Affinity ligands for purification of herpesvirus vaccine antigens

Abstract

Provided are human herpesvirus affinity ligands and related compositions including affinity resins.

Description

Docket No. 1580.00230WOAFFINITY LIGANDS FOR PURIFICATION OF HERPESVIRUS VACCINE ANTIGENSBACKGROUND

[0001] Protein-based vaccines have been used for the prophylaxis of bacterial and viral diseases in the United States for well over twenty-five years, the first vaccine including a viral protein having been approved in 1986. While protein-based vaccines are generally less immunogenic compared to vaccines comprised of either live-attenuated or inactivated whole organisms, their immunogenicity may be enhanced, for example, by mutation and / or by the use of adjuvants or other technologies including virus-like particles (VLPs) and protein micelles.

[0002] The polypeptide or protein component of a protein-based vaccine is typically manufactured using a recombinant eukaryotic expression systems, such as yeast, insect, or mammalian cells. It is important to obtain the recombinant polypeptide or protein at high purity for therapeutic use. Affinity purification allows for the efficient isolation of highly pure target polypeptide or protein. However, the use of this technique is limited by the availability of suitable affinity agents which selectively bind to the target polypeptide or protein with high affinity. In the absence of a suitable affinity agent, purification typically involves inefficient, labor intensive, and expensive processes, such as a multi-column process. The present invention addresses the need for affinity ligands against polypeptide antigens useful proteinbased vaccines, and related compositions.BRIEF SUMMARY

[0003] Provided are affinity ligands that bind to certain human herpesvirus antigens with high affinity and selectivity. Also provided are related compositions including affinity resins comprising the ligands described herein.

[0004] Provided are affinity ligands of Formula IL1-D1-L2-D2-L3-DE-DT-L4 (Formula I)whereinL1, L2, L3, and L4 are each independently absent or a linker;D1 and D2 are each independently an antigen-binding domain and D2 is optional; DE is an optional structural domain; andDT is an optional C-terminal tag domain.Docket No. 1580.00230WO

[0005] In aspects, at least one of D1 or D2 is defined by the amino acid sequence of Formula 1: VDAX1X2DX3X4X5X6X7AX8X9X10IX11X12LPNLX13X14X15QX16X17X18FIX19X20LX21X22DPSX 23X24X25X26LLX27X28AX29X30X31NX32X33QAPK (SEQ ID NO: 50), where X1 to X33 are as defined infra.

[0006] In aspects, at least one of D1 or D2 is defined by the amino acid sequence of Formula 1a:VDAX1FDX3EX5X6X7AX8X9EIX11X12LPNLNX14X15QX16X17AFIX19SLX21DDPSQSANLLAE AX29X30LNDAQAPK (SEQ ID NO: 51), where X1 to X30 are as defined infra. In aspects, at least one of D1 or D2 is defined by the amino acid sequence of any one of SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60, SEQ ID NO: 90, or SEQ ID NO: 91. In aspects, at least one of D1 or D2 comprises the amino acid sequence of any one of SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60, SEQ ID NO: 90, or SEQ ID NO: 91. In aspects, the affinity ligand comprises or consists of an amino acid sequence of SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 138 or SEQ ID NO: 139, or an amino acid sequence having at least 94% identity thereto.

[0007] In aspects, at least one of D1 or D2 is defined by the amino acid sequence of Formula 1b:VDAX1X2DX3X4LEX7ARX9X10IEX12LPNLTEEQRRAFIESLRDDPSQX24X25X26LLX27EAX29 X30LNX32AQAPK (SEQ ID NO: 52), where X1 to X32 are as defined infra. In aspects, at least one of D1 or D2 is defined by the amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, or SEQ ID NO: 70. In aspects, at least one of D1 orD2 comprises an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, or SEQ ID NO: 70, or an amino acid sequence having at least 94% identity thereto. In aspects, the affinity ligand comprises or consists of an amino acid sequence of SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, or an amino acid sequence having at least 94% identity thereto.

[0008] In aspects, at least one of D1 or D2 is defined by the amino acid sequence of FormulaDocket No. 1580.00230WO VDAKFDKELEEARAEIERLPNLTEX15QRRX18FIX19X20LRX22DPSX23SAX26LLAX28AX29X 30X31NDX33QAPK (SEQ ID NO: 53), where X15 to X33 are as defined infra. In aspects, at least one of D1 or D2 is defined by the amino acid sequence of any one of SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, or SEQ ID NO: 89.

[0009] Also provided are affinity ligands of Formula I where at least one of D1 or D2 is defined by the amino acid sequence of Formula 2:DEAX1RWX2X3LGX4AYX5X6RGDYDRAIEYYX7RALELDPNNAX8AWX9X10LGX11AYX12 X13RGDYDRAIEYYX14RALELDPX15X16AX17AWX18X19LGX20AYX21X22RGDYDRAIEYYX 23RX24LELDPX25X26EX27ARX28X29LEX30ARX31X32RX33 (SEQ ID NO: 54), where X1 to X33 are as defined infra. In aspects, at least one of D1 or D2 is defined by the amino acid sequence of any one of SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97. In aspects, at least one of D1 or D2 comprises an amino acid sequence of any one of SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94 or SEQ ID NO: 95, or an amino acid sequence having at least 94% identity thereto. In aspects, the affinity ligand comprises or consists of an amino acid sequence of any one of SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142 or SEQ ID NO: 143, or an amino acid sequence having at least 94% identity thereto.

[0010] The affinity ligand may also include where D2 is absent. In aspects, where D2 is absent, the affinity ligand may also include where Di is defined by the amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97.

[0011] The affinity ligand may also include where the two antigen-binding domains, Di and D2 are both present and each of Di and D2 is independently defined by the amino acid sequence of SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53 or SEQ ID NO: 54. The affinity ligand may also include where one of Di and D2 is defined by the amino acid sequence of SEQ ID NO: 77 and the remainder is defined by the amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95. The affinity ligand may also comprise an amino acid sequence as defined by SEQ ID NO: 42, SEQ ID NO: 43, or SEQ ID NO: 44. The affinity ligand may also include where one of Di and D2 is defined by the amino acid sequence of SEQ ID NO: 61 and the remainder is defined by the amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95. The affinity ligand may alsoDocket No. 1580.00230WOcomprise an amino acid sequence as defined by SEQ ID NO: 45, SEQ ID NO: 46, and SEQ ID NO: 47.

[0012] The affinity ligand may also include where at least one of D1 or D2 is defined by an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97, or an amino acid sequence having at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity thereto, optionally where any amino acid mutation of the reference protein defined by SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97 is a conservative amino acid substitution.

[0013] The affinity ligand may also include where the ligand includes two antigen-binding domains, Di and D2 and each of Di and D2 is independently defined by the amino acid sequence of SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53 or SEQ ID NO: 54. The affinity ligand may also include where each of Di and D2 independently includes an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 90, SEQ ID NO: 94 or SEQ ID NO: 95. The affinity ligand may also include wherein one of Di and D2 comprises or consists of an amino acid sequence of SEQ ID NO: 77 or SEQ ID NO: 90 and the remainder includes an amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95. The affinity ligand may also include where the affinity ligand comprises or consists of an amino acid sequence of SEQ ID NO: 42, SEQ ID NO: 43, or SEQ ID NO: 44. The affinity ligand may also include wherein one of Di and D2 comprises an amino acid sequence of SEQ ID NO: 61 and the remainder includes an amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95. The affinity ligand may also include where the affinity ligand comprises or consists of an amino acid sequence of any one of SEQ ID NO: 45, SEQ ID NO: 46, or SEQ ID NO: 47. The affinity ligand may also include where at least one of D1 or D2 comprises or consists of an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 90, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97, or an amino acid sequence having at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity thereto. The affinity ligand may also include where Di and D2 each independently comprises an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 90, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97, or an amino acid sequence having at least 94% identity thereto. The affinity ligand may also include where the affinity ligand comprises or consists of an amino acid sequence of any one of SEQ ID NO: 109, SEQ ID NO: 125, SEQ ID NO: 138, SEQ ID NO: 142 or SEQ ID NO: 143, or an aminoDocket No. 1580.00230WOacid sequence having at least 94% identity thereto. Other technical features may be readily apparent to one skilled in the art from the following figures, descriptions, and claims.

[0014] In accordance with aspects of any of the foregoing, the affinity ligand may also include where D2 is absent; where Li is a linker of from 1-6 amino acids; where L2. D2 and DT are absent; where L3 is absent or a linker of from 1-12 amino acids; where DE is present; where L4 is absent or a linker comprising from 1-15 amino acids; where the affinity ligand comprises a C-terminal thiol containing amino acid residue, optionally a cysteine or threonine residue, or one or more of the foregoing.

[0015] In accordance with aspects of any of the foregoing, the affinity ligand may also include where Li, L2, L3, and L4, when present, are each independently a single amino acid residue, a polypeptide, or a non-polypeptide molecule, optionally where the polypeptide is from 2-25, or from 2-12 amino acid residues in length. The affinity ligand may also include where L4, if present, is a C-terminal thiol containing amino acid residue, optionally a cysteine or threonine residue. The affinity ligand may also include where DT, when present, includes one or more peptide, polypeptide or protein tags selected from the group consisting of a bacteriophage T7 epitope (T7-tag), bacteriophage V5 epitope (V5-tag), biotin-carboxy carrier protein (BCCP), polyhistidine (His-tag), polyaspartate (Asp-tag), polycysteine (Cys-tag), polyphenylalanine (Phe-tag), glutathione S-transferase (GST), maltose binding protein (MBP), calmodulin binding peptide (CBP), intein-chitin binding domain (intein-CBD), streptavadin, streptavadin-binding peptide (SBP), Strep-tag, Protein A of Staphylococcus aureus and derivatives thereof, calmodulin-binding peptide (CBP), FLAG tag, human influenza hemagglutinin (HA), c-myc epitope, Glu-Glu, HSV epitope, and green fluorescent protein (GFP) and derivatives thereof.

