in certain aspects ±8%, in certain aspects ±7%, in certain aspects ±6%, in certain aspects ±5%, in certain aspects ±4%, in certain aspects ±3%, in certain aspects ±2%, in certain aspects ±1%, in certain aspects ±0.5%, and in certain aspects ±0.1% from the specified amount, as such variations are appropriate to perform the disclosed methods and / or employ the disclosed compositions. Accordingly, unless indicated to the contrary, the numerical parameters set forth in this specification and attached claims are approximations that can vary depending upon the desired properties sought to be obtained by the presently disclosed subject matter.

[0122] As used herein, the term “albumin binding domain” or “ABD” refers to a surface protein expressed by gram-positive bacteria that specifically binds to human serum albumin and includes both wild-type and naturally occurring or synthetic variants. Variants of albumin binding domains, e.g., having one or more amino acid residues modified compared to a wild-type albumin domain, are also disclosed herein and may also be referred to as albumin binding domains. Exemplary synthetic ABD sequences are provided in SEQ ID NO: 175 and SEQ ID NO: 176.

[0123] As used herein, the term “biantennary” refers to an N-linked glycan comprising the N-glycan core (Manal -6[Manal -3)]Man(P 1 -4)GlcNAc(P 1 -4)GlcNAc(P 1 -Asn-X-Ser / Thr) elongated with two GlcNAc residues linked to C-2 of the core mannose al -3 and the mannose al-6. This core structure can then be elongated or modified by various glycan structures.

[0124] As used herein, the term “conjugated” or “protein conjugate” refers to the attachment of another chemical group, molecule, moiety, or protein, typically to a protein. Attachment of the chemical group, molecule or protein can occur via covalent bonding, non- covalent bonding (i.e., hydrogen bonding, ionic interactions, Van der Waals interactions, and hydrophobic bond), use of a linker (e.g., peptide linker or chemical linker), fusion, or any other suitable method. Conjugation may be direct where, for example, one protein isAttorney Docket No. : CYR-009WO immediately connected to another protein, or it may be indirect, where one protein, for example, is connected via a linker to the other protein.

[0125] As used herein, the term “deletion” refers to the removal of one or more than one amino acid residue in the context of a reference amino acid sequence. A deletion can occur at the N-terminal or C-terminal position or anywhere between those terminal positions.Deletions may be interspersed either individually among residues in the reference sequence or in one or more contiguous groups within the reference sequence. When occurring at the N-terminal or C-terminal positions, a deletion of one or more than one contiguous amino acids can also be referred to as a “truncation.”

[0126] As used herein, the term “effective amount” refers to the amount of a drug (e.g., an IdeS variant, a polypeptide comprising an IdeS variant) sufficient to effect beneficial or desired results. An effective amount can be administered in one or more administrations, applications, or dosages and is not intended to be limited to a particular formulation or administration route.

[0127] As used herein, the term “functional fragment thereof’ refers to a portion of a protein or polypeptide that maintains the ability to perform a biological function of the whole protein or polypeptide. For example, a functional fragment of a polypeptide or protein of the present application maintains its ability to perform its catalytic activity.

[0128] As used herein, the term “heterologous” refers to a nucleic acid or polypeptide originating from a different genetic source or species.

[0129] As used herein, the term “human serum albumin” or “HSA” refers to a protein abundant in human blood plasma and where the wild-type mature form has the amino acid sequence: DAHKSEVAHRFKDLGEENFKALVLIAFAQYLQQCPFEDHVKLVNEVTEFAKTCVAD ESAENCDKSLHTLFGDKLCTVATLRETYGEMADCCAKQEPERNECFLQHKDDNPNL PRLVRPEVDVMCTAFHDNEETFL1<I<YLYEIARRHPYFYAPELLFFAI<RYI<AAFTECC QAADKAACLLPKLDELRDEGKASSAKQRLKCASLQKFGERAFKAWAVARLSQRFP KAEFAEVSKLVTDLTKVHTECCHGDLLECADDRADLAKYICENQDSISSKLKECCEK PLLEKSHCIAEVENDEMPADLPSLAADFVESKDVCKNYAEAKDVFLGMFLYEYARR HPDYSVVLLLRLAKTYETTLEKCCAAADPHECYAKVFDEFKPLVEEPQNLIKQNCEL FEQLGEYKFQNALLVRYTKKVPQVSTPTLVEVSRNLGKVGSKCCKHPEAKRMPCAEAttorney Docket No. : CYR-009WODYLSVVLNQLCVLHEKTPVSDRVTKCCTESLVNRRPCFSALEVDETYVPKEFNAETF TFHADICTLSEKERQIKKQTALVELVKHKPKATKEQLKAVMDDFAAFVEKCCKADD KETCFAEEGKKLVAASQAALGL (SEQ ID NO: 188).

[0130] In addition to the wild-type form, “human serum albumin” also refers to naturally occurring allelic variants. Known allelic variants are described in, e.g., Cardi et cd.. (2022), Intr. J. Mol. Set., 23(3): 1159, and at https: / / albumin.org / genetic-variants-of-human-serum- albumin / . When used in the context of IdeS polypeptide conjugates disclosed herein, “human serum albumin” or “human serum albumin moiety” may refer to both naturally occurring human serum albumin and variant human serum albumin as disclosed herein, depending on the context.

[0131] As used herein, the term “IdeS protein” or “IdeS peptide” refers to Immunoglobulin G-degrading enzyme of Streptococcus pyogenes which is an extracellular cysteine proteinase produced by S. pyogenes that cleaves human IgG in the hinge region with a high degree of specificity, producing an (Fab’)2 and an Fab fragment. An exemplary wildtype IdeS protein is provided in SEQ ID NO: 1 (with the signal sequence) and in SEQ ID NO:2 (mature form without the signal sequence). Naturally occurring variants of wild-type IdeS are also known. Further, variants from wild-type that are not naturally occurring (z.e., created by genetic engineering) are also disclosed herein. Accordingly, “IdeS protein” or “IdeS peptide” may refer to wild-type IdeS, to both wild-type IdeS and IdeS variants, or IdeS variants alone depending on the context.

[0132] As used herein, the term “IdeS polypeptide conjugate” refers to a chimeric protein including a IdeS protein moiety, such as wild-type IdeS or an IdeS variant, and a second protein moiety that is not an IdeS protein. The second moiety may be selected from (i) human serum albumin, (ii) variant human serum albumin, or (iii) an albumin binding domain. The first and second moieties may be connected via a linker sequence, such as one of the linker sequences provided in SEQ ID NOS: 162-171 as disclosed herein.

[0133] As used herein, the term “modified IdeS protein,” “IdeS protein variant,” “variant IdeS protein,” or “IdeS variant” refers to an IdeS protein that has one or more amino acid modifications (e.g., one or more substitutions, insertions, or deletions) as compared to the mature form of a wild-type IdeS protein, e.g., compared to residues 30-339 of SEQ ID NO: 1 (equivalent to SEQ ID NO:2) and retains activity associated with a wild-type IdeS protein,Attorney Docket No. : CYR-009WO e.g., IgG cleavage activity. For example, an IdeS variant may retain at least 5% of the IgG cleavage activity of a wild-type IdeS protein, e.g., the IdeS protein of SEQ ID NO:2.

[0134] As used herein, the terms “mutant” and “variant” are used interchangeably and refer to a protein, or enzyme having one or more than one mutation and / or deletion in the context of a reference sequence (e.g., a wild-type sequence).

[0135] As used herein, the terms “mutation,” “modification,” and “substitution” are used interchangeably and refer to the alteration of an amino acid, in the context of a reference amino acid sequence, to another amino acid. An alteration of an amino acid in a reference amino acid sequence can occur at the N-terminal or C-terminal position or anywhere between those terminal positions. Modifications may be interspersed either individually among residues in the reference sequence or in one or more contiguous groups within the reference sequence.

[0136] A substitution can be, but need not necessarily be, a conservative substitution. Twenty amino acids are commonly found in proteins. Those amino acids can be grouped into nine classes or groups based on the chemical properties of their side chains. Substitution of one amino acid residue for another within the same class or group is referred to herein as a “conservative” substitution. Conservative amino acid substitutions can frequently be made in a protein without significantly altering the conformation or function of the protein.Substitution of one amino acid residue for another from a different class or group is referred to herein as a “non-conservative” substitution. In contrast, non-conservative amino acid substitutions tend to modify conformation and function of a protein.Example of Amino Acid ClassificationAttorney Docket No. : CYR-009WO

[0137] In certain aspects, a conservative amino acid substitution includes substituting any of: glycine (G), alanine (A), isoleucine (I), valine (V), and leucine (L) for any other of these aliphatic amino acids; serine (S) for threonine (T) and vice versa; aspartic acid (D) for glutamic acid (E) and vice versa; glutamine (Q) for asparagine (N) and vice versa; lysine (K) for arginine (R) and vice versa; phenylalanine (F), tyrosine (Y) and tryptophan (W) for any other of these aromatic amino acids; and methionine (M) for cysteine (C) and vice versa. Other substitutions can also be considered conservative, depending on the environment of the particular amino acid and its role in the three-dimensional structure of the protein. For example, glycine (G) and alanine (A) can frequently be interchangeable, as can alanine (A) and valine (V). Methionine (M), which is relatively hydrophobic, can frequently be interchanged with leucine and isoleucine, and sometimes with valine. Lysine (K) and arginine (R) are frequently interchangeable in locations in which the significant feature of the amino acid residue is its charge and the differing pKs of these two amino acid residues are not significant. Still other changes can be considered “conservative” in particular environments (see, e.g., BIOCHEMISTRY at pp. 13-15, 2nd ed. Lubert Stryer ed. (Stanford University); Henikoff et al., Proc. Nat'l Acad. Sci. USA (1992) 89: 10915-10919; Lei et al., J. Biol. Chem. (1995) 270(20): 11882-11886).

[0138] As used herein, the term “neuraminic acid” refers to a 9-carbon monosaccharide, a derivative of a ketonanose.

[0139] As used herein, the term “N-glycan” refers to a core structure generally comprising two N-Acetyl-D-glucosamine (GlcNAc) and three mannose molecules. N-glycans can be added to an asparagine (Asn) side chain via N-linked glycosylation initiated by oligosaccharyltransferase complex in the endoplasmic reticulum membrane. N-glycans are attached to Asn located in a sequence of Asn-X-Serine / Threonine, whereby X can be any amino acid apart from proline. N-glycans may be expressed as: (Manal-6[Manal- 3)]Man(pi-4)GlcNAc(pi-4)GlcNAc(pi-Asn-X-Ser / Thr). The N-glycan core structure can be expanded through galactosylation, further GlcNAclyation, sialylation, fucosylation, or combinations thereof.

[0140] As used herein, the term “N-glycosylation” refers to the attachment of a sugar molecule oligosaccharide known as glycan to a nitrogen atom of an amino acid in a protein.Attorney Docket No. : CYR-009WO

[0141] As used herein, “nucleic acid” or “oligonucleotide” or “polynucleotide” or grammatical equivalents used herein means at least two nucleotides covalently linked together. The term “nucleic acid” includes single-, double-, or multiple-stranded DNA, RNA and analogs (derivatives) thereof. Oligonucleotides can be from about 5, 6, 7, 8, 9, 10, 12, 15, 25, 30, 40, 45 or more nucleotides in length, up to about 100 nucleotides in length. Nucleic acids and polynucleotides are polymers of any length, including longer lengths, e.g, 200, 300, 440, 1000, 2000, 3000, 4400, 7000, 10,000, etc. nucleotides in length.

[0142] As used herein, the term “O-glycosylation” refers to the attachment of a sugar molecule oligosaccharide (i.e., a glycan) to an oxygen atom of an amino acid in a protein.

[0143] As used herein, the term “pharmaceutical formulation” refers to the combination of an active agent (e.g, an IdeS variant) with a carrier, inert or active, making the composition especially suitable for diagnostic or therapeutic use in vivo or ex vivo.

[0144] As used herein, the term “pharmaceutically acceptable carrier” refers to any of the standard pharmaceutical carriers, such as a phosphate buffered saline solution, water, emulsions (e.g., such as an oil / water or water / oil emulsions), and various types of wetting agents. The compositions also can include stabilizers and preservatives. For examples of carriers, stabilizers and adjuvants, see e.g., Martin, Remington's Pharmaceutical Sciences, 15th Ed., Mack Publ. Co., Easton, PA

[1975] ,

[0145] The terms “protein” or “polypeptide” are used interchangeably herein and refer to a polymer of repeating structural units connected by a peptide bond. Typically, the repeating structural units of the peptide are amino acids including naturally occurring amino acids, non- naturally occurring amino acids, analogues of amino acids or any combination of these. In certain aspects, proteins or polypeptides may be post-translationally modified (e.g. glycosylated, phosphorylated, lapidated, acetylated, or conjugation with a labeling component).

[0146] The term “sequence identity” means the proportion of amino acid matches between two amino acid sequences. When sequence homology is expressed as a percentage, e.g., 85%, the percentage denotes the fraction of matches over the length of sequence that is compared to some other sequence. Gaps (in either of the two sequences) are permitted to maximize matching; for example, wherein gap lengths of 5 amino acids or less, optionally 3 amino acids or less, are usually used.Attorney Docket No. : CYR-009WO

[0147] Percent sequence identity can be any integer from 60% to 100%. Exemplary embodiments include at least: 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%, as compared to a reference sequence using the programs described herein; preferably BLAST using standard parameters, as described below. One of skill will recognize that these values can be appropriately adjusted to determine corresponding identity of proteins encoded by two nucleotide sequences by taking into account codon degeneracy, amino acid similarity, reading frame positioning and the like.

[0148] For sequence comparison, typically one sequence acts as a reference sequence to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are entered into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. Default program parameters can be used, or alternative parameters can be designated. The sequence comparison algorithm then calculates the percent sequence identities for the test sequences relative to the reference sequence, based on the program parameters.

[0149] Algorithms that are suitable for determining percent sequence identity and sequence similarity are the BLAST and BLAST 2.0 algorithms, which are described in Altschul et al. (1990) J. Mol. Biol. 215: 403-410 and Altschul et al. (1977) Nucleic Acids Res. 25: 3389-3402, respectively. Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information (NCBI) web site. The algorithm involves first identifying high scoring sequence pairs (HSPs) by identifying short words of length W in the query sequence, which either match or satisfy some positive-valued threshold score T when aligned with a word of the same length in a database sequence. T is referred to as the neighborhood word score threshold (Altschul et al, supra). These initial neighborhood word hits act as seeds for initiating searches to find longer HSPs containing them. The word hits are then extended in both directions along each sequence for as far as the cumulative alignment score can be increased. Cumulative scores are calculated using, for nucleotide sequences, the parameters M (reward score for a pair of matching residues; always >0) and N (penalty score for mismatching residues; always <0). For amino acid sequences, a scoring matrix is used to calculate the cumulative score. Extension of the word hits in each direction are halted when: the cumulative alignment score falls off by the quantity X from its maximum achieved value; the cumulative score goes to zero or below, due to the accumulation of one or more negative-scoring residue alignments; or the end of eitherAttorney Docket No. : CYR-009WO sequence is reached. The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses as defaults a word size (W) of 28, an expectation (E) of 10, M=l, N=-2, and a comparison of both strands. For amino acid sequences, the BLASTP program uses as defaults a word size (W) of 3, an expectation (E) of 10, and the BLOSUM62 scoring matrix (see Henikoff & Henikoff, Proc. Natl. Acad. Sci. USA 89: 10915 (1989)).

[0150] The BLAST algorithm also performs a statistical analysis of the similarity between two sequences (see, e.g., Karlin & Altschul, Proc. Nat'L Acad. Sci. USA 90:5873-5787 (1993)). One measure of similarity provided by the BLAST algorithm is the smallest sum probability (P(N)), which provides an indication of the probability by which a match between two nucleotide or amino acid sequences would occur by chance. For example, a nucleic acid is considered similar to a reference sequence if the smallest sum probability in a comparison of the test nucleic acid to the reference nucleic acid is less than about 0.01, more preferably less than about 10'5, and most preferably less than about IO'20.

[0151] As used herein, the term “sialylation” is the enzymatic addition of a neuraminic acid residue.

[0152] As used herein, the terms “subject” and “patient” refer to an organism to be treated by the methods and compositions described herein. Such organisms preferably include, but are not limited to, mammals (e.g., murines, simians, equines, bovines, porcines, canines, felines, and the like), and more preferably include humans.

[0153] As used herein, the term “tetraantennary” refers to an N-linked glycan comprising two additional GlcNAc residues that are added to either the C-4 of the core mannose al -3 or the C-6 of the core mannose al -6 of the biantennary core structure. This core structure can then be elongated or modified by various glycan structures.

[0154] As used herein, the terms “treat”, “treating,” or “treatment” includes inhibition, prevention, or postponement of the development of the symptoms associated with a disease or disorder and / or a reduction in the severity, frequency and / or duration of the symptoms of such a disorder. The terms further include ameliorating existing uncontrolled or unwanted symptoms, preventing additional symptoms, and ameliorating or preventing the underlying causes of such symptoms.Attorney Docket No. : CYR-009WO

[0155] As used herein, the term “triantennary” refers to an N-linked glycan comprising an additional GlcNAc residue is added to either the C-4 of the core mannose al-3 or the C-6 of the core mannose al-6 of the biantennary core structure. This structure can then be elongated or modified by various glycan structures.

[0156] As used herein, the term “variant human serum albumin” or “variant HSA” refers to a human serum albumin that has one or more amino acid modifications (e.g., one or more substitutions, insertions, or deletions) as compared to the mature form of wild-type HSA (SEQ ID NO: 188), and is synthetic, that is, not a naturally occurring variant. Variant human serum albumin molecules typically have at least 90% sequence identity with the amino acid sequence of the mature form of wild-type human serum albumin of SEQ ID NO: 188. For example, in some embodiments variant human serum albumin molecules have at least 99% sequence identity with the amino acid sequence of the mature form of wild-type human serum albumin of SEQ ID NO: 188. Variant HSA may include a mutation to remove a free cysteine, e.g., a cysteine corresponding to position 34 of SEQ ID NO: 188, such as by substitution with another amino acid, such as serine, alanine, or valine, e.g., serine.