[0016] In aspects, the affinity ligand binds to a human herpesvirus antigen where the antigen is a glycoprotein of a human herpesvirus. In aspects, the glycoprotein is selected from the group consisting of envelope glycoprotein H (gH), gL, gB, gC, gK, gG, gl, gD, gl, gE, glycoprotein pV (gpV), gpll, gpIII, gpIV, gpl, gp350 / 220, gp35, gp85, gp42, gpllO, ga, lEgp, and gp47 / 52, or a fragment or derivative of any of the foregoing.

[0017] In aspects, the affinity ligand binds to a human herpesvirus antigen, where the antigen is a polypeptide or protein of a human herpesvirus, optionally a glycoprotein. In aspects, the antigen is selected from the group consisting of capsid scaffolding protein (BVRF2 / BdRFl), capsid vertex component 1 (CVC1, BGLF1), capsid vertex component 2 (CVC2, BVRF1), envelope glycoprotein B (gB, BALF4), envelope glycoprotein GP350 (BLLF1), envelopeDocket No. 1580.00230WOglycoprotein H (gH, BXLF2), envelope glycoprotein L (gL, BKRF2), envelope glycoprotein M (gM, BBRF3), envelope glycoprotein N (gN, BLRF1), glycoprotein 42 (BZLF2), glycoprotein BDLF3 (BDLF3), glycoprotein BILF2 (BILF2), G-protein coupled receptor BILF1 (BFLF1), latent membrane protein 1 (LMP1, BNLF1), latent membrane protein 2 (LMP2), major capsid protein (MCP, BcLF1), major tegument protein (BNRF1), mRNA export factor ICP27 homolog (BMLF1, BSLF2), protein BDLF2 (BDLF2), protein BMRF2 (BMRF2), protein BNLF2a (BNLF2a), protein LF2 (LF2), protein UL24 homolog (BXRF1), putative BARFO protein (BARFO), secreted protein BARF1 (BARF1), triplex capsid protein 1 (TRX1, B0RF1), and triplex capsid protein 2 (TRX2, BDLF1), or a fragment or derivative of any of the foregoing. The affinity ligand may also include where the antigen is a polypeptide or protein of a human herpesvirus selected from the group consisting of glycoprotein gp42, glycoprotein gp35O, envelope glycoprotein H (gH), envelope glycoprotein L (gL), and latent membrane protein 2 (LMP2), or a fragment or derivative of any of the foregoing.

[0018] Also provided are affinity chromatography resins that include a solid support and a plurality of the affinity ligands as described herein covalently attached to the solid support. The affinity chromatography resin may also include where the solid support is in the form of discrete polymeric particles, ceramic or glass beads, a woven or non-woven membrane, or a polymeric monolith.

[0019] Other technical features may be readily apparent to one skilled in the art from the following figures, descriptions, and claims.BRIEF DESCRIPTION OF THE DRAWINGS

[0020] FIG. 1 is a line graph of absorbance (280 nm) versus volume (ml) for effluent fractions of vaccine antigen passed through a packed affinity column having resin functionalized with the ligand of SEQ ID NO: 40, which comprises a human herpesvirus antigen-binding domain having an amino acid sequence of SEQ ID NO: 94.

[0021] FIG. 2 is a line graph of percentage stability in NaOH normalized to DBCio at t=0 hrs versus simulated NaOH cycles for affinity resins prepared using ligands of SEQ ID NO: 7, SEQ ID NO: 23, SEQ ID NO: 40, and SEQ ID NO: 41. Each of these ligands comprises a human herpesvirus antigen-binding domain having an amino acid sequence of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, or SEQ ID NO: 95, respectively.Docket No. 1580.00230WODETAILED DESCRIPTION

[0022] Provided are human herpesvirus affinity ligands represented from N-terminus to C-terminus by Formula IL1-D1-L2-D2-L3-DE-DT-L4 (Formula I)whereinLi, L2, L3, and L4 are each independently absent or a linker;Di and D2 are each independently a human herpesvirus antigen-binding domain and D2 is optional;DE is an optional structural domain; andDT is an optional C-terminal tag domain.

[0023] Also provided are human herpesvirus affinity ligands of Formula IaL1-D1-L2-D2-L3 (Formula Ia)whereinLi, L2, and L3 are each independently absent or a linker;Di and D2 are each independently a human herpesvirus antigen-binding domain and D2 is optional.

[0024] In aspects of Formula I or Formula Ia, D1 or D2 comprises or consists of an amino acid sequence of Formula 1:VDAX1X2DX3X4X5X6X7AX8X9X10IX11X12LPNLX13X14X15QX16X17X18FIX19X20LX21X2 2DPSX23X24X25X26LLX27X28AX29X30X31NX32X33QAPK (SEQ ID NO: 50) whereinX1 is R or K; X18 is A, G, R, Y, D, F, W;X2 is F, R, Q, T, H, K, or E; X19 is R, S, H, Y, W, E, L, Q, T;X3 is R, K, H, A, Q, Y, I, F; X20 is S, A, K, H, W;X4 is E, Y, D, S, W, R; X21 is R, Y, A, I, V;X5 is L, V, I, F, W; X22 is W, V, Y, I, F, L, Q, S, D;X6 is W, F, E, A, H; X23 is Q, A, S, H, K, E, R, N;X7 is E, Y, R, F, K, W, T, D, Q, H; X24 is S, R, K, E, W;X8 is Q, L, Y, R; X25 is A, V, Y, H, R, W;X9 is A, K, H, N, V, G, T, S, E, W, Y, D; X26 is N, D, L, Y, F, W, K, R, T, H, Q, A, G, S, I; X10 is E, Y, I, V, K, R, H; X27 is A, N, L, H, W, Y, D;X11 is E, R, N, H, Q, W; X28 is E, F, S, R, L, A, H, K, T, I, Q, Y; X12 is D, Q, S, H, K, E, W, F, R; X29 is R, K, W, H, Y, Q, L, S, T;Docket No. 1580.00230WOX13 is N or T; X30 is R, K, F, D, Y, I, W, T, V, H, S, L, A; X14 is E, V, N, L, Y, W; X31 is L, K, R, H, D, Y, W, F, S;X15 is R, Y, W, Q, H, D, K, S, A, E; X32 is D, W, N, E, H, K, R, F; and X16 is R, K, W, Q; X33 is A, D, W, V, R, K, N, E, S, L.X17 is H, W, Y, G, S, T, E, R;

[0025] Also provided are affinity ligands of Formula I or Formula la where at least one of D1 or D2 comprises the amino acid sequence of Formula la:VDAX1FDX3EX5X6X7AX8X9EIX11X12LPNLNX14X15QX16X17AFIX19SLX21DDPSQSA NLLAEAX29X30LNDAQAPK (SEQ ID NO: 51) whereinX1 is R or K; X3 is R or K; X5 is L, V, I, F, W; X6 is W, F, E, A, H; X7 is E, T, D, Q, H; X8 is Q, L, Y; X9 is E, W, Y, D; X11 is R, N, H, Q, W; X12 is D, Q, S, H, K; X14 is V, N, L, Y, W; X15 is R, Q, H, K, S, A, E; X16 is R, K, W, Q; X17 is H, W, Y, G, S, T, E; X19 is R, H, Y, L, Q, T; X21 is R, Y, A, I, V; X29 is R or K; and X30 is R or K.

[0026] Also provided are affinity ligands of Formula Ia whereinX1 is R or K; X3 is R or K; X5 is L or V; X6 is A; X7 is H; X8 is Y; X9 is W; X11 is R; X12 is H; X14 is V or L; X15 is K; X16 is R; X17 is G or S; X19 is R; X21 is A or V; X29 is R or K; and X30 is R or K.

[0027] Also provided are affinity ligands of Formula I or Formula la where at least one of D1 or D2 comprises or consists of an amino acid sequence of Formula lb:VDAX1X2DX3X4LEX7ARX9X10IEX12LPNLTEEQRRAFIESLRDDPSQX24X25X26LLX27EAX29X30LNX32AQAPK (SEQ ID NO: 52) whereinX1 is R or K; X2 is R, Q, T, H, K, E; X3 is R, H, A, Q, Y, I, F; X4 is E, Y, D, S, W, R; X7 is Y, R, F, K, W; X9 is A, K, H, N, V, G, T, S, E; X10 is Y, I, V, K, R, H; X12 is S, E, W, F, R; X24 is R, K, E, W; X25 is V, Y, H, R, W; X26 is D, L, Y, F, W, K, R, T, H; X27 is A, N, L, H, W, Y, D; X29 is R, W, H, Y, Q, L, S, T; X30 is R, F, D, Y, R, I, W, T, V; and X32 is W, N, E, H, K, R, F.

[0028] Also provided are affinity ligands of Formula 1b whereX1 is R or K; X2 is R or K; X3 is Q; X4 is R; X7 is Y; X9 is K; X10 is K or R; X12 is S; X24 is K or R; X25 is W; X26 is R; X27 is D; X29 is R; X30 is T; and X32 is W.Docket No. 1580.00230WO

[0029] Also provided are affinity ligands of Formula I or Formula la where at least one of D1 or D2comprises or consists of an amino acid sequence of Formula 1c:VDAKFDKELEEARAEIERLPNLTEX15QRRX18FIX19X20LRX22DPSX23SAX26LLAX28AX29X30X31NDX33QAPK (SEQ ID NO: 53) whereinX15 is R, Y, W, Q, H, D, K, S, A; X18 is G, R, Y, D, F, W; X19 is R, S, H, Y, W; X20 is S, A, K, H, W; X22 is W, V, Y, I, F, L, Q, S; X23 is Q, A, S, H, K, E, R, N; X26 is N, D, L, W, K, R, T, H, Q, A, G, S, N, I; X28 is F, S, R, L, A, H, K, T, I, Q, Y; X29 is R or K; X30 is R, F, W, T, H, S, L, A; X31 is L, K, R, H, D, Y, W, F, S; and X33 is A, D, W, V, R, K, N, E, S, L.

[0030] Also provided are affinity ligands of Formula I or Formula la where at least one of D1 or D2comprises or consists of an amino acid sequence of Formula 2:DEAX1RWX2X3LGX4AYX5X6RGDYDRAIEYYX7RALELDPNNAX8AWX9X10LGX11 AYX12X13RGDYDRAIEYYX14RALELDPX15X16AX17AWX18X19LGX20AYX21X22RGDYDRAIEYYX23RX24LELDPX25X26EX27ARX28X29LEX30ARX31X32RX33 (SEQ ID NO: 54) wherein X1 is A or G; X18 is I, W, L, or G;X2 is D, W, or Q; X19 is D, L, H, or A;X3 is D, W, or Y; X20 is V, R, S, or E;X4 is W, T, F, or E; X21 is A or T;X5 is A, I, V, or L; X22 is A, S, or I;X6 is A, V, S, or Q; X23 is E or Q;X7 is E or Q; X24 is A or G;X8 is L, Y, A, or T; X25 is E or N;X9 is S, R, V, or I; X26 is D or N;X10 is I, L, or A; X27 is D, A, or R;X11 is R, D, or E; X28 is F, W, I, or N;X12 is A, D, or L; X29 is R, V, or Y;X13 is A, S, V or Y; X30 is A, D, or W;X14 is Q or E; X31 is K or R;X15 is N or D; X32 is A or S; andX16 is N or D; X33 is K or R.X17 is F, T, H, or W;

[0031] Also provided are affinity ligands of Formula 2 whereDocket No. 1580.00230WOX1 is A or G; X18 is I, W;X2 is D, W; X19 is D, L;X3 is W, or Y; X20 is V, R;X4 is W, T,; X21 is A or TX5 is A, I,; X22 is A, S;X6 is A, V; X23 is E;X7 is E; X24 is A or G;X8 is L, Y; X25 is E;X9 is S, R; X26 is D;X10 is I; X27 is D;X11 is R, D; X28 is F, W;X12 is A, D; X29 is R, V;X13 is A, S; X30 is A;X14 is E; X31 is K;X15 is D; X32 is A; andX16 is D; X33 is K.X17 is F, T;

[0032] In accordance with aspects of any of the foregoing, L1, L2, L3, and / or L4, if present, is a linker, as described in more detail infra. In some aspects, the linker is a peptide linker of from 1-12 amino acids in length, optionally from 1-10 amino acids in length or from 1-6 amino acids in length. In aspects, any one or more of L1, L2, L3, and / or L4, if present, is independently a poly(glycine) peptide, poly(alanine) peptide, poly((glycine)(alanine)) peptide, or a poly((glycine)(serine)) peptide. In aspects, a C-terminal linker, L3 or L4, if present, is a C-terminal thiol containing amino acid residue, optionally a cysteine or threonine residue.

[0033] In accordance with aspects of any of the foregoing, the structural domain DE is a nontarget molecule binding structural domain. In aspects, the structural domain may enhance soluble protein expression and / or serve as a tag domain for use in affinity purification of the target molecule. Exemplary structural domains include glutathione S-transferase (GST), maltose binding protein (MBP), small ubiquitin-like modifier (SUMO), Staphylococcal protein A (SpA), and derivatives of any of the foregoing. In aspects, Ds may comprise or consist of a polypeptide derivative of SpA that does not bind to the Fc region of immunoglobulin proteins. Additional exemplary structural domains include ecotin, the Z-domain of SpA, the albumin-Docket No. 1580.00230WObinding domain of protein G, the cellulose-binding domain of endoglucanase, disulfide bond oxidoreductase, and Barnase, an enzymatically inactive variant an RNase enzyme of Bacillus amyloliquefaciens. See review by A. Malik, 3 Biotech. 2016 Feb 4;6(1):44.

[0034] In aspects of any of the foregoing, the tag domain DT may include one or more C-terminal tags, as described in more detail infra. In aspects, DT comprises at least one affinity tag to facilitate isolation during manufacture and / or to facilitate detection of the affinity ligand.

[0035] In aspects, provided are affinity ligands of Formula la where at least one of D1 or D2 is defined by any one of SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60, SEQ ID NO: 90, or SEQ ID NO: 91.

[0036] In aspects, provided are affinity ligands of Formula lb where at least one of D1 or D2is defined by the amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, or SEQ ID NO: 70.

[0037] In aspects, provided are affinity ligands of Formula 1c where at least one of D1 or D2 is defined by the amino acid sequence of any one of SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, or SEQ ID NO: 89.

[0038] In aspects, provided are affinity ligands of Formula 2 where at least one of D1 or D2is defined by the amino acid sequence of any one of SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97.

[0039] Also provided are affinity ligands of Formula I or Formula la comprising two antigenbinding domains, Di and D2 where each of Di and D2is independently defined by the amino acid sequence of SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53 or SEQ ID NO: 54.

[0040] In aspects provided is an affinity ligand of Formula I or Formula la comprising two antigen-binding domains Di and D2 where one of Di and D2 is defined by the amino acid sequence of SEQ ID NO: 77 and the remainder is defined by the amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95. Representative affinity ligands include the proteins defined by the amino acid sequence of SEQ ID NO: 42, SEQ ID NO: 43, and SEQ ID NO: 44.Docket No. 1580.00230WO

[0041] In aspects provided is an affinity ligand of Formula I or Formula la comprising two antigen-binding domains Di and D2 where one of Di and D2 is defined by the amino acid sequence of SEQ ID NO: 61 and the remainder is defined by the amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95. Representative affinity ligands include the proteins defined by the amino acid sequence of SEQ ID NO: 45, SEQ ID NO: 46, and SEQ ID NO: 47.

[0042] In aspects, provided is an affinity ligand of Formula I or Formula la where at least one of D1 or D2 is defined by the amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97.

[0043] In aspects, provided is an affinity ligand of Formula 1 or Formula la where at least one of D1 or D2 is defined by an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97, or an amino acid sequence having at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity thereto, optionally wherein any amino acid mutation of the reference protein defined by SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97 is a conservative amino acid substitution.

[0044] The affinity ligands described here advantageously are able to withstand being subjected to conditions of high pH, including a pH of 13 or higher, for prolonged periods of time, without a significant decrease in their ligand-binding capacity. The alkali stability of the proteins described here is particularly advantageous for chromatographic processes incorporating a cleaning in place procedure utilizing highly alkaline solutions, including for example 0.1 M NaOH or 0.5 M NaOH solutions. For example, a resin functionalized with an affinity ligand as described herein may retain at least 80%, preferably at least 90%, of its ligand-binding capacity following exposure to 0.1 M NaOH or 0.5 M NaOH for at least 10 hours, or at least 20 hours, or at least 30 hours, where ligand-binding capacity is measured as a percentage of the dynamic binding capacity of the functionalized resin.

[0045] The affinity ligands described here also demonstrate high “dynamic binding capacity” or DBC, when coupled to a resin. DBC is defined as the mass loaded per unit of resin when there is 10% breakthrough in the column effluent. Typical units for DBC are given in milligrams protein per milliliter resin (mg / ml), or equivalently, grams per liter (g / L). Column DBC must be maintained at a high level during the manufacturing process in order to maintain overall process efficiency in terms of cost and capacity. Accordingly, in preferred aspects a resin functionalized with affinity ligands as described herein has a 10% breakthrough dynamicDocket No. 1580.00230WObinding capacity for viral antigen of at least 5-10 g / L or higher, for example at a residence time of between 4-10 minutes. In aspects, a resin functionalized with affinity ligands as described herein has a dynamic binding capacity (DBC) in the range of at least about 50 g / L, about 60 g / L, about 70 g / L, or about 80 g / L or higher for viral antigen. In aspects, a resin functionalized with affinity ligands as described herein has a DBC of from about 70-90 mg / ml, or from about 75-85 mg / ml for viral antigen.

[0046] Accordingly, provided are affinity ligands having chemical stability under alkali conditions such as commonly used for cleaning in place (CIP) procedures. Also provided are chromatography resins functionalized with the affinity ligands described here, which advantageously have a high dynamic binding capacity (DBC) for viral antigens of a human herpesvirus.Affinity ligands

[0047] Provided are affinity ligands of Formula I or or Formula la, as described above. The affinity ligands may be referred to interchangeably as “affinity agents” or “affinity ligands” or in some instances simply as “ligands”.