[0157] As used herein, the term “vector” refers to a nucleic acid molecule as introduced into a host cell, thereby producing a transformed host cell. A vector may include nucleic acid sequences that permit it to replicate in a host cell, such as an origin of replication. A vector may also include one or more selectable marker genes and other genetic elements known in the art. In certain aspects, the vector is a virus vector, such as a lentivirus vector.I. Modified IdeS proteins

[0158] Described herein, in certain embodiments, are modified IdeS proteins comprising one or more modifications as compared to wild-type IdeS sequence.

[0159] Wild-type IdeS with signal peptide (signal peptide underlined) MRKRCYSTSAAVLAAVTLFVLSVDRGVIADSFSANQEIRYSEVTPYHVTSVWTKGV TPPANFTQGEDVFHAPYVANQGWYDITKTFNGKDDLLCGAATAGNMLHWWFDQN KDQIKRYLEEHPEKQKINFNGEQMFDVKEAIDTKNHQLDSKLFEYFKEKAFPYLSTK HLGVFPDHVIDMFINGYRLSLTNHGPTPVKEGSKDPRGGIFDAVFTRGDQSKLLTSRH DFKEKNLKEISDLIKKELTEGKALGLSHTYANVRINHVINLWGADFDSNGNLKAIYV TDSDSNASIGMKKYFVGVNSAGKVAISAKEIKEDNIGAQVLGLFTLSTGQDSWNQTN (SEQ ID NO: 1)Attorney Docket No. : CYR-009WO

[0160] As used herein, references to amino acid residue positions are relative to the position of the residue in SEQ ID NO: 1 (i.e. full-length wild-type IdeS sequence including signal peptide) unless stated otherwise.

[0161] Wild-type IdeS (mature polypeptide) DSFSANQEIRYSEVTPYHVTSVWTKGVTPPANFTQGEDVFHAPYVANQGWYDITKT FNGKDDLLCGAATAGNMLHWWFDQNKDQIKRYLEEHPEKQKINFNGEQMFDVKE AIDTKNHQLDSKLFEYFKEKAFPYLSTKHLGVFPDHVIDMFINGYRLSLTNHGPTPVK EGSKDPRGGIFDAVFTRGDQSKLLTSRHDFKEKNLKEISDLIKKELTEGKALGLSHTY ANVRINHVINLWGADFDSNGNLKAIYVTDSDSNASIGMKKYFVGVNSAGKVAISAK EIKEDNIGAQVLGLFTLSTGQDSWNQTN (SEQ ID NO:2; corresponds to residues SO- 339 of SEQ ID NO: 1)7. IdeS Variants

[0162] Described herein are modified IdeS proteins. For example, the IdeS protein is modified compared to the mature form of a wild-type IdeS protein, e.g., a wild-type IdeS protein having the amino acid sequence of SEQ ID NO:2. In some embodiments, the IdeS protein is modified to have reduced presentation on HLA-II. In some embodiments, the IdeS protein is modified (e.g., by introduction of one or more glycosylation sites, or by fusion to serum albumin) to have reduced recognition by anti-IdeS antibodies. In some embodiments, the IdeS protein is modified (e.g., by fusion to serum albumin) to prolong half-life of the modified IdeS protein and / or IgG depletion. In some embodiments, the IdeS protein is modified in multiple ways.

[0163] In some embodiments, the present disclosure provides an IdeS variant comprising one or more modifications as compared to wild-type IdeS sequence, and having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, a sequence identity with SEQ ID NO: 1.

[0164] In some embodiments, the present disclosure provides an IdeS variant comprising one or more modifications as compared to wild-type IdeS sequence, and having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%,Attorney Docket No. : CYR-009WO at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.3%, or at least 99.6% sequence identity with SEQ ID NO:2.

[0165] In some embodiments, the present disclosure provides an IdeS variant comprising at least two modifications as compared to wild-type IdeS sequence, and having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, sequence identity with the amino acid sequence of SEQ ID NO: 1. In some embodiments, the present disclosure provides an IdeS variant having at least 87% sequence identity with SEQ ID NO: 1. In some embodiments, the present disclosure provides an IdeS variant having at least 95% sequence identity with SEQ ID NO: 1 and having one or more modifications. In some embodiments, the present disclosure provides an IdeS variant having at least 95% sequence identity with SEQ ID NO: 1 and having at least two modifications. In some embodiments, the IdeS variant has 10 or more modifications. In some embodiments, the IdeS variant has 12 or more modifications. In some embodiments, the IdeS variant is a mature form lacking the signal sequence.

[0166] In some embodiments, the present disclosure provides an IdeS variant comprising at least two modifications as compared to wild-type IdeS sequence, and having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.3%, or at least 99.6% sequence identity with the amino acid sequence of SEQ ID NO:2. In some embodiments, the present disclosure provides an IdeS variant having at least 96% sequence identity with SEQ ID NO:2 and having two or more modifications. In some embodiments, the present disclosure provides an IdeS variant having at least 96% sequence identity with SEQ ID NO:2 and having at least two modifications. In some embodiments, the IdeS variant has 10 or more modifications. In some embodiments, the IdeS variant has 12 or more modifications. In some embodiments, the IdeS variant is a mature form lacking the signal sequence.Attorney Docket No. : CYR-009WO

[0167] Described herein, in certain embodiments, are IdeS variant proteins comprising a modification at a position corresponding to position 189 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein comprises a modification of a threonine to a glycine at a position corresponding to position 189 of SEQ ID NO: 1.

[0168] In some embodiments, the IdeS variant proteins comprise a modification at a position corresponding to position 132 of SEQ ID NO: 1. In some embodiments, the modification is a modification of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1.

[0169] In some embodiments, the IdeS variant proteins comprise modifications at positions corresponding to positions 132 and 189 of SEQ ID NO: 1. In some embodiments, the modification is a modification of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1 and a threonine to a glycine at a position corresponding to position 189 of SEQ ID NO: 1.

[0170] In some embodiments, the IdeS variant protein comprises at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90% at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% to SEQ ID NO: 1. In some embodiments, the IdeS variant protein c an amino acid sequence having at least 87% sequence identity to SEQ ID NO: 1. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 1. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 1. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 99% sequence identity to SEQ ID NO: 1. In some embodiments, the IdeS variant is a mature form lacking the signal sequence.

[0171] In some embodiments, the IdeS variant protein comprises at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.3%, or at least 99.6% sequence identity with the amino acid sequence of SEQ ID NO:2. In some embodiments, the IdeS variant protein has at least 85% sequence identity with SEQ ID NO:2. In some embodiments, the IdeS variant protein has at least 87%Attorney Docket No. : CYR-009WO sequence identity with SEQ ID NO:2. In some embodiments, the IdeS variant protein has at least 90% sequence identity with SEQ ID NO:2. In some embodiments, the IdeS variant protein has at least 96% sequence identity with SEQ ID NO:2. In some embodiments, the IdeS variant protein has at least 98% sequence identity with SEQ ID NO:2.

[0172] In some embodiments, the IdeS variant protein further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 113,115, 116, 127, 128, 129, 130, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187, 188, 195,197, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258, 273, 274, 277, 278,288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.

[0173] In some embodiments, the IdeS variant protein further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111,113, 115, 116, 127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187,188, 189, 195, 197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258,273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316,318, 322, and 330. For example, the IdeS variant comprises modifications corresponding to two or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to three or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to four or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to five or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to six or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to seven or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to eight or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to nine or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to ten or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to eleven or more of the aforementioned positions. For example, the IdeS variant comprises modifications corresponding to twelve or more of the aforementioned positions.Attorney Docket No. : CYR-009WO

[0174] In some embodiments, the modification corresponding to position 31 of SEQ ID NO: l is a substitution of serine to aspartate or asparagine. For example, in some embodiments, the substitution is aspartate. For example, in other embodiments, the substitution is asparagine.

[0175] In some embodiments, the modification corresponding to position 32 of SEQ ID NO: 1 is a substitution of phenylalanine to lysine.

[0176] In some embodiments, the modification corresponding to position 33 of SEQ ID NO: 1 is a substitution of serine to glutamate.

[0177] In some embodiments, the modification corresponding to position 38 of SEQ ID NO: l is a substitution of isoleucine to valine.

[0178] In some embodiments, the modification corresponding to position 39 of SEQ ID NO: l is a substitution of arginine to asparagine or threonine.

[0179] In some embodiments, the modification corresponding to position 43 of SEQ ID NO: l is a substitution of valine to glutamate.

[0180] In some embodiments, the modification corresponding to position 44 of SEQ ID NO: l is a substitution of threonine to glutamate.

[0181] In some embodiments, the modification corresponding to position 45 of SEQ ID NO: 1 is a substitution of proline to glutamate.

[0182] In some embodiments, the modification corresponding to position 47 of SEQ ID NO: 1 is a substitution of histidine to lysine.

[0183] In some embodiments, the modification corresponding to position 54 of SEQ ID NO: 1 is a substitution of lysine to aspartate.

[0184] In some embodiments, the modification corresponding to position 57 of SEQ ID NO: l is a substitution of threonine to lysine or glutamine. For example, in some embodiments, the substitution is lysine. For example, in other embodiments, the substitution is glutamine.Attorney Docket No. : CYR-009WO

[0185] In some embodiments, the modification corresponding to position 60 of SEQ ID NO: l is a substitution of alanine to aspartate or glutamine. For example, in some embodiments, the substitution is aspartate. For example, in other embodiments, the substitution is glutamine.

[0186] In some embodiments, the modification corresponding to position 68 of SEQ ID NO: l is a substitution of valine to threonine.

[0187] In some embodiments, the modification corresponding to position 74 of SEQ ID NO: l is a substitution of valine to lysine.

[0188] In some embodiments, the modification corresponding to position 75 of SEQ ID NO: l is a substitution of alanine to asparagine or proline. For example, in some embodiments, the substitution is asparagine. For example, in other embodiments, the substitution is proline.

[0189] In some embodiments, the modification corresponding to position 75 of SEQ ID NO: l is a substitution of alanine to proline.

[0190] In some embodiments, the modification corresponding to position 77 of SEQ ID NO: l is a substitution of glutamine to glycine.

[0191] In some embodiments, the modification corresponding to position 82 of SEQ ID NO: l is a substitution of isoleucine to methionine.

[0192] In some embodiments, the modification corresponding to position 85 of SEQ ID NO: l is a substitution of threonine to glutamine.

[0193] In some embodiments, the modification corresponding to position 111 of SEQ ID NO:1 is a substitution of lysine to asparagine.

[0194] In some embodiments, the modification corresponding to position 113 of SEQ ID NO: l is a substitution of glutamine to aspartate.

[0195] In some embodiments, the modification corresponding to position 115 of SEQ ID NO:1 is a substitution of lysine to histidine.Attorney Docket No. : CYR-009WO

[0196] In some embodiments, the modification corresponding to position 116 of SEQ ID NO: 1 is a substitution of arginine to aspartate.

[0197] In some embodiments, the modification corresponding to position 127 of SEQ ID NO: l is a substitution of isoleucine to methionine.

[0198] In some embodiments, the modification corresponding to position 128 of SEQ ID NO: 1 is a substitution of asparagine to glycine.

[0199] In some embodiments, the modification corresponding to position 129 of SEQ ID NO: 1 is a substitution of phenylalanine to glutamate.

[0200] In some embodiments, the modification corresponding to position 130 of SEQ ID NO: l is a substitution of asparagine to glutamate

[0201] In some embodiments, the modification corresponding to position 132 of SEQ ID NO: 1 is a substitution of glutamate for glutamine.

[0202] In some embodiments, the modification corresponding to position 133 of SEQ ID NO: 1 is a substitution of glutamine to aspartate.

[0203] In some embodiments, the modification corresponding to position 148 of SEQ ID NO: 1 is a substitution of leucine to asparagine or arginine. For example, in some embodiments, the substitution is asparagine. For example, in other embodiments, the substitution is arginine.

[0204] In some embodiments, the modification corresponding to position 153 of SEQ ID NO: l is a substitution of phenylalanine to methionine or tyrosine. For example, in some embodiments, the substitution is methionine. For example, in other embodiments, the substitution is tyrosine.

[0205] In some embodiments, the modification corresponding to position 154 of SEQ ID NO: l is a substitution of glutamate to aspartate.

[0206] In some embodiments, the modification corresponding to position 159 of SEQ ID NO: 1 is a substitution of lysine to asparagine.Attorney Docket No. : CYR-009WO

[0207] In some embodiments, the modification corresponding to position 166 of SEQ ID NO: l is a substitution of threonine to arginine or a substitution of threonine to glycine. For example, in some embodiments, the substitution is arginine. For example, in other embodiments, the substitution is glycine.

[0208] In some embodiments, the modification corresponding to position 167 of SEQ ID NO: 1 is a substitution of lysine to proline.

[0209] In some embodiments, the modification corresponding to position 168 of SEQ ID NO: 1 is a substitution of histidine to aspartate or proline. For example, in some embodiments, the substitution is aspartate. For example, in other embodiments, the substitution is proline.

[0210] In some embodiments, the modification corresponding to position 175 of SEQ ID NO: 1 is a substitution of histidine to glutamine.

[0211] In some embodiments, the modification corresponding to position 187 of SEQ ID NO: l is a substitution of serine to aspartate or a substitution of serine to glutamate.

[0212] In some embodiments, the modification corresponding to position 188 of SEQ ID NO: 1 is a substitution of leucine to glycine or methionine.

[0213] In some embodiments, the modification corresponding to position 189 of SEQ ID NO: l is a substitution of threonine for glycine.

[0214] In some embodiments, the modification corresponding to position 195 of SEQ ID NO: 1 is a substitution of proline to aspartate.

[0215] In some embodiments, the modification corresponding to position 197 of SEQ ID NO: 1 is a substitution of lysine to glutamate.

[0216] In some embodiments, the modification corresponding to position 198 of SEQ ID NO: 1 is a substitution of glutamate to asparagine.

[0217] In some embodiments, the modification corresponding to position 210 of SEQ ID NO: l is a substitution of alanine to glycine.Attorney Docket No. : CYR-009WO

[0218] In some embodiments, the modification corresponding to position 213 of SEQ ID NO: l is a substitution of threonine to glutamate.

[0219] In some embodiments, the modification corresponding to position 218 of SEQ ID NO: 1 is a substitution of serine to aspartate.

[0220] In some embodiments, the modification corresponding to position 219 of SEQ ID NO: 1 is a substitution of lysine to glycine.

[0221] In some embodiments, the modification corresponding to position 220 of SEQ ID NO: 1 is a substitution of leucine to glutamine.

[0222] In some embodiments, the modification corresponding to position 228 of SEQ ID NO: 1 is a substitution of lysine to glutamine.

[0223] In some embodiments, the modification corresponding to position 233 of SEQ ID NO: 1 is a substitution of lysine to serine.

[0224] In some embodiments, the modification corresponding to position 236 of SEQ ID NO: 1 is a substitution of serine to cysteine.

[0225] In some embodiments, the modification corresponding to position 241 of SEQ ID NO: 1 is a substitution of lysine to aspartate.

[0226] In some embodiments, the modification corresponding to position 244 of SEQ ID NO: l is a substitution of threonine to lysine.

[0227] In some embodiments, the modification corresponding to position 245 of SEQ ID NO: l is a substitution of glutamate to aspartate.

[0228] In some embodiments, the modification corresponding to position 247 of SEQ ID NO: 1 is a substitution of lysine to asparagine.

[0229] In some embodiments, the modification corresponding to position 258 of SEQ ID NO: l is a substitution of valine to aspartate.

[0230] In some embodiments, the modification corresponding to position 273 of SEQ ID NO: 1 is a substitution of serine to aspartate.Attorney Docket No. : CYR-009WO

[0231] In some embodiments, the modification corresponding to position 274 of SEQ ID NO: 1 is a substitution of asparagine to glutamate.

[0232] In some embodiments, the modification corresponding to position 277 of SEQ ID NO: 1 is a substitution of leucine to cysteine.

[0233] In some embodiments, the modification corresponding to position 278 of SEQ ID NO: 1 is a substitution of lysine to aspartate or threonine. For example, in some embodiments, the substitution is aspartate. For example, in other embodiments, the substitution is threonine.

[0234] In some embodiments, the modification corresponding to position 288 of SEQ ID NO: l is a substitution of asparagine to aspartate, glycine, or glutamine. For example, in some embodiments, the substitution is aspartate. For example, in other embodiments, the substitution is glycine. For example, in yet other embodiments, the substitution is glutamine.

[0235] In some embodiments, the modification corresponding to position 289 of SEQ ID NO: l is a substitution of alanine to proline.

[0236] In some embodiments, the modification corresponding to position 297 of SEQ ID NO: 1 is a substitution of phenylalanine to aspartate.

[0237] In some embodiments, the modification corresponding to position 299 of SEQ ID NO: 1 is a substitution of glycine to aspartate.

[0238] In some embodiments, the modification corresponding to position 300 of SEQ ID NO: l is a substitution of valine to glutamate.

[0239] In some embodiments, the modification corresponding to position 302 of SEQ ID NO: 1 is a substitution of serine to aspartate, a substitution of serine to lysine, or a substitution of serine to glutamate. For example, in some embodiments, the substitution is aspartate. For example, in other embodiments, the substitution is lysine. For example, in yet other embodiments, the substitution is glutamate.

[0240] In some embodiments, the modification corresponding to position 303 of SEQ ID NO: 1 is a substitution of alanine to aspartate, a substitution of alanine to asparagine, or a substitution of alanine to glutamine. For example, in some embodiments, the substitution isAttorney Docket No. : CYR-009WO aspartate. For example, in other embodiments, the substitution is asparagine. For example, in yet other embodiments, the substitution is glutamine.

[0241] In some embodiments, the modification corresponding to position 306 of SEQ ID NO: l is a substitution of valine to threonine.

[0242] In some embodiments, the modification corresponding to position 307 of SEQ ID NO: l is a substitution of alanine to glycine or asparagine.

[0243] In some embodiments, the modification corresponding to position 308 of SEQ ID NO: l is a substitution of isoleucine to leucine.

[0244] In some embodiments, the modification corresponding to position 313 of SEQ ID NO: l is a substitution of isoleucine to proline.

[0245] In some embodiments, the modification corresponding to position 314 of SEQ ID NO: 1 is a substitution of lysine to glycine.