[0048] The affinity ligands described here bind with high affinity to a human herpesvirus antigen. In aspects, the affinity ligands described here bind with high affinity to an antigen which is a human herpesvirus polypeptide or protein, optionally a glycoprotein, or a fragment or derivative thereof. In aspects, the human herpesvirus antigen is a glycoprotein selected from the group consisting of envelope glycoprotein H (gH), gL, gB, gC, gK, gG, gJ, gD, gl, gE, glycoprotein pV (gpV), gpll, gpIII, gpIV, gpl, gp350 / 220, gp35, gp85, gp42, gpl 10, get, lEgp, and gp47 / 52, or a peptide or polypeptide fragment of any of the foregoing. In aspects, the human herpesvirus antigen is selected from capsid scaffolding protein (BVRF2 / BdRFl), capsid vertex component 1 (CVC1, BGLF1), capsid vertex component 2 (CVC2, BVRF1), envelope glycoprotein B (gB, BALF4), envelope glycoprotein GP350 (BLLF1), envelope glycoprotein H (gH, BXLF2), envelope glycoprotein L (gL, BKRF2), envelope glycoprotein M (gM, BBRF3), envelope glycoprotein N (gN, BLRF1), glycoprotein 42 (BZLF2), glycoprotein BDLF3 (BDLF3), glycoprotein BILF2 (BILF2), G-protein coupled receptor BILF1 (BILF1), latent membrane protein 1 (LMP1, BNLF1), latent membrane protein 2 (LMP2), major capsid protein (MCP, BcLF1), major tegument protein (BNRF1), mRNA export factor ICP27 homolog (BMLF1, BSLF2), protein BDLF2 (BDLF2), protein BMRF2 (BMRF2), protein BNLF2a (BNLF2a), protein LF2 (LF2), protein UL24 homolog (BXRF1), putative BARF0 proteinDocket No. 1580.00230WO(BARFO), secreted protein BARF1 (BARF1), triplex capsid protein 1 (TRX1, BORF1), and triplex capsid protein 2 (TRX2, BDLF1), or a fragment or derivative thereof. In aspects, provided is affinity ligand that binds to a human herpesvirus polypeptide or protein selected from glycoprotein gp42, glycoprotein gp350, envelope glycoprotein H (gH), envelope glycoprotein L (gL), and latent membrane protein 2 (LMP2), or a peptide or polypeptide fragment of any of the foregoing.

[0049] In aspects, provided is an affinity ligand comprising one or more units of a binding domain represented by Di or D2 in Formula 1, Formula la, Formula lb, Formula Ic, Formula II or Formula III. An affinity ligand comprising two or more units of a domain as described herein may also be referred to as a “multimer” or by the number of units of the triple helical bundle domain present in the protein. For example an affinity ligand having two units of the domain may be referred to as a dimer. Similarly, an affinity ligand having three, four, five, or six units of the domain may be referred to as a trimer, a quatramer, a pentamer, or a hexamer, respectively. In the context of a multimer, the two or more units of the domain are covalently attached to each other, either directly or via a linker molecule. Linker molecules are described in more detail infra, but generally a linker molecule is a single amino acid residue, a polypeptide, or a non-polypeptide molecule.

[0050] The affinity ligands described herein may be functionalized with one or more N- or C-terminal modifications to facilitate its purification during manufacture and / or to facilitate its covalent attachment to a solid support to form an affinity chromatography matrix or resin as described herein. In this context, the term “functionalized” refers to a modification by covalent attachment. The one or more C-terminal modifications may include a spacer or linker molecule or a peptide, polypeptide, or protein tag.

[0051] In the context of the present disclosure, the terms “spacer” and “linker” are used interchangeably. In aspects where an affinity ligand as described herein is functionalized with a linker, the linker may consist of a single amino acid residue, a polypeptide, or a non-polypeptide molecule. In aspects, the single amino acid spacer may be an alanine, glycine^ serine, proline, aspartic acid, glutamic acid, or any natural or non-natural amino acid residue. In aspects, the single amino acid spacer is a glycine, alanine, or serine residue. In aspects, the polypeptide linker may consist of a polypeptide comprising a majority of amino acids selected from glycine, alanine, proline, asparagine, glutamine, and lysine. In aspects, the linker may consist of a polypeptide comprising a majority of amino acids selected from glycine, alanine,Docket No. 1580.00230WOproline, asparagine, aspartic acid, threonine, glutamine, and lysine. In aspects, the linker may consist of a polypeptide comprising a majority of amino acids selected from glycine, serine, and / or alanine. In some aspects, the linker is selected from a poly(glycine) peptide, a poly(alanine) peptide, a poly((glycine)(alanine)) peptide and a poly((glycine)(serine)) peptide. Polypeptide linkers may be from about 1 to 50 amino acids, from about 1 to 20 amino acids, from about 1 to 15 amino acids, from about 1 to 10 amino acids, from about 1 to 5 amino acids, from about 2 to 20 amino acids, from about 2 to 15 amino acids, from about 2 to 10 amino acids, or from about 2 to 5 amino acids in length. In aspects, the poly(glycine) peptide, poly(alanine) peptide, poly((glycine)(alanine)) peptide or a poly((glycine)(serine)) peptide linker consists of from about 2 to 10 amino acids, or from 2 to 6 amino acids, or from 2 to 4 amino acids.

[0052] In other aspects, the spacer or linker may be a non-polypeptide molecule, for example a substituted or unsubstituted C2-C30 alkyl spacer or a polyethylene glycol (PEG) spacer. In aspects, the C2-C30 alkyl spacer is -NH-(CH2)n-C(O)-, where n is from 2 to 30. In aspects, the alkyl spacer is substituted by a lower alkyl, e.g., Ci-Ce alkyl, acyl, halogen, -CN, -NH2, or phenyl. In aspects, a PEG spacer has a molecular weight of from about 100 to 5000 kDa, or from about 100 to 500 kDa.

[0053] In aspects, an affinity ligand as described herein may be functionalized with a peptide, polypeptide, or protein tag, for example a C-terminal affinity tag, to facilitate its isolation during manufacture and / or its detection. The term “tag” refers to a polypeptide covalently attached to another polypeptide or protein, generally to facilitate isolation and / or detection of the tagged polypeptide or protein. Such tags may be from about 4 to 500 amino acids in length. Tags allow for isolation, detection and / or localization of the tagged polypeptide or protein using methods such as affinity purification or immunodetection with an appropriate antibody combined with detection e.g., by Western analyses, ELISA assays, or immunostaining. Exemplary tags that may be utilized include the bacteriophage T7 epitope (T7-tag), bacteriophage V5 epitope (V5-tag), biotin-carboxy carrier protein (BCCP), polyhistidine (His-tag), polyaspartate (Asp-tag), polycysteine (Cys-tag), polyphenylalanine (Phe-tag), glutathione S-transferase (GST), maltose binding protein (MBP), calmodulin binding peptide (CBP), intein-chitin binding domain (intein-CBD), a streptavidin / biotin-based tag such as streptavadin, streptavadin-binding peptide (SBP) or Strep-tag, a tandem affinity purification (TAP) tag, such as Protein A of Staphylococcus aureus and calmodulin-binding peptide (CBP),Docket No. 1580.00230WOor other TAP variants, a short epitope tag such as FLAG, human influenza hemagglutinin (HA), c-myc epitope, T7, Glu-Glu, HSV epitope, and small fluorescent proteins such as the green fluorescent protein (GFP) and derivatives thereof.

[0054] In aspects, an affinity ligand described here may include a C-terminal polyhistidine tag, also referred to as a “His-tag”, which may include, for example, 3-10 histidine residues, preferably about 5 or 6 histidine residues. In aspects, an affinity ligand described here may include one or more tags in the C-terminal tag domain.

[0055] Also provided are affinity chromatography matrices comprising a solid support functionalized with an affinity ligand as described herein. In this context, the term “functionalized” refers to a modification of the solid support by covalent attachment of a plurality of affinity ligands.

[0056] The affinity ligands may be covalently attached to the solid support via a reactive thiol- or nitrogen-containing amino acid. Alternatively or additionally, the affinity ligands may be covalently attached to the solid support via reactive epoxy or aldehyde groups of the support itself. Accordingly, in aspects, the polypeptides are covalently attached to the solid support directly, via a reactive thiol- or nitrogen-containing C-terminal amino acid of the polypeptide, or indirectly, via a polypeptide linker. In aspects, an affinity ligand as described herein may be modified to include a C-terminal reactive amino acid, e.g., a reactive thiol- or nitrogencontaining C-terminal amino acid, to facilitate covalent attachment of the polypeptide to the solid support. In aspects, the reactive amino acid is a lysine or cysteine. In aspects where the affinity ligand is covalently attached to the solid support via a C-terminal polypeptide linker, the C-terminal polypeptide linker may comprise from 2 to 50 amino acids, from 2 to 25 amino acids, from 2 to 15 amino acids, from 2 to 10 amino acids, or from 2 to 5 amino acids. In aspects, the C-terminal polypeptide linker comprises one or more reactive amino acids e.g., a reactive thiol- or nitrogen-containing amino acid, to facilitate covalent attachment of the linker to the solid support. In aspects, the one or more reactive amino acids includes one or more lysine and / or cysteine residues.

[0057] In aspects, the solid support is in the form of discrete polymeric particles, ceramic or glass beads, a woven or non-woven membrane, or a polymeric monolith. The discrete polymeric particles may be made from a polysaccharide such as agar, agarose, dextran, starch, cellulose, pullulan, etc., and stabilized variants and derivatives thereof; or from a synthetic polymer such as polystyrene, polyvinylether, polyvinyl alcohol, polyacrylate,Docket No. 1580.00230WOpolymethacrylate, polyacrylamide, etc. In aspects, the discrete particles may have a volume-weighted median diameter (d50,v or dv50) in the range of about 20 to 100 micrometers, or 40 to 90 micrometers, or 45 to 55 micrometers, or 80 to 90 micrometers. In aspects, the dv50 of the particles is 50 or 85 microns. In aspects, the particles are made from a material selected from agarose or a stabilized derivative thereof such as PraestoOPure or Praesto® Jetted A50, having a volume-weighted median diameter (d50,v) of about 50 or about 85 micrometers or having a d50v of from about 45 to 90 micrometers.

[0058] In aspects, provided is an affinity chromatography resin functionalized with at least about 1 to 20 mg / ml In aspects, an affinity chromatography resin functionalized with the affinity ligands described here may be subjected to a typical cleaning in place or “CIP” process without significant decrease in ligand-binding capacity. Typical CIP processes include subjecting the affinity resin to conditions of alkaline pH, including for example 0.01-0.05 M NaOH, acidic pH, or high concentration of a chaotropic agent such as 2-8 M urea or 2-6 M guanidine hydrochloride. In some aspects, an affinity chromatography resin functionalized with the affinity ligands described here may retain at least 80%, preferably at least 90%, of its ligand-binding capacity following exposure to 0.01-0.05 M NaOH or 6 M guanidine hydrochloride for at least 10 hours, or at least 20 hours, or at least 30 hours, where ligandbinding capacity is measured as a percentage of the dynamic binding capacity (DBC) of the functionalized resin. In aspects, an affinity chromatography resin functionalized with the affinity ligands described here are provided as single-use columns, as described herein, for example about 1 to 5 mg / ml or 5 to 15 mg / ml or 10-20 mg / ml or 15 to 20 mg / ml. In aspects, a column comprising the affinity chromatography resin has a dynamic binding capacity (DBC) of at least about 0.1 to 0.5 g / L or 0.1 to 0.5 mg / ml for ligand. In aspects, a column comprising the affinity chromatography resin has a dynamic binding capacity (DBC) of at least about 0.2 g / L or 0.2 mg / ml, 0.3 g / L or 0.3 mg / ml, 0.4 g / L or 0.4 mg / ml, or 0.5 g / L or 0.5 mg / ml for ligand.

[0059] Suitable solid supports include, for example, agarose and stabilized derivatives of agarose (e.g. Praesto®Pure, Praesto® Jetted A50, Sepharose 6B, Sepharose Fast Flow, etc.), cellulose or derivatives of cellulose, controlled pore glass, monolith (e.g. C1M® monoliths), silica, zirconium oxide (e.g. CM Zirconia or CPG®), titanium oxide, or synthetic polymers (e.g. polystyrene, polyvinylether, polyvinyl alcohol, monodisperse polyacrylate resin, polyhydroxyalkyl acrylates, polyhydroxyalkyl methacrylates, polyacrylamides, polymethacrylamides, etc.) and hydrogels of various compositions. In certain aspects, the solidDocket No. 1580.00230WOsupport comprises a polyhydroxy polymer, such as a polysaccharide. Examples of suitable polysaccharides include agar, agarose, dextran, starch, cellulose, pullulan, etc., and stabilized variants thereof. In some aspects, the solid support is made from agarose or a stabilized derivative thereof, such as PraestoOPure, Praesto® Jetted A50, or polyvinyl divinyl benzene, silica, or control pore glass.

[0060] In other aspects the solid support may be a microchip, e.g., a silicon, silicon-glass, or gold microchip. In aspects, the solid support may be formed from nitrocellulose, paper, plastic, nylon, metal, or any combination of the foregoing. In some aspects, the solid support may be a plate or dish, such as a microtiter plate or dish.

[0061] In aspects, an affinity chromatography resin functionalized with affinity ligands as described here may be subjected to a typical cleaning in place or “CIP” process without significant decrease in ligand-binding capacity. Typical CIP processes include subjecting the affinity resin to conditions of alkaline pH, including for example 0.01-0.05 M NaOH, acidic pH, or high concentration of a chaotropic agent such as 2-8 M urea or 2-6 M guanidine hydrochloride. In some aspects, an affinity chromatography resin functionalized with the affinity ligands described here may retain at least 80%, preferably at least 90%, of its ligandbinding capacity following exposure to 0.01-0.05 M NaOH or 6 M guanidine hydrochloride for at least 10 hours, or at least 20 hours, or at least 30 hours, where ligand-binding capacity is measured as a percentage of the dynamic binding capacity (DBC) of the functionalized resin. In aspects, an affinity chromatography resin functionalized with the ligand-binding polypeptides or chimeric proteins described here are provided as single-use columns.

[0062] Also provided are systems for purification of ligand-containing molecules in a manufacturing process. In aspects, the manufacturing system comprises at least one step of affinity chromatography utilizing an affinity chromatography resin functionalized with the affinity ligands described here.

[0063] The affinity ligands described herein may be produced synthetically e.g., by liquid or solid phase chemical synthetic methods, and / or other techniques such as polymerase chain reaction (PCR) based synthesis, concatemerization, seamless cloning, and recursive directional ligation (RDL). The affinity ligands may also be produced recombinantly via expression in a host cell from polynucleotides encoding the polypeptides. Such polynucleotides may also be produced synthetically or using recombinant techniques based on the amino acid sequences of the affinity ligands disclosed herein. It is understood that different nucleic acid sequences mayDocket No. 1580.00230WObe utilized to encode the same affinity ligand, for example where codon optimization is utilized to facilitate expression in a particular cell system. Methodologies and tools for constructing an optimized RNA sequence for protein expression are known in the art. See e.g., Strategies of codon optimization for high-level heterologous protein expression in microbial expression systems, Gene Reports 9:46-53 (2017); and “A new and updated resource for codon usage tables” Athey, J., Alexaki, A., Osipova, E. et al. A new and updated resource for codon usage tables. BMC Bioinformatics 18, 391 (2017).

[0064] Polynucleotides encoding the polypeptides described herein may comprise control elements such as promoters, enhancers, ribosomal binding sites, transcription termination signals, and polyadenylation signals, or may be inserted into an appropriate expression vector containing one or more such control elements for expression of the polynucleotides in a host cell such as a bacterial cell, yeast cell, insect cell, plant cell or mammalian cell. Examples of prokaryotic host expression systems, viral expression systems, yeast expression systems, plant expression systems, and mammalian expression systems are known in the art. Insertion of of the polynucleotides into a suitable expression vector can be accomplished using in vitro recombinant DNA techniques, synthetic techniques, or in vivo recombination / genetic recombination techniques. Suitable expression vectors are commercially available and may include, for example, plasmid vectors, single and double-stranded phage vectors, or single and double-stranded RNA or DNA viral vectors. Phage and viral vectors may also be introduced into host cells in the form of packaged or encapsulated viral particles using known techniques for infection and transduction. Alternatively, cell-free translation systems may also be used to produce the polypeptides.

[0065] In an exemplary embodiment, affinity ligands are produced recombinantly in an appropriate expression system such as an E. Coli Pichia Pastoris or in a mammalian cell system, using standard techniques. The recombinantly produced polypeptides are purified using multi-column chromatography. For example, histidine-tagged polypeptides may be purified using immobilized metal ion affinity chromatography (“IMAC”). The purity and identity of recombinant polypeptides may be assessed by a combination of gel electrophoresis, e.g., SDS-PAGE, reverse-phase ultra-high performance chromatography (“RP-UPLC”), quadrupole time-of-flight mass spectrometry, and size-exclusion chromatography (“SEC”).

[0066] It should be noted that the N-terminal methionine of the recombinantly produced polypeptides described here may be cleaved during expression resulting in polypeptides lackingDocket No. 1580.00230WOthe N-terminal methionine, or a mixture of polypeptides which contain or lack the N-terminal methionine. The presence or absence of the N-terminal methionine does not impact the function of the polypeptides or proteins described herein. Accordingly, the N-terminal methionine should be considered optional in the amino acid sequences of the affinity ligands described herein where it is present.Functional characterization of affinity ligands

[0067] The ability of the affinity ligands to bind to target ligand containing proteins is determined, for example, by measurement of binding affinity constants such as KA and / or KD using assays that may include one or more of competition analysis, equilibrium analysis, microcalorimetric analysis, and real-time interaction analysis based on surface plasmon resonance interaction (c.., using a BIACORE™ instrument). These methods are described, for example, in Neri etal. (1996) Tihtech 14:465-470 and Jansson el al. (1997) J Biol Chem 272:8189-8197. In aspects, an affinity ligand described here exhibits a dissociation constant (KD ) for target ligand containing proteins of about 1×10⁻⁴ M to about 1×10⁻⁵ M, about 1×10⁻⁵ M to about 1×10⁻⁶ M, about 1×10⁻⁶ M to about 1×10⁻⁷ M, about 1×10⁻⁷ M to about 1×10⁻⁸ M, about 1×10⁻⁸ M to about 1×10⁻⁹ M, about 1×10⁻⁹ M to about 1×10⁻¹⁰ M, about 1×10⁻¹⁰ M to about 1×10⁻¹¹ M, or about 1×10⁻¹¹ M to about 1×10⁻¹² M.

[0068] The determination of KD may be performed under different conditions of pH and salt concentration using suitable buffers. For example, low pH solutions (<pH 5.5) can be made using citrate buffers, glycine-HCl buffers, or succinic acid buffers. High pH solutions can be made, for example, using Tris-HCl buffers, phosphate buffers, or sodium bicarbonate buffers. A number of conditions may be used to determine KD and off-rates for the purpose of determining, for example, optimal pH and / or salt concentrations.