[0246] In some embodiments, the modification corresponding to position 315 of SEQ ID NO: l is a substitution of glutamate to proline.

[0247] In some embodiments, the modification corresponding to position 316 of SEQ ID NO: l is a substitution of aspartate to proline.

[0248] In some embodiments, the modification corresponding to position 318 of SEQ ID NO: l is a substitution of isoleucine to lysine, a substitution of isoleucine to aspartate, or a substitution of isoleucine to glycine. For example, in some embodiments, the substitution is lysine. For example, in other embodiments, the substitution is aspartate. For example, in yet other embodiments, the substitution is glycine.

[0249] In some embodiments, the modification corresponding to position 322 of SEQ ID NO: l is a substitution of valine to threonine.

[0250] In some embodiments, the modification corresponding to position 330 of SEQ ID NO: l is a substitution of threonine to aspartate.

[0251] In some embodiments, the IdeS variant protein comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ IDAttorney Docket No. : CYR-009WONO: 1 selected from 68, 75, 166, 187, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises two or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises three or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises four or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises five or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises six or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises modifications at positions corresponding to positions 68, 75, 166, 187, 213, 302, and 303 of SEQ ID NO: 1.

[0252] In some embodiments, the IdeS variant protein comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 68, 75, 132, 166, 187, 189, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises two or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 132, 166, 187, 189, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises three or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 132, 166, 187, 189, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises four or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 132, 166, 187, 189, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises five or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 132, 166, 187, 189, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises six or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 132, 166, 187, 189, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises seven or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 132, 166, 187, 189, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises eight or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 132, 166, 187, 189, 213,Attorney Docket No. : CYR-009WO302, and 303. In some embodiments, the IdeS variant protein further comprises modifications at positions corresponding to positions 68, 75, 132, 166, 187, 189, 213, 302, and 303 of SEQ ID NO: 1.

[0253] In some embodiments, the IdeS variant protein comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises two or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises three or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises four or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises five or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises six or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises seven or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises eight or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises nine or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein further comprises modifications at positions corresponding to positions 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303 of SEQ ID NO: 1.

[0254] For example, in some embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75Attorney Docket No. : CYR-009WO of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0255] For example, in some embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0256] In other embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0257] In other embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to positionAttorney Docket No. : CYR-009WO302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0258] For example, in some embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glycine at a position corresponding to position 189 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0259] For example, in some embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glycine at a position corresponding to position 189 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0260] For example, in some embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at aAttorney Docket No. : CYR-009WO position corresponding to position 187 of SEQ ID NO: 1, a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0261] For example, in some embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0262] For example, in some embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.Attorney Docket No. : CYR-009WO

[0263] For example, in some embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from: 68, 75, 166, 187, 213, 302, 303, or 306 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein further comprises two or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from: 68, 75, 166, 187, 213, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises three or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from: 68, 75, 166, 187, 213, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises four or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from: 68, 75, 166, 187, 213, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises five or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from: 68, 75, 166, 187, 213, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises six or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from: 68, 75, 166, 187, 213, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises seven or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from: 68, 75, 166, 187, 213, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises modifications at positions corresponding to positions 68, 75, 166, 187, 213, 302, 303, and 306 of SEQ ID NO: 1.

[0264] For example, in some embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 ofAttorney Docket No. : CYR-009WOSEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1, and a substitution of valine to threonine at a position corresponding to position 306 of SEQ ID NO: 1.

[0265] For example, in some embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1, and a substitution of valine to threonine at a position corresponding to position 306 of SEQ ID NO: 1.

[0266] In some embodiments, the IdeS variant protein further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, 306, or 318 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein further comprises two or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, 306, or 318 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein further comprises three or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, 306, or 318 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein further comprises four or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, 306, or 318 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein further comprises five or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, 306, or 318 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein further comprises six or more modifications atAttorney Docket No. : CYR-009WO positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, 306, or 318 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein further comprises seven or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 302, 306, or 318 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein comprises modifications at positions corresponding to positions 68, 75, 166, 187, 213, 302, 306 and 318 of SEQ ID NO: 1.

[0267] For example, in some embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, a substitution of valine to threonine at a position corresponding to position 306 of SEQ ID NO: 1, and a substitution of isoleucine to lysine at a position corresponding to position 318 of SEQ ID NO: 1.

[0268] For example, in some embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, a substitution of valine to threonine at a position corresponding to position 306 of SEQ ID NO: 1, and a substitution of isoleucine to lysine at a position corresponding to position 318 of SEQ ID NO: 1.

[0269] In some embodiments, the IdeS variant protein further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises two or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises three orAttorney Docket No. : CYR-009WO more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises four or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises five or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises six or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises seven or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises eight or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, 302, 303, or 306. In some embodiments, the IdeS variant protein further comprises nine or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, 302, 303, or 306. In some embodiments, the IdeS variant protein comprises modifications at positions corresponding to positions 68, 75, 166, 187, 213, 236, 277, 302, 303, and 306 of SEQ ID NO: 1.

[0270] For example, in some embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to cysteine at a position corresponding to position 236 of SEQ ID NO: 1, a substitution of leucine to cysteine at a position corresponding to position 277 of SEQ ID NO: 1, a substitution of serine at to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1, and a substitution of valine to threonine at a position corresponding to position 306 of SEQ ID NO: 1.Attorney Docket No. : CYR-009WO

[0271] For example, in some embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to cysteine at a position corresponding to position 236 of SEQ ID NO: 1, a substitution of leucine to cysteine at a position corresponding to position 277 of SEQ ID NO: 1, a substitution of serine at to glutamate at a position corresponding to position 302 of SEQ ID NO: 1, a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1, and a substitution of valine to threonine at a position corresponding to position 306 of SEQ ID NO: 1.

[0272] In some embodiments, the IdeS variant protein further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, or 308. In some embodiments, the IdeS variant protein further comprises two or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, or 308. In some embodiments, the IdeS variant protein further comprises three or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, or 308. In some embodiments, the IdeS variant protein further comprises four or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, or 308. In some embodiments, the IdeS variant protein further comprises five or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, or 308. In some embodiments, the IdeS variant protein further comprises six or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, or 308. In some embodiments, the IdeS variant protein further comprises seven or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 166, 187, 213, 236, 277, or 308 of SEQ ID NO: 1. In some embodiments, the IdeS variant protein further comprises modifications at positions corresponding to positions 68, 75, 166, 187, 213, 236, 277, or 308 of SEQ ID NO: 1.Attorney Docket No. : CYR-009WO

[0273] For example, in some embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to cysteine at a position corresponding to position 236 of SEQ ID NO: 1, a substitution of leucine to cysteine at a position corresponding to position 277 of SEQ ID NO: 1, and a substitution of isoleucine to leucine at a position corresponding to position 308 of SEQ ID NO: 1. In some embodiments, the IdeS variant comprises a modification at a position corresponding to position 68 is a substitution of valine to threonine.

[0274] For example, in some embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to cysteine at a position corresponding to position 236 of SEQ ID NO: 1, a substitution of leucine to cysteine at a position corresponding to position 277 of SEQ ID NO: 1, and a substitution of isoleucine to leucine at a position corresponding to position 308 of SEQ ID NO: 1. In some embodiments, the IdeS variant comprises a modification at a position corresponding to position 68 is a substitution of valine to threonine.

[0275] In some embodiments, the IdeS variant protein comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises two or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises three or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeSAttorney Docket No. : CYR-009WO variant protein comprises four or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises five or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises six or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises seven or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises eight or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises nine or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises ten or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises eleven or more modifications at positions corresponding to positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant protein comprises modifications at positions corresponding to positions 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303 of SEQ ID NO: 1.

[0276] For example, in some embodiments, the IdeS variant protein comprises one or more of a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1; a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1; a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1; a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1; a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1; a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1; a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1; a substitution of threonine to glycine at a position corresponding to position 189 of SEQ ID NO: 1; a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1; a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ IDAttorney Docket No. : CYR-009WONO: 1; a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0277] For example, in some embodiments, the IdeS variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1; a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1; a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1; a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1; a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1; a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1; a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1; a substitution of threonine to glycine at a position corresponding to position 189 of SEQ ID NO: 1; a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1; a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1; a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

[0278] In some embodiments, the IdeS variant proteins further comprise a substitution of the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with one of the following sequences:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).Attorney Docket No. : CYR-009WO

[0279] In some embodiments, the IdeS variant proteins have at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90% at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5% or 100% sequence identity with the amino acid sequence of residues 21-310 of SEQ ID NO:2 and comprise a substitution of the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with one of the following sequences:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).

[0280] In one embodiment, the IdeS variant comprises the sequence TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1- 20 of SEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111,113, 115, 116, 127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187,188, 189, 195, 197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258,273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316,318, 322, and 330. In some embodiments, the IdeS variant comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.Attorney Docket No. : CYR-009WO

[0281] In one embodiment, the IdeS variant comprises the sequence DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111, 113, 115, 116,127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187, 188, 189, 195,197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258, 273, 274, 277,278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.In some embodiments, the IdeS variant comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0282] In one embodiment, the IdeS variant comprises the sequence DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO:79) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111,113, 115, 116, 127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187,188, 189, 195, 197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258,273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316,318, 322, and 330. In some embodiments, the IdeS variant comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0283] In one embodiment, the IdeS variant comprises the sequence DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO:80) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1- 20 of SEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ IDAttorney Docket No. : CYR-009WONO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111,113, 115, 116, 127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187,188, 189, 195, 197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258,273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316,318, 322, and 330. In some embodiments, the IdeS variant comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0284] In one embodiment, the IdeS variant comprises the sequence SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111, 113, 115, 116,127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187, 188, 189, 195,197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258, 273, 274, 277,278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.In some embodiments, the IdeS variant comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0285] In one embodiment, the IdeS variant comprises the sequence AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO:82) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111, 113, 115, 116,127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187, 188, 189, 195,197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258, 273, 274, 277,278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.In some embodiments, the IdeS variant comprises at least 90% sequence identity withAttorney Docket No. : CYR-009WO residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0286] In one embodiment, the IdeS variant comprises the sequence GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111, 113, 115, 116,127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187, 188, 189, 195,197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258, 273, 274, 277,278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.In some embodiments, the IdeS variant comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0287] In one embodiment, the IdeS variant comprises the sequence AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111, 113, 115, 116,127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187, 188, 189, 195,197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258, 273, 274, 277,278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.In some embodiments, the IdeS variant comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0288] In one embodiment, the IdeS variant comprises the sequenceQEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 ofAttorney Docket No. : CYR-009WOSEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111,113, 115, 116, 127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187,188, 189, 195, 197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258,273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316,318, 322, and 330. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303. In some embodiments, the IdeS variant further comprises modifications at positions corresponding to positions 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303 of SEQ ID NO: 1. In some embodiments, the IdeS variant comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0289] In one embodiment, the IdeS variant comprises the sequence QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174) substituted for the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2. In some embodiments, the IdeS variant further comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111,113, 115, 116, 127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187,188, 189, 195, 197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258,273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316,318, 322, and 330 of SEQ ID NO: 1. In some embodiments, the IdeS variant comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2. In some embodiments, the IdeS variant comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

[0290] In some embodiments, the IdeS variant proteins comprise a substitution of the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with a sequence having at least 80%, at least 81%, at leastAttorney Docket No. : CYR-009WO82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90% at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5% or 100% sequence identity with one of the following sequences:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).In some embodiments, the substitution is SEQ ID NO:85.

[0291] In some embodiments, the IdeS variant proteins comprise a substitution of the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with a sequence having at least 80% sequence identity with one of the following sequences:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).In some embodiments, the substitution is SEQ ID NO:85.

[0292] In some embodiments, the IdeS variant proteins comprise a substitution of the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding toAttorney Docket No. : CYR-009WO residues 1-20 of SEQ ID NO:2 with a sequence having at least 90% sequence identity with one of the following sequences:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).In some embodiments, the substitution is SEQ ID NO:85.

[0293] In some embodiments, the IdeS variant proteins comprise a substitution of the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with a sequence having at least 95% sequence identity with one of the following sequences:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).In some embodiments, the substitution is SEQ ID NO:85.

[0294] In some embodiments, provided herein are IdeS variants having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90% at least 91%, at least 92%, at least 93%, at least 94%, atAttorney Docket No. : CYR-009WO least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5% or 100% sequence identity with the amino acid sequence of residues 21-310 of SEQ ID NO:2.

[0295] In some embodiments, provided herein are IdeS variants having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90% at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5% or 100% sequence identity to any one of the IdeS variant sequences listed in Table 1 A.

[0296] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 80% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0297] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 90% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0298] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 95% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0299] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 96% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0300] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 97% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0301] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 97.5% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0302] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 98% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.Attorney Docket No. : CYR-009WO

[0303] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 98.5% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0304] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 99% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0305] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 99.5% sequence identity with the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0306] In some embodiments, the IdeS variant protein comprises the amino acid sequence of any one of SEQ ID NOs: 3-75, 86-94, or 172-173.

[0307] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 96% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 97% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 97.5% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 98% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 98.5% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 99.5% sequence identity with the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises the amino acid sequence of SEQ IDAttorney Docket No. : CYR-009WONO: 172. In some embodiments, the IdeS variant protein comprises the amino acid sequence of SEQ ID NO: 172.

[0308] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 96% sequence identity with the amino acid sequence of SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 97% sequence identity with the amino acid sequence of SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 97.5% sequence identity with the amino acid sequence of SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 98% sequence identity with the amino acid sequence of SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 98.5% sequence identity with the amino acid sequence of SEQ ID NO: 172 or SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 172 or SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 99.5% sequence identity with the amino acid sequence of SEQ ID NO: 172 or SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises the amino acid sequence of SEQ ID NO: 172 or SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises the amino acid sequence of SEQ ID NO: 173.

[0309] In some embodiments, the IdeS variant is an IdeS variant disclosed in Table 1 A.Attorney Docket No. : CYR-009WOTable 1A. IdeS Variant SequencesAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WO* bold and underlined = chimeric sequences at the N-terminus; bold = modifications as compared to SEQ ID NO: 1

[0310] Described herein, in some embodiments, are IdeS variant proteins comprising the amino acid sequence according to any one of SEQ ID NOs: 3-75, 86-94, and 172-173. In some embodiments, the IdeS variant protein comprises a sequence having at least 80%, atAttorney Docket No. : CYR-009WO least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, or 100% sequence identity with the amino acid sequence according to any one of SEQ ID NOs: 3-75, 86-94, and 172-173. In some embodiments, the IdeS variant protein comprises a sequence having at least 95% sequence identity with the amino acid sequence according to any one of SEQ ID NOs: 3-75, 86-94, and 172-173 and having one or more than one modifications to these enumerated amino acid sequences.

[0311] Described herein, in some embodiments, are IdeS variant proteins comprising the amino acid sequence according to SEQ ID NO: 10. In some embodiments, the IdeS variant protein comprises a sequence having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, or 100% sequence identity with the amino acid sequence according to SEQ ID NO: 10. In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 95% sequence identity with the amino acid sequence according to SEQ ID NO: 10 and having one or more than one modification.

[0312] Described herein, in some embodiments, are IdeS variant proteins comprising the amino acid sequence according to SEQ ID NO: 11. In some embodiments, the IdeS variant protein comprises a sequence having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, or 100% sequence identity with the amino acid sequence according to SEQ ID NO: 11. In some embodiments, the IdeS variant protein comprises a sequence having at least 95% sequence identity with the amino acid sequence according to SEQ ID NO: 11 and having one or more than one modifications.

[0313] Described herein, in some embodiments, are IdeS variant proteins comprising the amino acid sequence according to SEQ ID NO: 12. In some embodiments, the IdeS variant protein comprises a sequence having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, or 100% sequence identity with the amino acidAttorney Docket No. : CYR-009WO sequence according to SEQ ID NO: 12. In some embodiments, the IdeS variant protein comprises a sequence having at least 95% sequence identity with the amino acid sequence according to SEQ ID NO: 12 and having one or more than one modifications.

[0314] Described herein, in some embodiments, are IdeS variant proteins comprising the amino acid sequence according to SEQ ID NO: 13. In some embodiments, the IdeS variant protein comprises a sequence having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, or 100% sequence identity with the amino acid sequence according to SEQ ID NO: 13. In some embodiments, the IdeS variant protein comprises a sequence having at least 95% sequence identity with the amino acid sequence according to SEQ ID NO: 13 and having one or more than one modification.

[0315] Described herein, in some embodiments, are IdeS variant proteins comprising the amino acid sequence according to SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises a sequence having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, or 100% sequence identity with the amino acid sequence according to SEQ ID NO: 172. In some embodiments, the IdeS variant protein comprises a sequence having at least 95% sequence identity with the amino acid sequence according to SEQ ID NO: 172 and having one or more than one modification. In some embodiments, the IdeS variant protein comprises a sequence having at least 98% sequence identity with the amino acid sequence according to SEQ ID NO: 172 and having one or more than one modification. In some embodiments, the IdeS variant protein comprises a sequence having at least 99% sequence identity with the amino acid sequence according to SEQ ID NO: 172 and having one or more than one modification.

[0316] Described herein, in some embodiments, are IdeS variant proteins comprising the amino acid sequence according to SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises a sequence having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, or 100% sequence identity with the amino acidAttorney Docket No. : CYR-009WO sequence according to SEQ ID NO: 173. In some embodiments, the IdeS variant protein comprises a sequence having at least 95% sequence identity with the amino acid sequence according to SEQ ID NO: 173 and having one or more than one modification. In some embodiments, the IdeS variant protein comprises a sequence having at least 98% sequence identity with the amino acid sequence according to SEQ ID NO: 173 and having one or more than one modification. In some embodiments, the IdeS variant protein comprises a sequence having at least 99% sequence identity with the amino acid sequence according to SEQ ID NO: 173 and having one or more than one modification.

[0317] In some embodiments, the IdeS variant proteins comprise one or more glycosylation modifications.