[0069] The ability of the affinity ligands to bind to target ligand containing proteins may also be determined, for example, using biolayer interferometry (BLI) analysis. BLI is a technique that measures the thickness of a biological layer at the tip of a sensor though interferometry. Thickness is correlated to binding events such as protein: protein interactions. In some aspects, an affinity ligand described here binds a target ligand containing protein with a biolayer interferometry (BLI) response of less than or equal to 15 nm, 10 nm, 5 nm, 3 nm, or 1 nm.

[0070] In aspects binding of the affinity ligands is characterized by one or more of sensogram, adsorption, SBC, and / or DBC analyses.Docket No. 1580.00230WOEXAMPLESProduction of recombinant protein ligands

[0071] Recombinant protein ligands were expressed in E. Coli and / or Pichia Pastoris using standard techniques. Ligands were purified using multi-column chromatography. For his-tagged ligands IMAC was used as the primary capture step. Biotinylated ligands were generated with the Avitag™ system (Avidity, Aurora, CO). Non-biotinylated ligands bearing the Avitag™ sequence were prepared by omitting exogenous biotin. The purity and identity of recombinant protein ligands was assessed by a combination of SDS-PAGE, RP UPLC, quadrupole time-of-flight mass spectrometry and SEC. In many instances the ligand is isolated without the N-terminal methionine residue, which is presumed to be cleaved during expression. In many instances a mixture is obtained with only a proportion of the purified ligand containing the N-terminal methionine. It is obvious to those skilled in the art that the presence or absence of the N-terminal methionine does not affect the conclusions herein. For clarity, we include the N-terminal methionine.Antigen binding selectivity of affinity ligands

[0072] Characterization of ligand binding to two selected human herpesvirus antigens was determined using biolayer interferometry (BLI) (ForteBio, Menlo park, CA). Biotinylated affinity ligands were immobilized on sensors and incubated with solutions containing each herpesvirus antigen at various concentrations. The binding curves were fitted with a 1:1 binding model using the ForteBio software to determine affinity. The results are shown in Table 1 below for representative affinity ligands within the scope of Formula I or la. Also shown are the sequence identifiers for the antigen-binding domain, Di and / or D2, of each ligand. The data show that all affinity agents bound both vaccine antigens with high affinity. In most cases, the binding affinity was less than 10 nM and in many cases less than 1 nM.

[0073] Table 1. Binding (Kd (nM) as measured by BLI) of representative affinity ligands to vaccine antigen 1 or vaccine antigen 2 of human herpesvirus.affinity ligand antigen-binding Antigen 1 Antigen 2domain (Di or D2)SEQ ID NO 4 SEQ ID NO: 58 < 1SEQ ID NO 5 SEQ ID NO: 59 < 1SEQ ID NO 6 SEQ ID NO: 60 < 1SEQ ID NO 9 SEQ ID NO: 63 < 1SEQ ID NO: 10 SEQ ID NO: 64 < 1Docket No. 1580.00230WOSEQ ID NO 11 SEQ ID NO 65 < 1SEQ ID NO 12 SEQ ID NO 66 < 1SEQ ID NO 13 SEQ ID NO 67 < 1SEQ ID NO 14 SEQ ID NO 68 < 1SEQ ID NO 15 SEQ ID NO 69 < 1SEQ ID NO 16 SEQ ID NO 70 < 1SEQ ID NO 18 SEQ ID NO 72 < 1SEQ ID NO 21 SEQ ID NO 75 < 1SEQ ID NO 25 SEQ ID NO 79 < 1SEQ ID NO 26 SEQ ID NO 80 < 1SEQ ID NO 27 SEQ ID NO 81 < 1 89SEQ ID NO 28 SEQ ID NO 82 < 1SEQ ID NO 29 SEQ ID NO 83 < 1SEQ ID NO 30 SEQ ID NO 84 < 1SEQ ID NO 31 SEQ ID NO 85 < 1SEQ ID NO 32 SEQ ID NO 86 < 1SEQ ID NO 33 SEQ ID NO 87 < 1SEQ ID NO 34 SEQ ID NO 88 < 1SEQ ID NO 35 SEQ ID NO 89 < 1SEQ ID NO 36 SEQ ID NO 90 < 1SEQ ID NO 38 SEQ ID NO 92 < 1SEQ ID NO 39 SEQ ID NO 93 < 1SEQ ID NO 40 SEQ ID NO 94 < 1SEQ ID NO 41 SEQ ID NO 95 < 1Measurement of affinity ligand binding on resin

[0074] Affinity resins described herein demonstrate high binding capacity toward selected human herpesvirus vaccine antigens. Select vaccine antigens were diluted to a concentration of 0.5 g / L and pumped over a packed column containing affinity resins prepared with from the ligand corresponding to SEQ ID NO: 40. A 0.3 x 5 cm (0.353 mL) column was operated according to Table 2.

[0075] Table 2Step Buffer CV Residence Time V (cm / hr)(min)Equilibration PBS 28 1.0 300 Load Sample Vaccine antigen in PBS 20 4.0 75 Chase PBS 10 4.0 75 Elution 1.0 M MgCh, 50 mM Tris, pH 8.0 11 4.0 75 Wash Water 14 1.0 300 PAB Strip PAB (120 mM Phosphoric acid, 167 11 1.4 212mM Acetic acid, 2.2% benzylalcohol)Docket No. 1580.00230WONaOH Strip 0.1 MNaOH 11 1.4 212Equilibration PBS 28 1.0 300

[0076] The materials were analyzed by measuring absorbance of each effluent fraction compared to the absorbance of the load solution. The aggregate mass of the vaccine antigen challenge at the point when the effluent measures 10% of the load solution is reported as the dynamic binding capacity of the resin. The data is shown in FIG. 1 for an exemplary affinity resin produced from the affinity ligand corresponding to SEQ ID NO: 40. Dynamic binding capacity (DBC) measured for exemplary affinity resins is reported in Table 3.

[0077] Table 3: DBC of affinity resins prepared with the indicated ligands, each comprising the indicated human herpesvirus binding domain, for binding to vaccine antigen 1 or vaccine antigen 2 of human herpesvirus.affinity ligand antigen-binding Antigen 1 DBC Antigen 2 DBC (g / LREs) domain (Di or (g / LREs)02) _SEQ ID NO 4 SEQ ID NO 58 0.7SEQ ID NO 7 SEQ ID NO 61 6.9SEQ ID NO 23 SEQ ID NO 77 7.3SEQ ID NO 40 SEQ ID NO 94 4.2SEQ ID NO 41 SEQ ID NO 95 4.1SEQ ID NO 43 SEQ ID NO: 97 4.0 2.2Affinity agent tolerance to sodium hydroxide measured on resin

[0078] This example demonstrates the binding characteristics and sodium hydroxide stability of affinity chromatography resins prepared from affinity ligands described herein. Affinity resins described herein demonstrated robust tolerance to 0.1 M NaOH, enabling its use as a CIP agent during industrial manufacturing of select vaccine antigens. FIG. 2 shows the dynamic binding capacity of exemplary affinity resins before and after repeated exposures to 0.1 M NaOH.

[0079] Resins were packed into a 0.353 mL column (0.3 x 5 cm), and initial binding capacity was obtained as described above. Resin was then held in a static hold of 0.1 M NaOH for some time, and binding capacity was re-assessed at the end of that time. NaOH stability is reported in % binding capacity of the initially measured binding capacity value.Production and characterization of affinity agentsDocket No. 1580.00230WO

[0080] This example demonstrates the production and characterization of affinity agents comprising ligands identified and described herein. Affinity resins were prepared by conjugating ligands to agarose beads. The following is a non-limiting example of affinity ligand conjugation to a commercially available base bead: Praesto® Jetted A50 beads (Purolite, King of Prussia, PA) were activated with disuccinimidyl carbonate and coupled with excess ethylenediamine. After washing, bromoacetate was conjugated to the aminated beads using EDC activation. After washing, ligands were conjugated to the beads at room temperature. Epoxy activated beads were either obtained from the vendor or prepared using standard methods well known in the art. Ligands were coupled to epoxy activated beads at 37-40 °C. Targeted ligand densities were varied from 5-30 g / L. Following washing, the beads were deactivated with excess thioglycerol. The actual ligand density for all resins was measured using a subtractive RP-HPLC method according to the following formula:Actual Ligand Density = (Measured [ligand] in feed - Measured [ligand] in effluent).

[0081] While the invention herein disclosed has been described by means of specific embodiments and applications thereof, modifications and variations could be made thereto by those skilled in the art without departing from the scope of the invention set forth in the claims.

[0082] It will be appreciated that the present invention is set forth in various levels of detail in this application. In certain instances, details not necessary for one of ordinary skill in the art to understand the invention, or that render other details difficult to perceive may have been omitted. The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting beyond the scope of the appended claims. Unless defined otherwise, technical terms used herein are to be understood as commonly understood by one of ordinary skill in the art to which the disclosure belongs.

[0083] Various features of a process system may be used independently of, or in combination, with each other. It will be appreciated that a system as disclosed herein may be embodied in different forms and should not be construed as limited to the illustrated embodiments of the figures.