[0318] In some embodiments, the IdeS variant proteins comprise one or more glycosylation modifications as compared to wild-type. In some embodiments, the glycosylation modification(s) is / are at one or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, or 319 of SEQ ID NO: 1. In some embodiments, the glycosylation modification is at two or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, or 319 of SEQ ID NO: 1. In some embodiments, the glycosylation modification is at three or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, or 319 of SEQ ID NO: 1. In some embodiments, the glycosylation modification is at four or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, or 319 of SEQ ID NO: 1. In some embodiments, the glycosylation modification is at five or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, or 319 of SEQ ID NO: 1. In some embodiments, the glycosylation modification is at six or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, or 319 of SEQ ID NO: 1. In some embodiments, the glycosylation modification is at seven or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, or 319 of SEQ ID NO: 1. In some embodiments, the glycosylation modification is at ten or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, orAttorney Docket No. : CYR-009WO319 of SEQ ID NO: 1. In some embodiments, the glycosylation modification is at twelve or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, or 319 of SEQ ID NO: 1. In some embodiments, the glycosylation modification is at fifteen or more positions corresponding to position(s) 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, or 319 of SEQ ID NO: 1.

[0319] In some embodiments, the glycosylation modification occurs at a position corresponding to position 31 of SEQ ID NO: 1 where a serine is substituted to asparagine.

[0320] In some embodiments, the glycosylation modification occurs at a position corresponding to position 37 of SEQ ID NO: 1 where a glutamate is substituted to asparagine.

[0321] In some embodiments, the glycosylation modification occurs at a position corresponding to position 39 of SEQ ID NO: 1 where arginine is substituted to threonine or asparagine.

[0322] In some embodiments, the glycosylation modification occurs at a position corresponding to position 42 of SEQ ID NO: 1 where glutamate is substituted to asparagine.

[0323] In some embodiments, the glycosylation modification occurs at a position corresponding to position 74 of SEQ ID NO: 1 where valine is substituted to asparagine.

[0324] In some embodiments, the glycosylation modification occurs at a position corresponding to position 76 of SEQ ID NO: 1 where asparagine is substituted to serine.

[0325] In some embodiments, the glycosylation modification occurs at a position corresponding to position 111 of SEQ ID NO: 1 where lysine is substituted to asparagine.

[0326] In some embodiments, the glycosylation modification occurs at a position corresponding to position 113 of SEQ ID NO: 1 where glutamine is substituted to threonine.

[0327] In some embodiments, the glycosylation modification occurs at a position corresponding to position 119 of SEQ ID NO: 1 where glutamate is substituted to asparagine.

[0328] In some embodiments, the glycosylation modification occurs at a position corresponding to position 121 of SEQ ID NO: 1 where a histidine is substituted to threonine.Attorney Docket No. : CYR-009WO

[0329] In some embodiments, the glycosylation modification occurs at a position corresponding to position 130 of SEQ ID NO: 1 where asparagine is substituted to serine.

[0330] In some embodiments, the glycosylation modification occurs at a position corresponding to position 142 of SEQ ID NO: 1 where aspartate is substituted to asparagine.

[0331] In some embodiments, the glycosylation modification occurs at a position corresponding to position 144 of SEQ ID NO: 1 where lysine is substituted to threonine.

[0332] In some embodiments, the glycosylation modification occurs at a position corresponding to position 147 of SEQ ID NO: 1 where glutamine is substituted to serine.

[0333] In some embodiments, the glycosylation modification occurs at a position corresponding to position 148 of SEQ ID NO: 1 where leucine is substituted to asparagine.

[0334] In some embodiments, the glycosylation modification occurs at a position corresponding to position 198 of SEQ ID NO: 1 where glutamate is substituted to asparagine.

[0335] In some embodiments, the glycosylation modification occurs at a position corresponding to position 233 of SEQ ID NO: 1 where lysine is substituted to serine.

[0336] In some embodiments, the glycosylation modification occurs at position 244 of SEQ ID NO: 1 where threonine is substituted to asparagine.

[0337] In some embodiments, the glycosylation modification occurs at a position corresponding to position 246 of SEQ ID NO: 1 where glycine is substituted to threonine.

[0338] In some embodiments, the glycosylation modification occurs at a position corresponding to position 311 of SEQ ID NO: 1 where lysine is substituted to asparagine.

[0339] In some embodiments, the glycosylation modification occurs at a position corresponding to position 313 of SEQ ID NO: 1 where isoleucine is substituted to threonine.

[0340] In some embodiments, the glycosylation modification occurs at a position corresponding to position 319 of SEQ ID NO: 1 where a glycine is substituted to serine.

[0341] In some embodiments, the IdeS variant protein comprises a substitution of glutamate to asparagine at a position corresponding to position 37 of SEQ ID NO: 1 and aAttorney Docket No. : CYR-009WO substitution of arginine to threonine at a position corresponding to position 39 of SEQ ID NO: 1.

[0342] In some embodiments, the IdeS variant protein comprises a substitution of valine to asparagine at a position corresponding to position 74 of SEQ ID NO: 1 and a substitution of asparagine to serine at a position corresponding to position 76 of SEQ ID NO: 1.

[0343] In some embodiments, the IdeS variant protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1 and a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1.

[0344] In some embodiments, the IdeS variant protein comprises a substitution of glutamate to asparagine at a position corresponding to position 119 of SEQ ID NO: 1 and a substitution of histidine to threonine at a position corresponding to position 121 of SEQ ID NO: 1.

[0345] In some embodiments, the IdeS variant protein comprises a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 1 and a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 1.

[0346] In some embodiments, the IdeS variant protein comprises a substitution of threonine to asparagine at a position corresponding to position 244 of SEQ ID NO: 1 and a substitution of glycine to threonine at a position corresponding to position 246 of SEQ ID NO: 1.

[0347] In some embodiments, the IdeS variant protein comprises a substitution of lysine to asparagine at a position corresponding to position 311 of SEQ ID NO: 1 and a substitution of isoleucine to threonine at a position corresponding to position 313 of SEQ ID NO: 1.

[0348] In some embodiments, provided herein are IdeS variants comprising at least one modification at a position corresponding to a position in SEQ ID NO: 12 selected from 47, 49, 51, 78, 111, 113, 123, 125, 126, 128, 142, 144, 148, 198, 273, 275, 278, 312, and 314 to introduce a glycosylation site. For example, the IdeS variant comprises modifications at at least one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen,Attorney Docket No. : CYR-009WO fifteen, sixteen, seventeen, eighteen, or nineteen of the aforementioned positions corresponding to positions in SEQ ID NO: 12.

[0349] In some embodiments, the modification is a substitution of threonine to asparagine at a position corresponding to position 47 of SEQ ID NO: 12.

[0350] In some embodiments, the modification is a substitution of glutamine to threonine or asparagine at a position corresponding to position 49 of SEQ ID NO: 12.

[0351] In some embodiments, the modification is a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12. In some embodiments, the modification is a substitution of glycine to serine at a position corresponding to position 78 of SEQ ID NO: 12.

[0352] In some embodiments, the modification is a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12.

[0353] In some embodiments, the modification is a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12.

[0354] In some embodiments, the modification is a substitution of glutamate to asparagine at a position corresponding to position 123 of SEQ ID NO: 12.

[0355] In some embodiments, the modification is a substitution of glutamine to threonine at a position corresponding to position 125 of SEQ ID NO: 12.

[0356] In some embodiments, the modification is a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12.

[0357] In some embodiments, the modification is a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12.

[0358] In some embodiments, the modification is a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 12.

[0359] In some embodiments, the modification is a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 12.Attorney Docket No. : CYR-009WO

[0360] In some embodiments, the modification is a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 12.

[0361] In some embodiments, the modification is a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0362] In some embodiments, the modification is a substitution of serine to asparagine at a position corresponding to position 273 of SEQ ID NO: 12.

[0363] In some embodiments, the modification is a substitution of glycine to serine at a position corresponding to position 275 of SEQ ID NO: 12.

[0364] In some embodiments, the modification is a substitution of lysine to serine at a position corresponding to position 278 of SEQ ID NO: 12.

[0365] In some embodiments, the modification is a substitution of glutamate to asparagine at a position corresponding to position 312 of SEQ ID NO: 12.

[0366] In some embodiments, the modification is a substitution of lysine to serine at a position corresponding to position 314 of SEQ ID NO: 12.

[0367] In some embodiments, the variant IdeS protein comprises a substitution of threonine to asparagine at a position corresponding to position 47 of SEQ ID NO: 12 and a substitution of glutamine to threonine at a position corresponding to position 49 of SEQ ID NO: 12.

[0368] In some embodiments, the variant IdeS protein comprises a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12 and a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12.

[0369] In some embodiments, the variant IdeS protein comprises a substitution of glutamate to asparagine at a position corresponding to position 123 of SEQ ID NO: 12 and a substitution of glutamine to threonine at a position corresponding to position 125 of SEQ ID NO: 12.Attorney Docket No. : CYR-009WO

[0370] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12 and a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12.

[0371] In some embodiments, the variant IdeS protein comprises a substitution of serine to asparagine at a position corresponding to position 273 of SEQ ID NO: 12 and a substitution of glycine to serine at a position corresponding to position 275 of SEQ ID NO: 12.

[0372] In some embodiments, the variant IdeS protein comprises a substitution of glutamate to asparagine at a position corresponding to position 312 of SEQ ID NO: 12 and a substitution of lysine to serine at a position corresponding to position 314 of SEQ ID NO: 12.

[0373] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 12, and a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 12.

[0374] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0375] In some embodiments, the variant IdeS protein comprises a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 12, a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0376] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 12, a substitution of lysine to threonine at a position corresponding to position 144 ofAttorney Docket No. : CYR-009WOSEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0377] In some embodiments, the variant IdeS protein comprises a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0378] In some embodiments, the variant IdeS protein comprises a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0379] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0380] In some embodiments, the variant IdeS protein comprises a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12, a substitution of valine to threonine at a position corresponding to position 51 of SEQ IDAttorney Docket No. : CYR-009WONO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 12, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0381] In some embodiments, the variant IdeS protein comprises a substitution of threonine to asparagine at a position corresponding to position 47 of SEQ ID NO: 12, a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0382] In some embodiments, the variant IdeS protein comprises a substitution of glutamine to asparagine at a position corresponding to position 49 of SEQ ID NO: 12, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0383] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of glutamate to asparagine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 125 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0384] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ IDAttorney Docket No. : CYR-009WONO: 12, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0385] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 12, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 12, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 12, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 12.

[0386] In some embodiments, provided herein are IdeS variants comprising at least one modification at a position corresponding to a position in SEQ ID NO: 1 selected from 47, 49, 51, 78, 111, 113, 123, 125, 126, 128, 142, 144, 148, 198, 273, 275, 278, 312, and 314 to introduce a glycosylation site. For example, the IdeS variant comprises modifications at at least one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, or nineteen of the aforementioned positions corresponding to positions in SEQ ID NO: 1.

[0387] In some embodiments, the modification is a substitution of histidine to asparagine at a position corresponding to position 47 of SEQ ID NO: 1.

[0388] In some embodiments, the modification is a substitution of threonine to asparagine at a position corresponding to position 49 of SEQ ID NO: 1

[0389] In some embodiments, the modification is a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 1. In some embodiments, the modification is a substitution of glycine to serine at a position corresponding to position 78 of SEQ ID NO: 1.

[0390] In some embodiments, the modification is a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1.

[0391] In some embodiments, the modification is a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1.Attorney Docket No. : CYR-009WO

[0392] In some embodiments, the modification is a substitution of glutamate to asparagine at a position corresponding to position 123 of SEQ ID NO: 1.

[0393] In some embodiments, the modification is a substitution of glutamine to threonine at a position corresponding to position 125 of SEQ ID NO: 1.

[0394] In some embodiments, the modification is a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 1.

[0395] In some embodiments, the modification is a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 1.

[0396] In some embodiments, the modification is a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 1.

[0397] In some embodiments, the modification is a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 1.

[0398] In some embodiments, the modification is a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 1.

[0399] In some embodiments, the modification is a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0400] In some embodiments, the modification is a substitution of serine to asparagine at a position corresponding to position 273 of SEQ ID NO: 1.

[0401] In some embodiments, the modification is a substitution of glycine to serine at a position corresponding to position 275 of SEQ ID NO: 1.

[0402] In some embodiments, the modification is a substitution of lysine to serine at a position corresponding to position 278 of SEQ ID NO: 1.

[0403] In some embodiments, the modification is a substitution of glutamate to asparagine at a position corresponding to position 312 of SEQ ID NO: 1.

[0404] In some embodiments, the modification is a substitution of lysine to serine at a position corresponding to position 314 of SEQ ID NO: 1.Attorney Docket No. : CYR-009WO

[0405] In some embodiments, the variant IdeS protein comprises a substitution of histidine to asparagine at a position corresponding to position 47 of SEQ ID NO: 1 and a substitution of threonine to asparagine at a position corresponding to position 49 of SEQ ID NO: 1.

[0406] In some embodiments, the variant IdeS protein comprises a substitution of threonine to asparagine at a position corresponding to position 49 of SEQ ID NO:2 and a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 1.

[0407] In some embodiments, the variant IdeS protein comprises a substitution of glutamate to asparagine at a position corresponding to position 123 of SEQ ID NO: 1 and a substitution of glutamine to threonine at a position corresponding to position 125 of SEQ ID NO: 1.

[0408] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 1 and a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 1.

[0409] In some embodiments, the variant IdeS protein comprises a substitution of serine to asparagine at a position corresponding to position 273 of SEQ ID NO: 1 and a substitution of glycine to serine at a position corresponding to position 275 of SEQ ID NO: 1.

[0410] In some embodiments, the variant IdeS protein comprises a substitution of glutamate to asparagine at a position corresponding to position 312 of SEQ ID NO: 1 and a substitution of lysine to serine at a position corresponding to position 314 of SEQ ID NO: 1.

[0411] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 1, and a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 1.

[0412] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, and aAttorney Docket No. : CYR-009WO substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0413] In some embodiments, the variant IdeS protein comprises a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 1, a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0414] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of aspartate to asparagine at a position corresponding to position 142 of SEQ ID NO: 1, a substitution of lysine to threonine at a position corresponding to position 144 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0415] In some embodiments, the variant IdeS protein comprises a substitution of threonine to asparagine at a position corresponding to position 49 of SEQ ID NO: 1, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 1, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 1, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0416] In some embodiments, the variant IdeS protein comprises a substitution of threonine to asparagine at a position corresponding to position 49 of SEQ ID NO: 1, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of leucine to asparagine at a position corresponding to positionAttorney Docket No. : CYR-009WO148 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0417] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 1, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 1, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0418] In some embodiments, the variant IdeS protein comprises a substitution of threonine to asparagine at a position corresponding to position 49 of SEQ ID NO: 1, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 1, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0419] In some embodiments, the variant IdeS protein comprises a substitution of histidine to asparagine at a position corresponding to position 47 of SEQ ID NO: 1, a substitution of threonine to asparagine at a position corresponding to position 49 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0420] In some embodiments, the variant IdeS protein comprises a substitution of threonine to asparagine at a position corresponding to position 49 of SEQ ID NO: 1, a substitution of valine to threonine at a position corresponding to position 51 of SEQ ID NO: 12, a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to positionAttorney Docket No. : CYR-009WO113 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0421] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of glutamate to asparagine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 125 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0422] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of lysine to asparagine at a position corresponding to position 126 of SEQ ID NO: 1, a substitution of asparagine to serine at a position corresponding to position 128 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0423] In some embodiments, the variant IdeS protein comprises a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1, a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1, a substitution of leucine to asparagine at a position corresponding to position 148 of SEQ ID NO: 1, and a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1.

[0424] In some embodiments, provided herein are IdeS variants having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90% at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5% or 100% sequence identity to any one of the IdeS variant sequences listed in Table IB.

[0425] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 80% sequence identity with any one of SEQ ID NOs: 117-153.Attorney Docket No. : CYR-009WO

[0426] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 90% sequence identity any one of SEQ ID NOs: 117-153.

[0427] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 95% sequence identity with any one of SEQ ID NOs: 117-153.

[0428] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 96% sequence identity with any one of SEQ ID NOs: 117-153.

[0429] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 96.5% sequence identity with any one of SEQ ID NOs: 117-153.

[0430] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 97% sequence identity with any one of SEQ ID NOs: 117-153.

[0431] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 97.5% sequence identity with any one of SEQ ID NOs: 117-153.

[0432] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 98% sequence identity with any one of SEQ ID NOs: 117-153.

[0433] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 98.5% sequence identity with any one of SEQ ID NOs: 117-153.

[0434] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 99% sequence identity with any one of SEQ ID NOs: 117-153.

[0435] In some embodiments, the IdeS variant protein comprises an amino acid sequence having at least 99.5% sequence identity with any one of SEQ ID NOs: 117-153.

[0436] In some embodiments, the IdeS variant protein comprises the amino acid sequence of any one of SEQ ID NOs: 117-153.

[0437] In some embodiments, the IdeS variant protein comprises a IdeS glycosylation variant in Table IB.Attorney Docket No. : CYR-009WOTable IB. IdeS Glycosylation VariantsAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WO* Mutation / s in bold; site of N-glycosylation is underlined

[0438] Further described herein, in some embodiments, are IdeS variant proteins comprising one or more glycans. In some embodiments, the one or more glycans are glycans of a mammalian glycoprotein. In some embodiments, the one or more glycans are N-linked glycans (z.e., N-glycans) or O-linked glycans (z.e., O-glycans) of a mammalian glycoprotein. In some embodiments, the one or more glycans are N-linked glycans (z.e., N-glycans). In some embodiments the one or more glycans comprise an N-glycan core structure. In some embodiments the one or more glycans comprise an N-glycan core structure of: (Manal- 6[Manal-3)]Man(pi-4)GlcNAc(pi-4)GlcNAc(pi-Asn-X-Ser / Thr). In some embodiments, the one or more glycans are selected from the group consisting of: a biantennary glycan, a triantennary glycan, and a tetraantennary glycan.

[0439] In some embodiments, the one or more glycans terminate with a galactose, an N- acetylgalactosamine, a glucose, an N-acetylglucosamine, a fucose, an N-acetylneuraminic acid, a mannose or a sialic acid. In some embodiments, the one or more glycans terminate with a galactose. In some embodiments, the one or more glycans terminate with an N- acetylgalactosamine. In some embodiments, the one or more glycans terminate with a glucose. In some embodiments, the one or more glycans terminate with an N- acetylglucosamine. In some embodiments, the one or more glycans terminate with a fucose. In some embodiments, the one or more glycans terminate with an N-acetylneuraminic acid. In some embodiments, the one or more glycans terminate with a mannose. In some embodiments, the one or more glycans terminate with a sialic acid.

[0440] In some embodiments, the one or more glycans is selected from the group consisting of: a biantennary glycan that terminates with mannose, a biantennary glycan thatAttorney Docket No. : CYR-009WO terminates with sialic acid, a triantennary glycan that terminates with a mannose, a triantennary glycan that terminates with a sialic acid, a tetraantennary glycan that terminates with mannose, and a tetraantennary glycan that terminates with sialic acid. In some embodiments, the one or more glycans is a biantennary glycan that terminates with a galactose. In some embodiments, the one or more glycans is a biantennary glycan that terminates with an N-acetylgalactosamine. In some embodiments, the one or more glycans is a biantennary glycan that terminates with a glucose. In some embodiments, the one or more glycans is a biantennary glycan that terminates with an N-acetylglucosamine. In some embodiments, the one or more glycans is a biantennary glycan that terminates with a fucose. In some embodiments, the one or more glycans is a biantennary glycan that terminates with an N-acetylneuraminic acid. In some embodiments, the one or more glycans is a biantennary glycan that terminates with mannose. In some embodiments, the one or more glycans is a biantennary glycan that terminates with sialic acid. In some embodiments, the one or more glycans is a triantennary glycan that terminates with a galactose. In some embodiments, the one or more glycans is a triantennary glycan that terminates with an N-acetylgalactosamine. In some embodiments, the one or more glycans is a triantennary glycan that terminates with a glucose. In some embodiments, the one or more glycans is a triantennary glycan that terminates with an N-acetylglucosamine. In some embodiments, the one or more glycans is a triantennary glycan that terminates with a fucose. In some embodiments, the one or more glycans is a triantennary glycan that terminates with an N-acetylneuraminic acid. In some embodiments, the one or more glycans is a triantennary glycan that terminates with a mannose. In some embodiments, the one or more glycans is a triantennary glycan that terminates with a sialic acid. In some embodiments, the one or more glycans is a tetraantennary glycan that terminates with a galactose. In some embodiments, the one or more glycans is a tetraantennary glycan that terminates with an N-acetylgalactosamine. In some embodiments, the one or more glycans is a tetraantennary glycan that terminates with a glucose. In some embodiments, the one or more glycans is a tetraantennary glycan that terminates with an N-acetylglucosamine. In some embodiments, the one or more glycans is a tetraantennary glycan that terminates with a fucose. In some embodiments, the one or more glycans is a tetraantennary glycan that terminates with an N-acetylneuraminic acid. In some embodiments, the one or more glycans is a tetraantennary glycan that terminates with mannose, and a tetraantennary glycan that terminates with sialic acid. In some embodiments, the one or more glycans is a tetraantennary glycan that terminates with sialic acid.Attorney Docket No. : CYR-009WO

[0441] In some embodiments, the glycan is selected from the glycans listed in Table 10. In some embodiments, the glycan is selected from the group consisting of Man(al-6)[Man(al-3)]Man(pi-4)GlcNAc(pi-4)GlcNAc(pi-ASN) (i.e., Glycan 1), Neu5Ac(a2-6)Gal(pi-4)GlcNAc(P 1 -2)Man(al -6)[Neu5 Ac(a2-6)Gal(P 1 -4)GlcNAc(P 1 -2)Man(al -3)]Man(P 1 - 4)GlcNAc(pi-4)GlcNAc(pi-ASN) (i.e., Glycan 2), and Neu5Ac(a2-6)Gal(pi-4)GlcNAc(pi- 6) [Neu5 Ac(a2-6)Gal(P 1 -4)GlcNAc(P 1 -2)]Man(al -6)[Neu5 Ac(a2-6)Gal(P 1 -4)GlcNAc(P 1 - 2)[Neu5 Ac(a2-6)Gal(P 1 -4)GlcNAc(P 1 -4)]Man(al -3)]Man(P 1 -4)GlcNAc(P 1 -4)GlcNAc(P 1 - ASN) (i.e., Glycan 3).

[0442] In some embodiments the one or more glycans terminate with an N- acetylglucosamine i.e. GlcNAc).

[0443] In some embodiments, the one or more glycans shield the surface area of the IdeS variant protein. In some embodiments, the one or more glycans shield at least 10-64% of the total accessible surface area of the IdeS variant protein. In some embodiments, the one or more glycans shield at least 20-50% of the total accessible surface area of the IdeS variant protein. In some embodiments, the one or more glycans shield at least 25-45% of the total accessible surface area of the IdeS variant protein. In some embodiments, the one or more glycans shield at least 20-35% of the total accessible surface area of the IdeS variant protein. In some embodiments, the one or more glycans shield at least 25% of the total accessible surface area of the IdeS variant protein. In some embodiments, the one or more glycans shield at least 30% of the total accessible surface area of the IdeS variant protein. In some embodiments, the one or more glycans shield at least 35% of the total accessible surface area of the IdeS variant protein. In some embodiments, the one or more glycans shield at least 40% of the total accessible surface area of the IdeS variant protein. In some embodiments, the one or more glycans shield at least 45% of the total accessible surface area of the IdeS variant protein.

[0444] In some embodiments, the IdeS variant proteins comprising one or more glycans have reduced immunogenicity compared to aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, or wild-type IdeS. In some embodiments, the IdeS variant proteins comprising one or more glycans have increased serum stability compared to aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, or wild-type IdeS. In some embodiments, the IdeS variant proteins comprising one or more glycans persist longer in serum than aglycosylated IdeS variant proteins, IdeS variant proteins with fewerAttorney Docket No. : CYR-009WO glycans, or wild-type IdeS. In some embodiments, the IdeS variant proteins comprising one or more glycans have a serum half-life approximately two times longer than the serum halflife of aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, or wildtype IdeS. In some embodiments, the IdeS variant proteins comprising one or more glycans have increased size (z.e., Molecular Weight (MW)) compared to aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, or wild-type IdeS. In some embodiments, the IdeS variant proteins comprising one or more glycans have reduced renal elimination compared to aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, or wild-type IdeS. In some embodiments, reduced renal elimination is due to an increase in the IdeS variant protein’s size (z.e., MW) as compared to aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, or wild-type IdeS. In some embodiments, the IdeS variant proteins comprising one or more glycans have a longer duration of IgG depletion compared to aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, or wild-type IdeS. In some embodiments, the longer duration of IgG depletion is correlated with a greater extent of IdeS variant protein glycosylation.

[0445] Further described herein, in some embodiments, are IdeS variant proteins fused to Human Serum Albumin (HSA). In some embodiments, the IdeS variant proteins fused to HSA comprise one or more glycans. In some embodiments, the IdeS variant proteins fused to HSA have a reduced EC50 value for polyclonal anti-IdeS reactivity compared to the EC50 value of aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, IdeS variant proteins not fused to HSA, or wild-type IdeS. In some embodiments, the EC50 value of IdeS variant proteins fused to HSA is approximately half the EC50 value of aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, IdeS variant proteins not fused to HSA, or wild-type IdeS. In some embodiments, the IdeS variant proteins fused to HSA have a longer duration of IgG depletion compared to aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, IdeS variant proteins not fused to HSA, or wild-type IdeS. In some embodiments, the IdeS variant proteins fused to HSA have a serum half-life approximately two, three, four, five, six, or seven times longer than the serum halflife of aglycosylated IdeS variant proteins, IdeS variant proteins with fewer glycans, IdeS variant proteins not fused to HSA, or wild-type IdeS.Attorney Docket No. : CYR-009WO2. IdeS Polypeptides Conjugates

[0446] Described herein, in certain embodiments, are IdeS polypeptide conjugates. In some embodiments, the IdeS polypeptide conjugates are polypeptides comprising an IdeS protein moiety (e.g., a Ides variant or wild-type IdeS protein) conjugated to either a human serum albumin moiety (e.g., naturally occurring or variant serum albumin), or an albumin binding domain moiety. The IdeS protein moiety may be conjugated to the human serum albumin moiety by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171. Any of the IdeS variants described in the Section 1.1 above entitled “IdeS Variants" is suitable for inclusion as the IdeS protein moiety in an IdeS polypeptide conjugate as disclosed herein.

[0447] For example, disclosed are IdeS polypeptide conjugates comprising a variant Ides protein comprising at least 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1-75 or 172-173 conjugated to a human serum albumin or a variant human serum albumin. In some embodiments, the human serum albumin has the amino acid sequence of SEQ ID NO: 188. In some embodiments, the human serum albumin variant has the amino acid sequence of SEQ ID NO: 189. In some embodiments, the human serum albumin variant has the amino acid sequence of SEQ ID NO: 190. In some embodiments, the human serum albumin has at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least, at least 99.8%, or at least 99.9% sequence identity with the amino acid sequence of SEQ ID NO: 188. In some embodiments, the variant human serum albumin has at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least, at least 99.8%, or at least 99.9% sequence identity with the amino acid sequence of SEQ ID NO: 188. In some embodiments, the human serum albumin or human serum albumin variant is conjugated to a N-terminus of the IdeS variant protein. In some embodiments, the human serum albumin or human serum albumin variant is conjugated to a C-terminus of the IdeS variant protein. In some embodiments, the IdeS polypeptide conjugates comprise at least 90% sequence identity to the amino acid sequence of SEQ ID NOs: 172 or 173, e.g., 173. The IdeS variant protein may be connected to the human serum albumin or human serumAttorney Docket No. : CYR-009WO albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0448] For example, in certain embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising at least 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1-75 or 172-173 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 188. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0449] For example, in other embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising at least 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1-75 or 172-173 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 189. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0450] For example, in other embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising at least 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1-75 or 172-173 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 190. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0451] For example, in other embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising at least 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1-75 and 172-173 conjugated to a human serum albumin having at least at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least, at least 99.8%, or at least 99.9% with the amino acid sequence of SEQ ID NO: 188. In some embodiments, the IdeS orAttorney Docket No. : CYR-009WOIdeS variant protein comprises at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.5% or 100% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1-75 and 172-173. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0452] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of any one of SEQ ID NOs: 1-75 and 172-173 conjugated to a human serum albumin having at least at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least, at least 99.8%, or at least 99.9% with the amino acid sequence of SEQ ID NO: 188. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0453] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of any one of SEQ ID NOs: 1-75 and 172-173 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 188. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0454] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of any one of SEQ ID NOs: 1-75 and 172-173 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 189. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0455] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of any one ofAttorney Docket No. : CYR-009WOSEQ ID NOs: 1-75 and 172-173 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 190. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0456] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 172 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 188. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0457] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 173 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 188. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0458] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 172 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 189. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0459] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 173 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 189. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0460] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 172 conjugated to a human serum albumin having the amino acid sequence of SEQ IDAttorney Docket No. : CYR-009WONO: 190. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0461] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 173 conjugated to a human serum albumin having the amino acid sequence of SEQ ID NO: 190. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0462] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 172 conjugated to a human serum albumin having at least at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least, at least 99.8%, or at least 99.9% with the amino acid sequence of SEQ ID NO: 188. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0463] For example, in some embodiments, the disclosed IdeS polypeptide conjugates comprise an IdeS or IdeS variant protein comprising the amino acid sequence of SEQ ID NO: 173 conjugated to a human serum albumin having at least at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least, at least 99.8%, or at least 99.9% with the amino acid sequence of SEQ ID NO: 188. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0464] Further described herein, in certain embodiments, are IdeS polypeptide conjugates comprising an IdeS variant protein comprising at least 90% sequence identity to the aminoAttorney Docket No. : CYR-009WO acid sequence of SEQ ID NO: 172 conjugated to a human serum albumin or a variant human serum albumin. In some embodiments, the human serum albumin has at least at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least, at least 99.8%, or at least 99.9% with the amino acid sequence of SEQ ID NO: 188. In some embodiments, the human serum albumin has the amino acid sequence of SEQ ID NO: 188. In some embodiments, the human serum albumin variant has the amino acid sequence of SEQ ID NO: 189. In some embodiments, the human serum albumin variant has the amino acid sequence of SEQ ID NO: 190. In some embodiments, the human serum albumin is conjugated to an N-terminus of the IdeS variant protein. In some embodiments, the human serum albumin is conjugated to a C-terminus of the IdeS variant protein. In some embodiments, the IdeS or IdeS variant protein comprises at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.5% or 100% sequence identity to the amino acid sequence of SEQ ID NO: 172. In some embodiments, the IdeS variant comprises the amino acid sequence of SEQ ID NO: 172 and the human serum albumin comprises the amino acid sequence of SEQ ID NO: 189. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0465] Further described herein, in certain embodiments, are IdeS polypeptide conjugates comprising an IdeS variant protein comprising at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 173 conjugated to a human serum albumin or a variant human serum albumin. In some embodiments, the human serum albumin has at least at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least, at least 99.8%, or at least 99.9% with the amino acid sequence of SEQ ID NO: 188. In some embodiments, the human serum albumin has the amino acid sequence of SEQ ID NO: 188. In some embodiments, the human serum albumin variant has the amino acid sequence of SEQ ID NO: 189. In some embodiments, the human serum albumin has the amino acid sequence of SEQ ID NO: 190. In some embodiments, the human serum albuminAttorney Docket No. : CYR-009WO is conjugated to a N-terminus of the IdeS variant protein. In some embodiments, the human serum albumin is conjugated to a C-terminus of the IdeS variant protein. In some embodiments, the IdeS or IdeS variant protein comprises at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 95%, at least 95.5%, at least 96%, at least 96.5%, at least 97%, at least 97.5%, at least 98%, at least 98.5%, at least 99%, at least 99.5% or 100% sequence identity to the amino acid sequence of SEQ ID NO: 173. In some embodiments, the IdeS variant comprises the amino acid sequence of SEQ ID NO: 173 and the human serum albumin comprises the amino acid sequence of SEQ ID NO: 189. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0466] In some embodiments, the IdeS polypeptide conjugates comprise an Ides variant protein and a human serum albumin. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having at least 80% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186 or 187. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having at least 90% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186, or 187. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having at least 95% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186, or 187. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having at least 96% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186, or 187. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having at least 97% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186, or 187. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having at least 98% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186, or 187. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having at least 98.5% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186, or 187. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having at least 99% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186, or 187. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having at least 99.5% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186, or 187. In some embodiments, the IdeS polypeptide conjugate comprises an amino acid sequence having 100% sequence identity with any one of SEQ ID NOs: 154-161, 177, 186, or 187.Attorney Docket No. : CYR-009WO

[0467] In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table 1 A and the human serum albumin comprises the amino acid sequence of SEQ ID NO: 188. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table IB and the human serum albumin comprises the amino acid sequence of SEQ ID NO: 188. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein is selected from any of the variants disclosed in Table 1 A and the human serum albumin comprises the amino acid sequence of SEQ ID NO: 189. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein is selected from any of the variants disclosed in Table IB and the human serum albumin comprises the amino acid sequence of SEQ ID NO: 189. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein is selected from any of the variants disclosed in Table 1 A and the human serum albumin comprises the amino acid sequence of SEQ ID NO: 190. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein is selected from any of the variants disclosed in Table IB and the human serum albumin comprises the amino acid sequence of SEQ ID NO: 190. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0468] In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table 1 A and the human serum albumin comprises at least 90% sequence identity, e.g. at least 95% sequence identity, with the amino acid sequence of SEQ ID NO: 188. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table IB and the human serum albumin comprises at least 90% sequence identity, e.g. at least 95% sequence identity, with the amino acid sequence of SEQ ID NO: 188. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0469] In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table 1 A and the human serum albumin comprises at least 90% sequence identity, e.g. at least 95% sequence identity, with the aminoAttorney Docket No. : CYR-009WO acid sequence of SEQ ID NO: 189. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table IB and the human serum albumin comprises at least 90% sequence identity, e.g. at least 95% sequence identity, with the amino acid sequence of SEQ ID NO: 189. The IdeS variant protein may be connected to the human serum albumin or human serum albumin variant by a peptide linker disclosed herein, e.g., a linker having an amino acid sequence according to any one of SEQ ID NOS: 162-171.

[0470] In some embodiments, the IdeS polypeptide conjugate comprises an albumin binding domain. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 80% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 90% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 95% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 96% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 97% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 97.5% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 98% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 98.5% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 99% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having at least 99.5% sequence identity with any one of SEQ ID NOs: 178-184. In some embodiments, the IdeS polypeptide comprises an amino acid sequence having 100% sequence identity with any one of SEQ ID NOs: 178-184. The IdeS variant protein may be connected to the albumin binding domain by a peptide linker disclosed herein.

[0471] In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 95% identity with SEQ ID NO: 186. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 96% identity withAttorney Docket No. : CYR-009WOSEQ ID NO: 186. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 97% identity with SEQ ID NO: 186. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 98% identity with SEQ ID NO: 186. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99% identity with SEQ ID NO: 186. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99.5% identity with SEQ ID NO: 186. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99.7% identity with SEQ ID NO:186. In some embodiments, the Ides polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 186.

[0472] In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 95% identity with SEQ ID NO: 187. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 96% identity with SEQ ID NO: 187. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 97% identity with SEQ ID NO: 187. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 98% identity with SEQ ID NO: 187. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99% identity with SEQ ID NO: 187. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99.5% identity with SEQ ID NO: 187. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99.7% identity with SEQ ID NO:187. In some embodiments, the Ides polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 187.

[0473] In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 95% identity with SEQ ID NO: 177. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 96% identity with SEQ ID NO: 177. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 97% identity with SEQ ID NO: 177. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 98% identity with SEQ ID NO: 177. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99% identity with SEQ ID NO: 177. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having atAttorney Docket No. : CYR-009WO least 99.5% identity with SEQ ID NO: 177. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99.7% identity with SEQ ID NO: 177. In some embodiments, the Ides polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 177.