[0084] It should be understood that, as described herein, an “embodiment” (such as illustrated in the accompanying Figures) may refer to an illustrative representation of an environment or article or component in which a disclosed concept or feature may be provided or embodied, orDocket No. 1580.00230WOto the representation of a manner in which just the concept or feature may be provided or embodied. However such illustrated embodiments are to be understood as examples (unless otherwise stated), and other manners of embodying the described concepts or features, such as may be understood by one of ordinary skill in the art upon learning the concepts or features from the present disclosure, are within the scope of the disclosure. In addition, it will be appreciated that while the Figures may show one or more embodiments of concepts or features together in a single embodiment of an environment, article, or component incorporating such concepts or features, such concepts or features are to be understood (unless otherwise specified) as independent of and separate from one another and are shown together for the sake of convenience and without intent to limit to being present or used together. For instance, features illustrated or described as part of one embodiment can be used separately, or with one or more other features to yield a still further embodiment. Thus, it is intended that the present subject matter covers such modifications and variations as come within the scope of the appended claims and their equivalents.

[0085] In the foregoing description and the following claims, the following will be appreciated. The phrases “at least one”, “one or more”, and “and / or”, as used herein, are open-ended expressions that are both conjunctive and disjunctive in operation. The terms “a”, “an”, “the”, “first”, “second”, etc., do not preclude a plurality. For example, the term “a” or “an” entity, as used herein, refers to one or more of that entity. As such, the terms “a” (or “an”), “one or more” and “at least one” can be used interchangeably herein.

[0086] The term “about” when used before a numerical designation, e.g., temperature, time, amount, concentration, and such other, including a range, indicates approximations which may vary by ( + ) or ( - ) 10%, 5%, 1%, or any subrange or subvalue there between. Preferably, the term “about” means that the value may vary by + / - 10%.

[0087] In the claims, the term “comprises / comprising” does not exclude the presence of other elements, components, features, regions, integers, steps, operations, etc. Additionally, although individual features may be included in different claims, these may possibly advantageously be combined, and the inclusion in different claims does not imply that a combination of features is not feasible and / or advantageous. In addition, singular references do not exclude a plurality. Reference signs in the claims are provided merely as a clarifying example and shall not be construed as limiting the scope of the claims in any way. By contrast, the transitional phrase “consisting of’ excludes any element, step, or ingredient not specified in the claim. TheDocket No. 1580.00230WOtransitional phrase “consisting essentially of’ limits the scope of a claim to the specified materials or steps “and those that do not materially affect the basic and novel character! stic(s)” of the claimed invention.

[0088] The term “dissociation constant” or “Kd” refers to the dissociation equilibrium constant of the particular interaction between a first protein and a second protein. A ligandbinding protein is considered to bind to a target ligand-containing protein where it has a dissociation constant Kd of less than about 15 uM or less than about 10 uM. In some aspects, a ligand-binding protein as described herein may have a Kd of 500 nM or less, 100 nM or less, 50 nM or less, 20 nM or less, or 10 nM or less for a target ligand-containing protein or a reference ligand-containing protein. Kd may be determined, for example by Surface Plasmon Resonance (SPR) using a commercially available kit such as the Biacore™ assay sold by Cytiva Life Sciences.

[0089] The terms “protein” and “polypeptide” refer to any linear molecular chain of two or more amino acids linked by peptide bonds and does not refer to a specific length of the product. Thus, “peptides”, “protein”, “amino acid chain,” or any other term used to refer to a chain of two or more amino acids, are included within the definition of “polypeptide,” and the term “polypeptide” may be used instead of, or interchangeably with any of these terms. The term “polypeptide” is also intended to refer to the products of post-translational modifications of the polypeptide, including without limitation glycosylation, acetylation, phosphorylation, amidation, proteolytic cleavage, modification by non-naturally occurring amino acids and similar modifications which are well-known in the art. Thus, Ig binding proteins comprising two or more protein domains also fall under the definition of the term “protein” or “polypeptides”.

[0090] The terms “alkaline stable” or “alkaline stability” may be used interchangeably herein and refer to the ability of the Ig binding protein or Ig binding domain of the invention to withstand alkaline conditions without significantly losing the ability to bind to immunoglobulins. The skilled person in this field can easily test alkaline stability by incubating an Ig binding protein or Ig binding domain with, for example, sodium hydroxide solutions, e.g., as described in the Examples, and subsequent testing of the binding capacity or binding activity to immunoglobulin by routine experiments known to someone skilled in the art, for example, by chromatographic approaches. The alkaline stability may be determined by coupling the Ig binding protein or Ig binding domain of the invention to a surface plasmon resonance (SPR)Docket No. 1580.00230WOsensor chip, and assaying the binding capacity or binding activity for immunoglobulin before and after exposure to an alkaline solution. The alkaline treatment can be performed, for instance, in 0.5 M NaOH for an extended period of time, e.g., at least 5 hours, at least 8 hours, at least 12 hours, at least 16 hours, or at least 20 hours.

[0091] The term “percent (%) identity”, in the context of two or more polypeptide sequences, refer to two or more sequences or subsequences that are the same or have a specified percentage of amino acid residues or nucleotides that are the same, when compared and aligned for maximum correspondence, as measured using one of the following sequence comparison algorithms or by visual inspection. Percent identity may be determined using a computer algorithms such as the Basic Local Alignment Search Tool (“BLAST”) or a related tool, including other BLAST-based tools available at the US National Library of Medicine, National Center for Biotechnology Information website. The BLAST and related algorithms have been described, for example, in Altschul et al., 1990, J. Mol. Biol. 215:3, 403-410; and Altschul et al., 1997, Nucleic Acids Res. 25:17, 3389-402.

[0092] The term “substitution” in the context of an amino acid substitution refers to an exchange of an amino acid at a particular position in a polypeptide sequence with a different amino acid. Similarly, the term “deletion” in the context of an amino acid deletion refers to the removal of an amino acid at a particular position in a polypeptide sequence; and the term “insertion” in the context of amino acid insertion refers to the addition of an amino acid to the polypeptide sequence.

[0093] The term “chromatography” refers to separation technologies which employ a mobile phase and a stationary phase to separate one type of molecules (e.g., immunoglobulins) from other molecules (e.g. contaminants or other immunoglobulins) in a sample. A liquid mobile phase contains a mixture of molecules and transports these across or through a stationary phase (such as a solid resin, support, or matrix, which terms are used interchangeably). Due to the differential interaction of the different molecules in the mobile phase with the stationary phase, molecules in the mobile phase can be separated. The term “affinity chromatography” refers to a specific type of chromatography in which a ligand having a specific affinity for a target molecule is coupled to the stationary phase. The ligand interacts with the target molecule in the mobile phase thereby separating it from the mobile phase. Generally, the target molecule is eluted from the stationary phase in a separate step. The terms “solid support” or “solid matrix” or “resin” are used interchangeably to refer to the stationary phase.Docket No. 1580.00230WO

[0094] A “conservative” amino acid substitution is one in which one amino acid residue is replaced with another having similar side chain chemistry and / or size. Families of amino acid residues having similar side chain chemistry have been defined in the art. For example, those having basic side chains, e.g., lysine (K), arginine (R), histidine (H); acidic side chains, e.g., aspartic acid (D), glutamic acid (E); uncharged polar side chains, e.g., glycine (G), asparagine (N), glutamine (Q), serine (S), threonine (T), tyrosine (Y), cysteine (C); nonpolar side chains, e.g., alanine (A), valine (V), leucine (L), isoleucine (I), proline (P), phenylalanine (F), methionine (M), tryptophan (W); beta-branched side chains, e.g., threonine (T), valine (V), isoleucine (I); and aromatic side chains, e.g., tyrosine (Y), phenylalanine (F), tryptophan (W), histidine (H).Additional numbered embodiments

[0095] Embodiment 1. A human herpesvirus affinity ligand of Formula TL1-D1-L2-D2-L3-DE-DT-L4 (Formula I)whereinL1, L2, L3, and L4 are each independently absent or a linker;Di and D2 are each independently an antigen-binding domain and D2 is optional;DE is an optional structural domain; andDT is an optional C-terminal tag domain.

[0096] Embodiment 2. The affinity ligand of Embodiment 1, wherein at least one of D1 or D2is defined by the amino acid sequence of Formula 1:VDAX1X2DX3X4X5X6X7AX8X9X10IX11X12LPNLX13X14X15QX16X17X18FIX19X20LX21X2 2DPSX23X24X25X26LLX27X28AX29X30X31NX32X33QAPK (SEQ ID NO: 50)whereinXi is R or K; Xis is A, G, R, Y, D, F, W;X2is F, R, Q, T, H, K, E; Xi9is R, S, H, Y, W, E, L, Q, T;X3is R, K, H, A, Q, Y, 1, F; X2ois S, A, K, H, W;X4 is E, Y, D, S, W, R; X21 is R, Y, A, I, V;X5is L, V, I, F, W; X22is W, V, Y, I, F, L, Q, S, D;Xsis W, F, E, A, H; X23is Q, A, S, H, K, E, R, N;X7 is E, Y, R, F, K, W, T, D, Q, H; X24 is S, R, K, E, W;X8is Q, L, Y, R; X25 is A, V, Y, H, R, W;Docket No. 1580.00230WOX9is A, K, H, N, V, G, T, S, E, W, Y, D; X26 is N, D, L, Y, F, W, K, R, T, H, Q, A, G, S, I;Xiois E, Y, I, V, K, R, H; X27 i s A, N, L, H, W, Y, D;Xn is E, R, N, H, Q, W; X28 is E, F, S, R, L, A, H, K, T, I, Q, Y; Xi2is D, Q, S, H, K, E, W, F, R; X29 is R, K, W, H, Y, Q, L, S, T;X13 is N or T; X30 is R, K, F, D, Y, I, W, T, V, H, S, L, A Xi4 is E, V, N, L, Y, W; X3iis L, K, R, H, D, Y, W, F, S;X15 is R, Y, W, Q, H, D, K, S, A, E; X32 is D, W, N, E, H, K, R, F; and X16 is R, K, W, Q; X33 is A, D, W, V, R, K, N, E, S, L. X17 is H, W, Y, G, S, T, E, R;

[0097] Embodiment 3. The affinity ligand of Embodiment 1 or 2, wherein at least one of D1 or D2 is defined by the amino acid sequence of Formula 1a:VDAX1FDX3EX5X6X7AX8X9EIX11X12LPNLNX14X15QX16X17AFIX19SLX21DDPSQSA NLLAEAX29X30LNDAQAPK (SEQ ID NO: 51)whereinXi is R or K;X3 is R or K;X5is L, V, I, F, W;Xeis W, F, E, A, H;X7is E, T, D, Q, H;X8is Q, L, Y;X9is E, W, Y, D;Xn is R, N, H, Q, W;Xi2is D, Q, S, H, K;X14 is V, N, L, Y, W;X15 is R, Q, H, K, S, A, E;Xi6 is R, K, W, Q;X17 is H, W, Y, G, S, T, E;Xi9is R, H, Y, L, Q, T;X2iis R, Y, A, I, V;X29 is R or K; andDocket No. 1580.00230WOXao is R or K.