[0474] In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 95% identity with the amino acid sequence of any of SEQ ID NO: 154-161. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 96% identity with the amino acid sequence of any of SEQ ID NO: 154-161. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 97% identity with the amino acid sequence of any of SEQ ID NO: 154-161. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 98% identity with the amino acid sequence of any of SEQ ID NO: 154-161. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99% identity with the amino acid sequence of any of SEQ ID NO: 154-161. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99.5% identity with the amino acid sequence of any of SEQ ID NO: 154-161. In some embodiments, the Ides polypeptide conjugate comprises an amino acid sequence having at least 99.7% identity with the amino acid sequence of any of SEQ ID NO: 154-161. In some embodiments, the Ides polypeptide conjugate comprises the amino acid sequence of one of SEQ ID NO: 154-161.

[0475] Described herein, in certain embodiments, are IdeS polypeptide conjugates comprising a) an IdeS variant protein or wild-type IdeS; and b) an albumin binding domain conjugated to the IdeS variant protein or the wild-type IdeS, wherein the albumin binding domain comprises an amino acid sequence having at least 90% sequence identity with SEQ ID NO: 175 or SEQ ID NO: 176. In some embodiments, the albumin binding domain comprises an amino acid sequence having at least 95% sequence identity with SEQ ID NO: 175 or SEQ ID NO: 176. In some embodiments, the albumin binding domain comprises an amino acid sequence having at least 98% sequence identity with SEQ ID NO: 175 or SEQ ID NO: 176. In some embodiments, the albumin binding domain comprises an amino acid sequence having at least 99% sequence identity with SEQ ID NO: 175 or SEQ ID NO: 176. In some embodiments, the albumin binding domain comprises the amino acid sequence of SEQ ID NO: 175. In other embodiments, the albumin binding domain comprises the amino acidAttorney Docket No. : CYR-009WO sequence of SEQ ID NO: 176. In some embodiments, the albumin binding domain is conjugated to an N-terminus of the wild-type IdeS or IdeS variant protein. In some embodiments, the albumin binding domain is conjugated to a C-terminus of the wild-type IdeS or IdeS variant protein. The IdeS variant protein may be connected to the albumin binding domain by a peptide linker disclosed herein.

[0476] In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table 1 A and the albumin binding domain comprises the amino acid sequence of SEQ ID NO: 175. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table IB and the albumin binding domain comprises the amino acid sequence of SEQ ID NO: 175. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein is selected from any of the variants disclosed in Table 1 A and the albumin binding domain comprises the amino acid sequence of SEQ ID NO: 176. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein is selected from any of the variants disclosed in Table IB and the albumin binding domain comprises the amino acid sequence of SEQ ID NO: 176. The IdeS variant protein may be connected to the albumin binding domain by a peptide linker disclosed herein.

[0477] In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table 1 A and the albumin binding domain comprises at least 90% sequence identity, e.g. at least 95% sequence identity, with the amino acid sequence of SEQ ID NO: 175. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein selected from any of the variants disclosed in Table IB and the albumin binding domain comprises at least 90% sequence identity, e.g. at least 95% sequence identity, with the amino acid sequence of SEQ ID NO: 175. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein is selected from any of the variants disclosed in Table 1 A and the albumin binding domain comprises at least 90% sequence identity, e.g. at least 95% sequence identity, with the amino acid sequence of SEQ ID NO: 176. In some embodiments, the Ides polypeptide conjugate comprises an IdeS variant protein is selected from any of the variants disclosed in Table IB and the albumin binding domain comprises at least 90% sequence identity, e.g. at least 95% sequence identity, with the amino acid sequence of SEQ ID NO: 176. The IdeS variant protein may be connected to the albumin binding domain by a peptide linker disclosed herein.Attorney Docket No. : CYR-009WO

[0478] In some embodiments, provided herein are IdeS polypeptide conjugates comprising an IdeS variant protein having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90% at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5% or 100% sequence identity to any one of the polypeptide sequences listed in Table 1C.

[0479] In some embodiments, the IdeS polypeptide conjugate comprises one of the polypeptide sequences listed in Table 1C. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 154. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 155. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 156. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 157. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 158. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 159. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 160. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 161. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 177. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 178. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 179. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 180. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 181. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 182. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 183. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 184. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 185. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 186. For example, the IdeS polypeptide conjugate comprises the amino acid sequence of SEQ ID NO: 187.

[0480] In one embodiment, the IdeS polypeptide conjugate comprises the amino acid sequence of any one of SEQ ID NOS: 154-161 or 177, except that in the human serum albumin moiety, a free cysteine at a position corresponding to position 34 of SEQ ID NO: 188Attorney Docket No. : CYR-009WO is substituted with an amino acid selected from serine, valine, or alanine. For example, the substitution is cysteine to serine. For example, the substitution is cysteine to alanine. For example, in one embodiment, the IdeS polypeptide conjugate comprises the amino acid sequence of any one of SEQ ID NOS: 154-161 or 177, except that the human serum albumin moiety has been modified to correspond with the amino acid sequence of SEQ ID NO: 189. For example, in one embodiment, the IdeS polypeptide conjugate comprises the amino acid sequence of any one of SEQ ID NOS: 154-161 or 177, except that the human serum albumin moiety has been modified to correspond with the amino acid sequence of SEQ ID NO: 190.Table 1C. IdeS-HSA, IdeS-RSA, and IdeS-albumin binding domain (ABD) polypeptidesAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WOAttorney Docket No. : CYR-009WO* IdeS variant - linker - HSA or IdeS variant - linker - albumin binding domain (ABD)

[0481] In some embodiments, the IdeS protein or variant IdeS protein is conjugated to the human serum albumin using a linker. In some embodiments, the linker is a peptide linker. In some embodiments, the peptide linker comprises a flexible peptide linker. In some embodiments, the peptide linker comprises a rigid peptide linker. In some embodiments, the peptide linker comprises a cleavable peptide linker.

[0482] In some embodiments, the linker comprises at least 2 to about 30 amino acids. In some embodiments, the linker comprises about 2 to about 35 amino acids, about 2 to about 40 amino acids, about 2 to about 35 amino acids, about 2 to about 35 amino acids, about 2 to about 30 amino acids, about 2 to about 25 amino acids, about 2 to about 20 amino acids, about 2 to about 15 amino acids, about 2 to about 10 amino acids, about 5 to about 30 amino acids, about 5 to about 30 amino acids, about 5 to about 25 amino acids, about 5 to about 20 amino acids, about 5 to about 15 amino acids, or about 5 to about 10 amino acids.

[0483] In some embodiments, the linker comprises a sequence selected from the group consisting of (GS)n (SEQ ID NO:162), (G2S)n (SEQ ID NO:163), (G3S)n (SEQ ID NO: 164), (G4S)n (SEQ ID NO: 165), and (G)n (SEQ ID NO: 166), and wherein n is an integer from 1 to 20. In some embodiments, n is an integer from 1 to 18, from 1 to 16, from 1 to 14, from 1 to 12, from 1 to 10, from 1 to 8, from 1 to 6, from 1 to 4, from 4 to 20, from 6 to 20, from 8 to 20, from 10 to 20, from 12 to 20, from 14 to 20, from 16 to 20, or from 18 to 20.

[0484] In some embodiments, the linker comprises a sequence selected from the group consisting of (GGSGGD)n (SEQ ID NO: 167) or (GGSGGE)n (SEQ ID NO: 168), and wherein n is an integer from 1 to 6.Attorney Docket No. : CYR-009WO

[0485] In some embodiments, the linker comprises a sequence selected from the group consisting of (GGGSGSGGGGS)n (SEQ ID NO: 169) and (GGGGGPGGGGP)n (SEQ ID NO: 170), and wherein n is an integer from 1 to 3.

[0486] In some embodiments, the linker comprises a sequence selected from the group consisting of (GX)n, (GGX)n, (GGGX)n, (GGGGX)n (SEQ ID NO: 171), and (GzX)n, wherein z is between 1 and 20. In some embodiments, z is between 1 and 18, 1 and 16, 1 and 14, 1 and 12, 1 and 10, 1 and 8, 1 and 6, 1 and 4, 4 and 20, 6 and 20, 8 and 20, 10 and 20, 12 and 20, 14 and 20, 16 and 20, or 18 and 20. In some embodiments, X is serine, aspartic acid, glutamic acid, threonine, or proline.

[0487] In some embodiments, the linker comprises GSGGGSG (SEQ ID NO: 113), GSGSGSGS (SEQ ID NO: 114), GSGGGSGGGSG (SEQ ID NO: 115), or GSGSGSGSGSGS (SEQ ID NO: 116). In some embodiments, the linker comprises GS.

[0488] In some embodiments, the IdeS protein or variant IdeS protein is fused to the human serum albumin or variant human serum albumin.

[0489] In some embodiments, the IdeS variant proteins or IdeS polypeptide conjugates described herein comprise increased levels of expression when expressed by a cell as compared to the level of expression of wild-type IdeS by the same cell. In some embodiments the IdeS variant protein or IdeS polypeptide conjugate comprises an increase of at least or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 125%, 150%, 200%, 250%, 300%, 350%, 400%, 450%, 500% or more than 500% in the level of expression as compared to the level of expression of wildtype IdeS by the same cell. In some embodiments, the IdeS variant protein or IdeS polypeptide conjugate comprises an increase in a range of about 5-95%, 10-90%, 20-80%, 30-70%, 40-60%, 50-95%, 65-85%, 75-95%, 10-200%, 20-200%, 30-200%, 40-200%, 50- 200%, 75-200%, 100-200%, 150-200%, 10-300%, 20-300%, 30-300%, 40-300%, 50-300%, 75-300%, 100-300%, 150-300%, 10-400%, 20-400%, 30-400%, 40-400%, 50-400%, 75- 400%, 100-400%, 150-400%, 10-500%, 20-500%, 30-500%, 40-500%, 50-500%, 75-500%, 100-500%, or 150-500% in the level of expression when expressed by a cell as compared to the level of expression of wild-type IdeS.

[0490] In some embodiments, the IdeS variant protein or IdeS polypeptide conjugate described herein comprises levels of expression in a cell ± about 1-20%, ± about 2-20%, ±Attorney Docket No. : CYR-009WO about 4-20%, ± about 6-20%, ± about 8-20%, ± about 10-20%, ± about 12-20%, ± about 14- 20%, ± about 16-20%, ± about 18-20%, about 1-18%, ± about 2-18%, ± about 4-18%, ± about 6-18%, ± about 8-18%, ± about 10-18%, ± about 12-18%, ± about 14-18%, ± about 16- 18%, 1-16%, ± about 2-16%, ± about 4-16%, ± about 6-16%, ± about 8-16%, ± about 10- 16%, ± about 12-16%, ± about 14-16%, about 1-14%, ± about 2-14%, ± about 4-14%, ± about 6-14%, ± about 8-14%, ± about 10-14%, ± about 12-14%, ± about 1-12%, ± about 2- 12%, ± about 4-12%, ± about 6-12%, ± about 8-12%, ± about 10-12%, ± about 1-10%, ± about 2-10%, ± about 4-10%, ± about 6-10%, ± about 8-10%, ± about 1-8%, ± about 2-8%, ± about 4-8%, ± about 6-8%, ± about 1-6%, ± about 2-6%, ± about 4-6%, ± about 1-4%, ± about 2-4%, or about 1-2% the levels of expression of wild-type IdeS in the same cell.

[0491] In some embodiments, the IdeS variant protein or IdeS polypeptide conjugate described herein comprises increased stability of the IdeS variant protein as compared to the stability of wild-type IdeS. In some embodiments, the IdeS variant protein or IdeS polypeptide conjugate comprises an increase of at least or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 125%, 150%, 200%, 250%, 300%, 350%, 400%, 450%, 500% or more than 500% in the stability as compared to the stability of wild-type IdeS. In some embodiments, the IdeS variant protein or IdeS polypeptide conjugate comprises an increase in a range of about 5- 95%, 10-90%, 20-80%, 30-70%, 40-60%, 50-95%, 65-85%, 75-95%, 10-200%, 20-200%, 30-200%, 40-200%, 50-200%, 75-200%, 100-200%, 150-200%, 10-300%, 20-300%, 30- 300%, 40-300%, 50-300%, 75-300%, 100-300%, 150-300%, 10-400%, 20-400%, 30-400%, 40-400%, 50-400%, 75-400%, 100-400%, 150-400%, 10-500%, 20-500%, 30-500%, 40- 500%, 50-500%, 75-500%, 100-500%, or 150-500% increase in the stability as compared to the stability of wild-type IdeS.

[0492] In some embodiments, the IdeS variant protein or IdeS polypeptide conjugate described herein comprises an increased enzymatic activity as compared to the activity of wild-type IdeS. In some embodiments, the enzymatic activity is cleavage (e.g., proteolytic) activity of immunoglobulins. In some embodiments, the IdeS variant protein or IdeS polypeptide conjugate comprises an increase of at least or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 125%, 150%, 200%, 250%, 300%, 350%, 400%, 450%, 500% or more than 500% in the enzymatic activity as compared to the activity of wild-type IdeS. In some embodiments, theAttorney Docket No. : CYR-009WOIdeS variant protein or IdeS polypeptide conjugate comprises an increase in a range of about 5-95%, 10-90%, 20-80%, 30-70%, 40-60%, 50-95%, 65-85%, 75-95%, 10-200%, 20-200%, 30-200%, 40-200%, 50-200%, 75-200%, 100-200%, 150-200%, 10-300%, 20-300%, 30- 300%, 40-300%, 50-300%, 75-300%, 100-300%, 150-300%, 10-400%, 20-400%, 30-400%, 40-400%, 50-400%, 75-400%, 100-400%, 150-400%, 10-500%, 20-500%, 30-500%, 40- 500%, 50-500%, 75-500%, 100-500%, or 150-500% in the enzymatic activity as compared to the activity to wild-type IdeS.

[0493] In some embodiments, the IdeS variant protein or IdeS polypeptide conjugate described herein comprises decreased presentation of epitopes on human leukocyte antigen (HLA) as compared to a wild-type IdeS protein. In some embodiments, the IdeS variant protein or IdeS polypeptide conjugate comprises a decrease of a...

Claims

Attorney Docket No.: CYR-009WOCLAIMSWHAT IS CLAIMED IS:

1. An IdeS variant protein comprising a modification at a position corresponding to position 189 of SEQ ID NO: 1.

2. The IdeS variant protein of claim 1, wherein the a modification is a substitution of threonine to glycine.

3. The IdeS variant protein of claim 1 or claim 2, wherein the IdeS variant protein comprises at least 90% sequence identity to SEQ ID NO:2.

4. The IdeS variant protein of claim 1 or claim 2, wherein the IdeS variant protein comprises at least 95% sequence identity to SEQ ID NO:2.

5. The IdeS variant protein of claim 1 or claim 2, wherein the IdeS variant protein comprises at least 98% sequence identity to SEQ ID NO:2.

6. The IdeS variant protein of any one of claims 1-5, further comprising a modification at a position corresponding to position 132 of SEQ ID NO: 1.

7. The IdeS variant protein of claim 6, wherein the modification is a substitution of glutamate to glutamine.

8. The IdeS variant protein of any one of claims 1-8, further comprising one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 166, 187, 213, 236, 277, 302, 303, 306, and 318.

9. The IdeS variant protein of claim 8, wherein the variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1, a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1, a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1, a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1, a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1, a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to positionAttorney Docket No.: CYR-009WO10. The IdeS variant protein of any one of claims 1-7, further comprising one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 111, 113, 166, 187, 198, 213, 302, and 303.

11. The IdeS variant protein of claim 10, wherein the variant protein comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1; a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1; a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ IDNO: 1; a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1; a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1; a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1; a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1; a substitution of threonine to glycine at a position corresponding to position 189 of SEQ ID NO: 1; a substitution of glutamate to asparagine at a position corresponding to position 198 of SEQ ID NO: 1; a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1; a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

12. The IdeS variant protein of any one of claims 1-11, wherein the variant protein further comprises a substitution of the amino acid residues DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with an amino acid sequence selected from the group consisting of:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).

13. The IdeS variant protein of any one of claims 1-12, wherein the variant protein further comprises a substitution of the amino acid residues DSFSANQEIRYSEVTPYHVTAttorney Docket No.: CYR-009WO (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with the amino acid sequence QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO: 85).

14. The IdeS variant protein of claim 12 or 13, wherein the IdeS variant protein comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2.

15. The IdeS variant protein of claim 12 or 13, wherein the IdeS variant protein comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2.

16. The IdeS variant protein of claim 12 or 13, wherein the IdeS variant protein comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

17. An IdeS variant protein comprising an amino acid sequence comprising at least 90% sequence the amino acid sequence according to SEQ ID NO: 172 or SEQ ID NO: 173.

18. The IdeS variant protein of claim 17, wherein the IdeS variant protein comprises at least 95% identity with the amino acid sequence according to SEQ ID NO: 172 or SEQ ID NO: 173.

19. The IdeS variant protein of claim 17, wherein the IdeS variant protein comprises at least 99% identity with the amino acid sequence according to SEQ ID NO: 172 or SEQ ID NO: 173.

20. The IdeS variant protein of claim 17, wherein the IdeS variant comprises the amino acid sequence of SEQ ID NO: 172.

21. The IdeS variant protein of claim 17, wherein the IdeS variant comprises the amino acid sequence of SEQ ID NO: 173.

22. An IdeS polypeptide conjugate comprising an IdeS variant of any one of claims 1-18 conjugated to a human serum albumin or a variant human serum albumin.

23. An IdeS polypeptide conjugate comprising an IdeS variant protein comprising at least 90% sequence identity to SEQ ID NO: 172 or SEQ ID NO: 173 conjugated to a human serum albumin or a variant human serum albumin.

24. The IdeS polypeptide conjugate of claim 22 or 23, wherein the human serum albumin or variant human serum albumin is conjugated to an N-terminal of the IdeS variant protein.Attorney Docket No.: CYR-009WO25. The Ides polypeptide conjugate of claim 22 or 23 wherein the human serum albumin or variant human serum albumin is conjugated to a C-terminal of the IdeS variant protein.

26. The Ides polypeptide conjugate of any one of claims 22-25, wherein the IdeS variant protein is conjugated to the human serum albumin or variant human serum albumin using a linker.

27. The IdeS polypeptide conjugate of claim 26, wherein the linker comprises GS, GSGGGSG (SEQ ID NO: 113), GSGSGSGS (SEQ ID NO: 114), GSGGGSGGGSG (SEQ ID NO: 115), or GSGSGSGSGSGS (SEQ ID NO: 116).

28. The IdeS polypeptide conjugate of any one of claims 22-27, wherein the IdeS variant protein is fused to the human serum albumin or the variant human serum albumin.

29. The IdeS polypeptide conjugate of any one of claims 22-28, wherein the human serum albumin or variant human serum albumin is selected from i) the amino acid sequence of SEQ ID NO: 188, ii) the amino acid sequence of SEQ ID NO: 189; or iii) the amino acid sequence of SEQ ID NO: 190.

30. The IdeS polypeptide conjugate of any one of claims 22-29, wherein the variant human serum albumin comprises an amino acid sequence having at least 99% sequence identity with SEQ ID NO: 188.

31. The IdeS polypeptide conjugate of any one of claims 22-30, wherein the IdeS variant protein comprises at least 95% sequence identity to SEQ ID NO: 172 or SEQ ID NO: 173.

32. An IdeS polypeptide conjugate comprising the amino acid sequence of any one of SEQ ID NOS: 154-161, 177, 186, or 18733. The IdeS polypeptide conjugate of claim 32, wherein the polypeptide comprises the amino acid sequence of SEQ ID NO: 177.

34. The IdeS polypeptide conjugate of claim 32, wherein the polypeptide comprises the amino acid sequence of SEQ ID NO: 186.

35. The IdeS polypeptide conjugate of claim 32, wherein the polypeptide comprises the amino acid sequence of SEQ ID NO: 187.Attorney Docket No.: CYR-009WO36. An IdeS polypeptide conjugate comprising an IdeS variant of any one of claims 1-21 conjugated to an albumin binding domain.

37. The IdeS polypeptide conjugate of claim 36, wherein the albumin binding domain comprises an amino acid sequence with at least 90% sequence identity to SEQ ID NO: 175 or 176.

38. The IdeS polypeptide conjugate of claim 36, wherein the albumin binding domain comprises the amino acid sequence of SEQ ID NO: 175 or 176.

39. The IdeS polypeptide conjugate of any one of claims 36-38, wherein the albumin binding domain is conjugated to an N-terminus of the IdeS variant protein.

40. The IdeS polypeptide conjugate of any one of claims 36-38, wherein the albumin binding domain is conjugated to a C-terminus of the IdeS variant protein.

41. The Ides polypeptide conjugate of any one of claims 36-40, wherein the IdeS variant protein is conjugated to the albumin binding domain with a linker selected from GS, GSGGGSG (SEQ ID NO: 113), GSGSGSGS (SEQ ID NO: 114), GSGGGSGGGSG (SEQ ID NO: 115), or GSGSGSGSGSGS (SEQ ID NO: 116).

42. The Ides polypeptide conjugate of claim 36, wherein the conjugate comprises the amino acid sequence of any one of SEQ ID NOS: 178-184.

43. The Ides polypeptide conjugate of claim 36, wherein the conjugate comprises the ab amino acid sequence comprising at least 90% sequence identity with the amino acid sequence of any one of SEQ ID NOS: 178-184.

44. A polypeptide comprising: a) an IdeS variant protein or wild-type IdeS; and b) an albumin binding domain conjugated to the IdeS variant protein or the wild-type IdeS, wherein the albumin binding domain comprises an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 175 or SEQ ID NO: 176.

45. A polypeptide comprising: a) an IdeS variant protein; and b) a human serum albumin or a human serum albumin variant.Attorney Docket No.: CYR-009WO46. The polypeptide of claim 44 or 45, wherein the IdeS variant protein comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 113, 115, 116, 127, 128, 129, 130, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187, 188, 195, 197, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258, 273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.

47. The polypeptide of claim 44 or 45, wherein the IdeS variant protein comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 31, 32, 33, 38, 39, 43, 44, 45, 47, 54, 57, 60, 68, 74, 75, 77, 82, 85, 111, 113, 115, 116, 127, 128, 129, 130, 132, 133, 148, 153, 154, 159, 166, 167, 168, 175, 187, 188, 189, 195, 197, 198, 210, 213, 218, 219, 220, 228, 233, 236, 241, 244, 245, 247, 258, 273, 274, 277, 278, 288, 289, 297, 299, 300, 302, 303, 306, 307, 308, 313, 314, 315, 316, 318, 322, and 330.

48. The polypeptide of any one of claims 44-47, wherein the IdeS variant protein comprises one or more modifications at a position or positions corresponding to a position or positions in SEQ ID NO: 1 selected from the group consisting of: 68, 75, 111, 113, 132, 166, 187, 189, 198, 213, 302, and 303.

49. The polypeptide of any one of claims 44-48, wherein: i) the modification corresponding to position 68 of SEQ ID NO: 1 is a substitution of valine to threonine; ii) the modification corresponding to position 75 of SEQ ID NO: 1 is a substitution of alanine to proline; iii) the modification corresponding to position 166 of SEQ ID NO: 1 is a substitution of threonine to arginine or a substitution of threonine to glycine; iv) the modification corresponding to position 187 of SEQ ID NO: 1 is a substitution of serine to aspartate or a substitution of serine to glutamate; v) the modification corresponding to position 213 of SEQ ID NO: 1 is a substitution of threonine to glutamate; vi) the modification corresponding to position 236 of SEQ ID NO: 1 is a substitution of serine to cysteine; vii)the modification corresponding to position 277 of SEQ ID NO: 1 is a substitution of leucine to cysteine;Attorney Docket No.: CYR-009WO viii) the modification corresponding to position 302 of SEQ ID NO: 1 is a substitution of serine to aspartate, a substitution of serine to lysine, or a substitution of serine to glutamate; ix) the modification corresponding to position 303 of SEQ ID NO:1 is a substitution of alanine to aspartate, a substitution of alanine to asparagine, or a substitution of alanine to glutamine; x) the modification corresponding to position 306 of SEQ ID NO: 1 is a substitution of valine to threonine. xi) the modification corresponding to position 318 of SEQ ID NO: 1 is a substitution of isoleucine to lysine, a substitution of isoleucine to aspartate, or a substitution of isoleucine to glycine; xii)the modification corresponding to position 308 of SEQ ID NO:1 is a substitution of isoleucine to leucine; xiii) the modification corresponding to position 31 of SEQ ID NO: 1 is a substitution of serine to aspartate or asparagine; xiv) the modification corresponding to position 32 of SEQ ID NO: 1 is a substitution of phenylalanine to lysine; xv)the modification corresponding to position 33 of SEQ ID NO:1 is a substitution of serine to glutamate; xvi) the modification corresponding to position 38 of SEQ ID NO: 1 is a substitution of isoleucine to valine; xvii) the modification corresponding to position 39 of SEQ ID NO: 1 is a substitution of arginine to asparagine or threonine; xviii) the modification corresponding to position 43 of SEQ ID NO: 1 is a substitution of valine to glutamate; xix) the modification corresponding to position 44 of SEQ ID NO: 1 is a substitution of threonine to glutamate; xx)the modification corresponding to position 45 of SEQ ID NO:1 is a substitution of proline to glutamate; xxi) the modification corresponding to position 47 of SEQ ID NO: 1 is a substitution of histidine to lysine; xxii) the modification corresponding to position 54 of SEQ ID NO: 1 is a substitution of lysine to aspartate; xxiii) the modification corresponding to position 57 of SEQ ID NO: 1 is a substitution of threonine to lysine or glutamine;Attorney Docket No.: CYR-009WO xxiv) the modification corresponding to position 60 of SEQ ID NO: 1 is a substitution of alanine to aspartate or glutamine; xxv) the modification corresponding to position 74 of SEQ ID NO: 1 is a substitution of valine to lysine; xxvi) the modification corresponding to position 75 of SEQ ID NO: 1 is a substitution of alanine to asparagine or proline; xxvii) the modification corresponding to position 77 of SEQ ID NO: 1 is a substitution of glutamine to glycine; xxviii) the modification corresponding to position 82 of SEQ ID NO: 1 is a substitution of isoleucine to methionine; xxix) the modification corresponding to position 85 of SEQ ID NO: 1 is a substitution of threonine to glutamine; xxx) the modification corresponding to position 111 of SEQ ID NO: 1 is a substitution of lysine to asparagine; xxxi) the modification corresponding to position 113 of SEQ ID NO: 1 is a substitution of glutamine to aspartate; xxxii) the modification corresponding to position 115 of SEQ ID NO: 1 is a substitution of lysine to histidine; xxxiii) the modification corresponding to position 116 of SEQ ID NO: 1 is a substitution of arginine to aspartate; xxxiv) the modification corresponding to position 127 of SEQ ID NO: 1 is a substitution of isoleucine to methionine; xxxv) the modification corresponding to position 128 of SEQ ID NO: 1 is a substitution of asparagine to glycine; xxxvi) the modification corresponding to position 129 of SEQ ID NO: 1 is a substitution of phenylalanine to glutamate; xxxvii) the modification corresponding to position 130 of SEQ ID NO: 1 is a substitution of asparagine to glutamate; xxxviii) the modification corresponding to position 132 of SEQ ID NO: 1 is a substitution of glutamate for glutamine; xxxix) the modification corresponding to position 133 of SEQ ID NO: 1 is a substitution of glutamine to aspartate; xl) the modification corresponding to position 148 of SEQ ID NO: 1 is a substitution of leucine to asparagine or arginine;Attorney Docket No.: CYR-009WO xli)the modification corresponding to position 153 of SEQ ID NO: 1 is a substitution of phenylalanine to methionine or tyrosine; xlii) the modification corresponding to position 154 of SEQ ID NO: 1 is a substitution of glutamate to aspartate; xliii) the modification corresponding to position 159 of SEQ ID NO: 1 is a substitution of lysine to asparagine; xliv) the modification corresponding to position 167 of SEQ ID NO: 1 is a substitution of lysine to proline; xlv) the modification corresponding to position 168 of SEQ ID NO: 1 is a substitution of histidine to aspartate or proline; xlvi) the modification corresponding to position 175 of SEQ ID NO: 1 is a substitution of histidine to glutamine; xlvii) the modification corresponding to position 188 of SEQ ID NO: 1 is a substitution of leucine to glycine or methionine; xlviii) the modification corresponding to position 189 of SEQ ID NO: 1 is a substitution of threonine for glycine xlix) the modification corresponding to position 195 of SEQ ID NO: 1 is a substitution of proline to aspartate;1) the modification corresponding to position 198 of SEQ ID NO: 1 is a substitution of glutamate to asparagine; li) the modification corresponding to position 197 of SEQ ID NO: 1 is a substitution of lysine to glutamate; lii) the modification corresponding to position 210 of SEQ ID NO: 1 is a substitution of alanine to glycine; liii)the modification corresponding to position 218 of SEQ ID NO: 1 is a substitution of serine to aspartate; liv)the modification corresponding to position 219 of SEQ ID NO: 1 is a substitution of lysine to glycine;Iv) the modification corresponding to position 220 of SEQ ID NO: 1 is a substitution of leucine to glutamine; lvi)the modification corresponding to position 228 of SEQ ID NO: 1 is a substitution of lysine to glutamine;Ivii) the modification corresponding to position 233 of SEQ ID NO: 1 is a substitution of lysine to serine;Attorney Docket No.: CYR-009WOIviii) the modification corresponding to position 241 of SEQ ID NO: 1 is a substitution of lysine to aspartate; lix)the modification corresponding to position 244 of SEQ ID NO: 1 is a substitution of threonine to lysine; lx) the modification corresponding to position 245 of SEQ ID NO: 1 is a substitution of glutamate to aspartate; lxi)the modification corresponding to position 247 of SEQ ID NO: 1 is a substitution of lysine to asparagine;Ixii) the modification corresponding to position 258 of SEQ ID NO: 1 is a substitution of valine to aspartate;Ixiii) the modification corresponding to position 273 of SEQ ID NO: 1 is a substitution of serine to aspartate;Ixiv) the modification corresponding to position 274 of SEQ ID NO: 1 is a substitution of asparagine to glutamate;Ixv) the modification corresponding to position 278 of SEQ ID NO: 1 is a substitution of lysine to aspartate or threonine;Ixvi) the modification corresponding to position 288 of SEQ ID NO: 1 is a substitution of asparagine to aspartate, glycine, or glutamine;Ixvii) the modification corresponding to position 289 of SEQ ID NO: 1 is a substitution of alanine to proline;Ixviii) the modification corresponding to position 297 of SEQ ID NO: 1 is a substitution of phenylalanine to aspartate;Ixix) the modification corresponding to position 299 of SEQ ID NO: 1 is a substitution of glycine to aspartate;Ixx) the modification corresponding to position 300 of SEQ ID NO: 1 is a substitution of valine to glutamate;Ixxi) the modification corresponding to position 307 of SEQ ID NO: 1 is a substitution of alanine to glycine or asparagine;Ixxii) the modification corresponding to position 313 of SEQ ID NO: 1 is a substitution of isoleucine to proline;Ixxiii) the modification corresponding to position 314 of SEQ ID NO: 1 is a substitution of lysine to glycine;Ixxiv) the modification corresponding to position 315 of SEQ ID NO: 1 is a substitution of glutamate to proline;Attorney Docket No.: CYR-009WO Ixxv) the modification corresponding to position 316 of SEQ ID NO: 1 is a substitution of aspartate to proline;Ixxvi) the modification corresponding to position 322 of SEQ ID NO: 1 is a substitution of valine to threonine; and / orIxxvii) the modification corresponding to position 330 of SEQ ID NO: 1 is a substitution of threonine to aspartate.

50. The polypeptide of any one of claims 44-49, wherein the IdeS variant protein comprises: i) a substitution of valine to threonine at a position corresponding to position 68, a substitution of alanine to proline at a position corresponding to position 75, a substitution of threonine corresponding to arginine at a position to corresponding to position 166, a substitution of serine to aspartate at a position corresponding to position 187, a substitution of threonine to glutamate at a position corresponding to position 213, a substitution of serine to lysine at a position corresponding to position 302, and a substitution of alanine to aspartate at a position corresponding to position 303; ii) a substitution of valine to threonine at a position corresponding to position 68, a substitution of alanine to proline at a position corresponding to position 75, a substitution of threonine to arginine at a position corresponding to position 166, a substitution of serine to aspartate at a position corresponding to position 187, a substitution of threonine to glutamate at a position corresponding to position 213, a substitution of serine to glutamate at a position corresponding to position 302, and a substitution of alanine to aspartate at a position corresponding to position 303; iii) a substitution of valine to threonine at a position corresponding to position 68, a substitution of alanine to proline at a position corresponding to position 75, a substitution of threonine to arginine at a position corresponding to position 166, a substitution of serine to aspartate at a position corresponding to position 187, a substitution of threonine to glutamate at a position corresponding to position 213, a substitution of serine to lysine at a position corresponding to position 302, a substitution of alanine to aspartate at a position corresponding to position 303, and a substitution of valine to threonine at a position corresponding to position 306; iv)a substitution of valine to threonine at a position corresponding to position 68, a substitution of alanine to proline at a position corresponding to position 75, a substitution ofAttorney Docket No.: CYR-009WO threonine to arginine at a position corresponding to position 166, a substitution of serine to aspartate at a position corresponding to position 187, a substitution of threonine to glutamate at a position corresponding to position 213, a substitution of serine to glutamate at a position corresponding to position 302, a substitution of valine to threonine at a position corresponding to position 306, and a substitution of isoleucine to lysine at a position corresponding to position 318; v) a substitution of valine to threonine at a position corresponding to position 68, a substitution of alanine to proline at a position n corresponding to position 75, a substitution of threonine to arginine at a position corresponding to position 166, a substitution of serine to aspartate at a position corresponding to position 187, a substitution of threonine to glutamate at a position corresponding to position 213, a substitution of serine to cysteine at a position corresponding to position 236, a substitution of leucine to cysteine at a position corresponding to position 277, a substitution of serine to glutamate at a position corresponding to position 302, a substitution of alanine to aspartate at a position corresponding to position 303, and a substitution of valine to threonine at a position corresponding to position 306; or vi)a substitution of valine to threonine at a position corresponding to position 68, a substitution of alanine to proline at a position corresponding to position 75, a substitution of threonine to arginine at a position corresponding to position 166, a substitution of serine to aspartate at a position corresponding to position 187, a substitution of threonine to glutamate at a position corresponding to position 213, a substitution of serine to cysteine at a position corresponding to position 236, a substitution of leucine to cysteine at a position corresponding to position 277, and a substitution of isoleucine to leucine at a position corresponding to position 308.

51. The polypeptide of any one of claims 44-49, wherein the IdeS variant comprises a substitution of valine to threonine at a position corresponding to position 68 of SEQ ID NO: 1; a substitution of alanine to proline at a position corresponding to position 75 of SEQ ID NO: 1; a substitution of lysine to asparagine at a position corresponding to position 111 of SEQ ID NO: 1; a substitution of glutamine to threonine at a position corresponding to position 113 of SEQ ID NO: 1; a substitution of glutamate to glutamine at a position corresponding to position 132 of SEQ ID NO: 1; a substitution of threonine to arginine at a position corresponding to position 166 of SEQ ID NO: 1; a substitution of serine to aspartate at a position corresponding to position 187 of SEQ ID NO: 1; a substitution of threonine to glycine at a position corresponding to position 189 of SEQ ID NO: 1; a substitution of glutamate to asparagine at aAttorney Docket No.: CYR-009WO position corresponding to position 198 of SEQ ID NO: 1; a substitution of threonine to glutamate at a position corresponding to position 213 of SEQ ID NO: 1; a substitution of serine to lysine at a position corresponding to position 302 of SEQ ID NO: 1, and a substitution of alanine to aspartate at a position corresponding to position 303 of SEQ ID NO: 1.

52. The polypeptide of any one of claims 44-51, further comprising one or more glycosylation modifications as compared to wild-type.

53. The polypeptide of claim 52, wherein the glycosylation modification is at a position corresponding to a position in SEQ ID NO: 1 selected from the group consisting of: 31, 37, 39, 42, 74, 76, 111, 113, 119, 121, 130, 142, 144, 147, 148, 198, 233, 244, 246, 311, 313, and 319.

54. The polypeptide of claim 53, wherein: i) the glycosylation modification occurs at a position corresponding to position 31 of SEQ ID NO: 1 where there is a substitution of serine to asparagine; ii) the glycosylation modification occurs at a position corresponding to position 37 of SEQ ID NO: 1 where there is a substitution of glutamate to asparagine; iii) the glycosylation modification occurs at a position corresponding to position 39 of SEQ ID NO: 1 where there is a substitution of arginine to threonine or asparagine; iv) the glycosylation modification occurs at a position corresponding to position 42 of SEQ ID NO: 1 where there is a substitution of glutamate to asparagine; v) the glycosylation modification occurs at a position corresponding to position 74 of SEQ ID NO: 1 where there is a substitution of valine to asparagine; vi) the glycosylation modification occurs at a position corresponding to position 76 of SEQ ID NO: 1 where there is a substitution of asparagine to serine; vii) the glycosylation modification occurs at a position corresponding to position 111 of SEQ ID NO: 1 where there is a substitution of lysine to asparagine; viii) the glycosylation modification occurs at a position corresponding to position 113 of SEQ ID NO: 1 where there is a substitution of glutamine to threonine; ix) the glycosylation modification occurs at a position corresponding to position 119 of SEQ ID NO: 1 where there is a substitution of glutamate to asparagine; x) the glycosylation modification occurs at a position corresponding to position 121 of SEQ ID NO: 1 where there is a substitution of histidine to threonine;Attorney Docket No.: CYR-009WO xi) the glycosylation modification occurs at a position corresponding to position 130 of SEQ ID NO: 1 where there is a substitution of asparagine to serine; xii) the glycosylation modification occurs at a position corresponding to position 142 of SEQ ID NO: 1 where there is a substitution of aspartate to asparagine; xiii) the glycosylation modification occurs at a position corresponding to position 144 of SEQ ID NO: 1 where there is a substitution of lysine to threonine; xiv) the glycosylation modification occurs at a position corresponding to position 147 of SEQ ID NO: 1 where there is a substitution of glutamine to serine; xv) the glycosylation modification occurs at a position corresponding to position 148 of SEQ ID NO: 1 where there is a substitution of leucine to asparagine; xvi) the glycosylation modification occurs at a position corresponding to position 198 of SEQ ID NO: 1 where there is a substitution of glutamate to asparagine; xvii) the glycosylation modification occurs at a position corresponding to position 233 of SEQ ID NO: 1 where there is a substitution of lysine to serine; xviii) the glycosylation modification occurs at a position corresponding to position 244 of SEQ ID NO: 1 where there is a substitution of threonine to asparagine; xix) the glycosylation modification occurs at a position corresponding to position 246 of SEQ ID NO: 1 where there is a substitution of glycine to threonine; xx) the glycosylation modification occurs at a position corresponding to position 311 of SEQ ID NO: 1 where there is a substitution of lysine to asparagine; xxi) the glycosylation modification occurs at a position corresponding to position 313 of SEQ ID NO: 1 where there is a substitution of isoleucine to threonine; and / or xxii) the glycosylation modification occurs at a position corresponding to position 319 of SEQ ID NO: 1 where there is a substitution of glycine to serine.

55. The polypeptide of any one of claims 44-54, wherein the IdeS variant protein comprises: i) a substitution of glutamate at position 37 to asparagine and a substitution of arginine at position 39 to threonine; ii) a substitution of valine at position 74 to asparagine and a substitution of asparagine at position 76 to serine;Attorney Docket No.: CYR-009WO iii) a substitution of lysine at position 111 to asparagine and a substitution of glutamine at position 113 to threonine; iv) a substitution of glutamate at position 119 to asparagine and a substitution of histidine at position 121 to threonine; v) a substitution of aspartate at position 142 to asparagine and a substitution of lysine at position 144 to threonine; vi) a substitution of threonine at position 244 to asparagine and a substitution of glycine at position 246 to threonine; and / or vii) a substitution of lysine at position 311 to asparagine and a substitution of isoleucine at position 313 to threonine.

56. The polypeptide of any one of claims 44-55, further comprising at least one modification at a position corresponding to a position in SEQ ID NO: 12 selected from the group consisting of: 47, 49, 51, 78, 111, 113, 123, 125, 126, 128, 142, 144, 148, 198, 273, 275, 278, 312, and 314 to introduce a glycosylation site.

57. The polypeptide of claim 56, wherein: i) the modification at a position corresponding to position 47 is a substitution of threonine to asparagine; ii) the modification at a position corresponding to position 49 is a substitution of glutamine to threonine or asparagine; iii) the modification at a position corresponding to position 51 is a substitution of valine to threonine; iv) the modification at a position corresponding to position 78 is a substitution of glycine to serine; v) the modification at a position corresponding to position 111 is a substitution of lysine to asparagine; vi) the modification at a position corresponding to position 113 is a substitution of glutamine to threonine; vii) the modification at a position corresponding to position 123 is a substitution of glutamate to asparagine; viii) the modification at a position corresponding to position 125 is a substitution of glutamine to threonine;Attorney Docket No.: CYR-009WO ix) the modification at a position corresponding to position 126 is a substitution of lysine to asparagine; x) the modification at a position corresponding to position 128 is a substitution of asparagine to serine; xi) the modification at a position corresponding to position 142 is a substitution of aspartate to asparagine; xii) the modification at a position corresponding to position 144 is a substitution of lysine to threonine; xiii) the modification at a position corresponding to position 148 is a substitution of leucine to asparagine; xiv) the modification at a position corresponding to position 198 is a substitution of glutamate to asparagine; xv) the modification at a position corresponding to position 273 is a substitution of serine to asparagine; xvi) the modification at a position corresponding to position 275 is a substitution of glycine to serine; xvii) the modification at a position corresponding to position 278 is a substitution of lysine to serine; xviii) the modification at a position corresponding to position 312 is a substitution of glutamate to asparagine; or xix) the modification at a position corresponding to position 314 is a substitution of lysine to serine.

58. The polypeptide of any one of claims 44-57, wherein the variant protein comprises: i) a substitution of threonine to asparagine at a position corresponding to position 47 and a substitution of glutamine to threonine at a position corresponding to position 49; ii) a substitution of glutamine to asparagine at a position corresponding to position 49 and a substitution of valine to threonine at a position corresponding to position 51; iii) a substitution of glutamate to asparagine at a position corresponding to position 123 and a substitution of glutamine to threonine at a position corresponding to position 125;Attorney Docket No.: CYR-009WO iv)a substitution of lysine to asparagine at a position corresponding to position 126 and a substitution of asparagine to serine at a position corresponding to position 128; v) a substitution of serine to asparagine at a position corresponding to position 273 and a substitution of glycine to serine at a position corresponding to position 275; vi)a substitution of glutamate to asparagine at a position corresponding to position 312 and a substitution of lysine to serine at a position corresponding to position 314; vii) a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, a substitution of aspartate to asparagine at a position corresponding to position 142, and a substitution of lysine to threonine at a position corresponding to position 144; viii) a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, and a substitution of glutamate to asparagine at a position corresponding to position 198; ix)a substitution of aspartate to asparagine at a position corresponding to position 142, a substitution of lysine to threonine at a position corresponding to position 144, and a substitution of glutamate to asparagine at a position corresponding to position 198; x) a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, a substitution of aspartate to asparagine at a position corresponding to position 142, a substitution of lysine to threonine at a position corresponding to position 144, and a substitution of glutamate to asparagine at a position corresponding to position 198; xi)a substitution of glutamine to asparagine at a position corresponding to position 49, a substitution of valine to threonine at a position corresponding to position 51, a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, a substitution of lysine to asparagine at a position corresponding to position 126, a substitution of asparagine to serine at a position corresponding to position 128, a substitution of leucine to asparagine at a position corresponding to position 148, and a substitution of glutamate to asparagine at a position corresponding to position 198;Attorney Docket No.: CYR-009WO xii) a substitution of glutamine to asparagine at a position corresponding to position 49, a substitution of valine to threonine at a position corresponding to position 51, a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, a substitution of leucine to asparagine at a position corresponding to position 148, and a substitution of glutamate to asparagine at a position corresponding to position 198; xiii) a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, a substitution of lysine to asparagine at a position corresponding to position 126, a substitution of asparagine to serine at a position corresponding to position 128, a substitution of leucine to asparagine at a position corresponding to position 148, and a substitution of glutamate to asparagine at a position corresponding to position 198; xiv) a substitution of glutamine to asparagine at a position corresponding to position 49, a substitution of valine to threonine at a position corresponding to position 51, a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, a substitution of lysine to asparagine at a position corresponding to position 126, a substitution of asparagine to serine at a position corresponding to position 128, and a substitution of glutamate to asparagine at a position corresponding to position 198; xv) a substitution of threonine to asparagine at a position corresponding to position 47, a substitution of glutamine to asparagine at a position corresponding to position 49, a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, and a substitution of glutamate to asparagine at a position corresponding to position 198; xvi) a substitution of glutamine to asparagine at a position corresponding to position 49, a substitution of valine to threonine at a position corresponding to position 51, a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, and a substitution of glutamate to asparagine at a position corresponding to position 198; xvii) a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, a substitution of glutamate to asparagine at a position corresponding to position 123, aAttorney Docket No.: CYR-009WO substitution of glutamine to threonine at a position corresponding to position 125, and a substitution of glutamate at to asparagine position 198; xviii) a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine at to threonine position 113, a substitution of lysine to asparagine at a position corresponding to position 126, a substitution of asparagine to serine at a position corresponding to position 128, and a substitution of glutamate to asparagine at a position corresponding to position 198; or xix) a substitution of lysine to asparagine at a position corresponding to position 111, a substitution of glutamine to threonine at a position corresponding to position 113, a substitution of leucine to asparagine at a position corresponding to position 148, and a substitution of glutamate to asparagine at a position corresponding to position 198.

59. The polypeptide of any one of claims 44-58, wherein the IdeS variant protein comprises a modification at a position corresponding to position 189 of SEQ ID NO: 1.

60. The polypeptide of claim 59, wherein the modification is a substitution of threonine to a glycine at a position corresponding to position 189 of SEQ ID NO: 1.

61. The polypeptide of any one of claims 44-60, wherein the IdeS variant protein comprises a modification at a position corresponding to position 132 of SEQ ID NO: 1.

62. The polypeptide of claim 61, wherein the modification is a substitution of glutamate to glutamine at a position corresponding to a position 132 of SEQ ID NO: 1.

63. The polypeptide of any one of claims 44-62, wherein the IdeS variant protein further comprises a substitution of the amino acid sequence DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with a sequence selected from the group consisting of:(i) TTIQAETSKHTISKKDETLHQNQLSISKTAT (SEQ ID NO:77),(ii) DDYQRNAMEAYAKEVPHQIT (SEQ ID NO:78),(iii) DPNNENEVSNLEKIKKLYPKGFQYGN (SEQ ID NO: 79),(iv) DQKLKDYLKNDQLKGTELGKFLEEQGVTK (SEQ ID NO: 80),(v) SAKVETGLPGELAPEEFSFPN (SEQ ID NO:81),(vi) AQNKNPVTHYVNQFDGEEKEII (SEQ ID NO: 82),(vii) GSIGEKWDLLLDGIGLNSHRSS (SEQ ID NO:83),(viii) AEPITLIWENYLSDSVSPDRDIR (SEQ ID NO:84),Attorney Docket No.: CYR-009WO(ix) QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85); and(x) QEEIAEGRRNPLRTAEWPMTKSTNDT (SEQ ID NO: 174).

64. The polypeptide of claim 63, wherein the IdeS variant protein further comprises a substitution of the amino acid residues DSFSANQEIRYSEVTPYHVT (SEQ ID NO:76) corresponding to residues 1-20 of SEQ ID NO:2 with the amino acid sequence QEEIAEGRRNPLRTAEWPMTKSTTDQ (SEQ ID NO:85).

65. The polypeptide of claim 63 or 64, wherein the IdeS variant protein comprises at least 90% sequence identity with residues 21-310 of SEQ ID NO:2.

66. The polypeptide of claim 63 or 64, wherein the IdeS variant protein comprises at least 95% sequence identity with residues 21-310 of SEQ ID NO:2.

67. The polypeptide of claim 63 or 64, wherein the IdeS variant protein comprises at least 98% sequence identity with residues 21-310 of SEQ ID NO:2.

68. The polypeptide of claim 44 or 45, wherein the wild-type IdeS comprises the amino acid sequence according to SEQ ID NO:2.

69. The polypeptide of claim 44 or 45, wherein the IdeS variant protein comprises an amino acid sequence having at least 90% sequence identity with any one of SEQ ID NOs: 3-75, 86-94, 117-153 and 172-173.

70. The polypeptide of claim 44 or 45, wherein the IdeS variant comprises an amino acid sequence having at least 95% sequence identity with any one of SEQ ID NOs: 3-75, 86-94, 117-153, and 172-173.

71. The polypeptide of any one of claims 44 or 46-70, wherein the albumin binding domain comprises the amino acid sequence of SEQ ID NO: 175 or 176.

72. The polypeptide of any one of claims 45-70, wherein the human serum albumin comprises the amino acid sequence of SEQ ID NO: 188.

73. The polypeptide of any one of claims 45-70, wherein the variant human serum albumin comprises the amino acid sequence of SEQ ID NO: 189.

74. The polypeptide of any one of claims 45-70, wherein the variant human serum albumin comprises the amino acid sequence of SEQ ID NO: 190.Attorney Docket No.: CYR-009WO75. The polypeptide of any one of claims 45-70, wherein the variant human serum albumin comprises an amino acid sequence having at least 99% sequence identity with SEQ ID NO: 188.

76. An IdeS variant comprising the amino acid sequence of any one of SEQ ID NOS:3- 75, 86-94, 117-153, 172, or 173.

77. The IdeS variant protein of any one of claims 1-21 or 76, the IdeS polypeptide conjugate of any one of claims 22-43 or the polypeptide of any one of claims 44-75, wherein the IdeS variant protein, the IdeS polypeptide conjugate or the polypeptide has decreased presentation of epitopes on human leukocyte antigen (HLA) as compared to a wild-type IdeS protein.

78. The IdeS variant protein of any one of claims 1-21 or 76, the IdeS polypeptide conjugate of any one of claims 22-43 or the polypeptide of any one of claims 44-75, wherein the IdeS variant protein, the IdeS polypeptide conjugate or the polypeptide has increased cleavage activity of immunoglobulins as compared to a wild-type IdeS protein.

79. The IdeS variant protein of any one of claims 1-21 or 76, the IdeS polypeptide conjugate of any one of claims 22-43 or the polypeptide of any one of claims 44-75, wherein the IdeS variant protein, the IdeS polypeptide conjugate or the polypeptide has a decrease in immunogenicity score more than 0 relative to SEQ ID NO: 1 or SEQ ID NO:2.

80. A polynucleotide encoding the IdeS variant protein of any one of claims 1-21 or 76, the IdeS polypeptide conjugate of any one of claims 22-43 or the polypeptide of any one of claims 44-75.

81. An expression plasmid comprising the polynucleotide as in claim 80 and a promoter.

82. A cell comprising the polynucleotide as in claim 80.

83. A pharmaceutical composition comprising the IdeS variant protein of any one of claims 1-21 or 76, the IdeS polypeptide conjugate of any one of claims 22-43 or the polypeptide of any one of claims 44-75, and a pharmaceutically acceptable carrier.

84. A method of treating an autoantibody-mediated autoimmune disease or disorder comprising administering an effective amount of the IdeS variant protein of any one of claims 1-21 or 76, the IdeS polypeptide conjugate of any one of claims 22-43, the polypeptide of anyAttorney Docket No.: CYR-009WO one of claims 44-75, or the pharmaceutical composition of claim 83 to a subject in need thereof.

85. The method of claim 84, wherein the autoantibody-mediated autoimmune disease is selected from Addison’s disease, Anti-glomerular basement membrane antibody disease (ABM, also known as Goodpasture’s disease), Axial Spondyloarthritis, Bullous Pemphigoid, celiac disease, chronic inflammatory demyelinating polyneuropathy (CIDP), dermatomyositis, Fetal and Neonatal Alloimmune Thrombocytopenia (FNAIT), generalized myasthenia gravis (gMG) and myasthenic crisis, Grave’s Disease, Guillain-Barre syndrome (GBS), Hashimoto’s thyroiditis, hemolytic disease of the fetus and newborn (HDFN), heparin induced thrombocytopenia (HIT), immune thrombocytopenia (ITP), inflammatory bowel disease (IBD) including Crohn’s disease and ulcerative colitis, Lupus Nephritis, Membranous Nephritis, multiple sclerosis, Myasthenia Gravis, myositis, MOG antibody disease (MOGAD), Obstetric Antiphospholipid Syndrome (OAPS), Pemphigus, Relapsing Polychondritis, psoriatic arthritis, psoriasis, rheumatoid Arthritis, scleroderma, Sjogren’s syndrome, stiff person syndrome, systemic lupus erythematosus (SLE), systemic sclerosis, Thrombotic Thrombocytopenic Purpura (TTP), thyroid eye disease (TED), vitiligo, vasculitis, warm autoimmune hemolytic anemia (wAIHA). .

86. The method of claim 84 or 85, wherein the autoantibody-mediated autoimmune disease is heparin induced thrombocytopenia (HIT) or immune thrombocytopenia (ITP).

87. The method of any one of claims 84-86, wherein the IdeS variant protein, polypeptide, or pharmaceutical composition is co-administered with a gene therapy.

88. A method for reducing presentation on HLA-II of IdeS comprising administering an effective amount of IdeS variant protein of any one of claims 1-21 or 76, the IdeS polypeptide conjugate of any one of claims 22-43, the polypeptide of any one of claims 44-75, or the pharmaceutical composition of claim 83.

89. A method for prolonging serum half-life of IdeS comprising administering an effective amount of the IdeS variant protein of any one of claims 1-21 or 76, the IdeS polypeptide conjugate of any one of claims 22-43, the polypeptide of any one of claims 44-75, or the pharmaceutical composition of claim 83.

90. A method for prolonging IgG depletion of IdeS comprising administering an effective amount of the IdeS variant protein of any one of claims 1-21 or 76, the IdeSAttorney Docket No.: CYR-009WO polypeptide conjugate of any one of claims 22-43, the polypeptide of any one of claims 44-75, or the pharmaceutical composition of claim 83.

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