[0098] Embodiment 4. The affinity ligand of Embodiment 3, wherein at least one of D1 or D2is defined by the amino acid sequence of any one of SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60, SEQ ID NO: 90, or SEQ ID NO: 91.

[0099] Embodiment 5. The affinity ligand of Embodiment 1 or 2, wherein at least one of D1 or D2 is defined by the amino acid sequence of Formula 1b:VDAXiX2DX3X4LEX7ARX9XioIEXi2LPNLTEEQRRAFIESLRDDPSQX24X25X26LLX27EAX29 X30LNX32AQAPK (SEQ ID NO: 52)whereinXi is R or K;X2is R, Q, T, H, K, E;X3is R, H, A, Q, Y, I, F;X4is E, Y, D, S, W, R;X7is Y, R, F, K, W;Xg is A, K, H, N, V, G, T, S, E;Xiois Y, I, V, K, R, H;Xi2is S, E, W, F, R;X24is R, K, E, W;X25 is V, Y, H, R, W;X26 is D, L, Y, F, W, K, R, T, H;X27is A, N, L, H, W, Y, D;X29is R, W, H, Y, Q, L, S, T;X30is R, F, D, Y, R, I, W, T, V; andX32is W, N, E, H, K, R, F.

[0100] Embodiment 6. The affinity ligand of Embodiment 5, wherein at least one of D1 or D2is defined by the amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, or SEQ ID NO: 70.

[0101] Embodiment 7. The affinity ligand of Embodiment 1 or 2, wherein at least one of D1 or D2 is defined by the amino acid sequence of Formula 1c:Docket No. 1580.00230WO VDAKFDKELEEARAEIERLPNLTEX15QRRX18FIX19X20LRX22DPSX23SAX26LLAX28AX29X 30X31NDX33QAPK (SEQ ID NO: 53)whereinX15 is R, Y, W, Q, H, D, K, S, A;X18is G, R, Y, D, F, W;Xi9is R, S, H, Y, W;X2ois S, A, K, H, W;X22is W, V, Y, I, F, L, Q, S;X23 is Q, A, S, H, K, E, R, N;X26is N, D, L, W, K, R, T, H, Q, A, G, S, N, I;X28is F, S, R, L, A, H, K, T, I, Q, Y;X29 is R or K;X3ois R, F, W, T, H, S, L, A;X31 is L, K, R, H, D, Y, W, F, S; andX33 is A, D, W, V, R, K, N, E, S, L.

[0102] Embodiment 8. The affinity ligand of Embodiment 7, wherein at least one of D1 or D2is defined by the amino acid sequence of any one of SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, or SEQ ID NO: 89.

[0103] Embodiment 9. The affinity ligand of Embodiment 1, wherein at least one of D1 or D2is defined by the amino acid sequence of Formula 2:DEAXIRWX2X3LGX4AYX5X6RGDYDRAIEYYX7RALELDPNNAXSAWX9XIOLGXII AYX12X13RGDYDRAIEYYX14RALELDPX15X16AX17AWX18X19LGX20AYX21X22RGDYDRA IEYYX23RX24LELDPX25X26EX27ARX28X29LEX3OARX31X32RX33 (SEQ ID NO: 54) whereinXi is A or G; X18is I, W, L, or G;X2is D, W, or Q; X19 is D, L, H, or A;X3is D, W, or Y; X2o is V, R, S, or E;X4 is W, T, F, or E; X21 is A or T;X5is A, I, V, orL; X22is A, S, or I;Docket No. 1580.00230WOX6 is A, V, S, or Q; X23 is E or Q;X7 is E or Q; X24 is A or G;Xs is L, Y, A, or T; X25 is E or N;X9is S, R, V, or I; X26 is D or N;X10 is I, L, or A; X27is D, A, or R;X11 is R, D, or E; X28 is F, W, I, or N;X12 is A, D, or L; X29 is R, V, or Y;X13 is A, S, V or Y; X30 is A, D, or W;X14 is Q or E; X31 is K or R;X15 is N or D; X32 is A or S; andX16 is N or D; X33 is K or R.X17 is F, T, H, or W;

[0104] Embodiment 10. The affinity ligand of Embodiment 9, wherein at least one of D1 or D2is defined by the amino acid sequence of any one of SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97.

[0105] Embodiment 11. The affinity ligand of any one of Embodiments 1 to 10, wherein Li, L2, L3, and L4, when present, are each independently a single amino acid residue, a polypeptide, or a non-polypeptide molecule, optionally wherein the polypeptide is from 2-25, or from 2-12 amino acid residues in length.

[0106] Embodiment 12. The affinity ligand of Embodiment 11, wherein L4, if present, is a C-terminal thiol containing amino acid residue, optionally a cysteine or threonine residue.

[0107] Embodiment 13. The affinity ligand of Embodiment 11, wherein DT, when present, comprises one or more peptide, polypeptide or protein tags selected from the group consisting of a bacteriophage T7 epitope (T7-tag), bacteriophage V5 epitope (V5-tag), biotin-carboxy carrier protein (BCCP), polyhistidine (His-tag), polyaspartate (Asp-tag), polycysteine (Cys-tag), polyphenylalanine (Phe-tag), glutathione S-transferase (GST), maltose binding protein (MBP), calmodulin binding peptide (CBP), intein-chitin binding domain (intein-CBD), streptavadin, streptavadin-binding peptide (SBP), Strep-tag, Protein A of Staphylococcus aureus and derivatives thereof, calmodulin-binding peptide (CBP), FLAG tag, human influenza hemagglutinin (HA), c-myc epitope, Glu-Glu, HSV epitope, and green fluorescent protein (GFP) and derivatives thereof.Docket No. 1580.00230WO

[0108] Embodiment 14. The affinity ligand of any one of Embodiments 1 to 13, wherein D2 is absent.

[0109] Embodiment 15. The affinity ligand of Embodiment 14, wherein Di is defined by the amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97.

[0110] Embodiment 16. The affinity ligand of Embodiment 1, wherein the protein comprises two antigen-binding domains, Di and D2 and each of Di and D2 is independently defined by the amino acid sequence of SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53 or SEQ ID NO: 54.

[0111] Embodiment 17. The affinity ligand of Embodiment 16, wherein one of Di and D2 is defined by the amino acid sequence of SEQ ID NO: 77 and the remainder is defined by the amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95.

[0112] Embodiment 18. The affinity ligand of Embodiment 17, wherein the affinity ligand comprises or consists of an amino acid sequence as defined by SEQ ID NO: 42, SEQ ID NO: 43, or SEQ ID NO: 44.

[0113] Embodiment 19. The affinity ligand of Embodiment 16, wherein one of Di and D2 is defined by the amino acid sequence of SEQ ID NO: 61 and the remainder is defined by the amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95.

[0114] Embodiment 20. The affinity ligand of Embodiment 19, wherein the affinity ligand comprises or consists of an amino acid sequence as defined by SEQ ID NO: 45, SEQ ID NO: 46, and SEQ ID NO: 47.

[0115] Embodiment 21. The affinity ligand of Embodiment 1, wherein at least one of D1 or D2is defined by an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97, or an amino acid sequence having at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity thereto, optionally wherein any amino acid mutation of the reference protein defined by SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97 is a conservative amino acid substitution.

[0116] Embodiment 22. The affinity ligand of any one of Embodiments 1 to 21, wherein the affinity ligand binds to a human herpesvirus antigen selected from the group consisting of envelope glycoprotein H (gH), gL, gB, gC, gK, gG, gJ, gD, gl, gE, glycoprotein pV (gpV),Docket No. 1580.00230WOgpll, gpIII, gpiv, gpl, gp350 / 220, gp35, gp85, gp42, gpllO, ga, lEgp, and gp47 / 52, or a fragment or derivative of any of the foregoing.

[0117] Embodiment 23. The affinity ligand of any one of Embodiments 1 to 21, wherein the affinity ligand binds to a human herpesvirus antigen selected from the group consisting of capsid scaffolding protein (BVRF2 / BdRFl), capsid vertex component 1 (CVC1, BGLF1), capsid vertex component 2 (CVC2, BVRF1), envelope glycoprotein B (gB, BALF4), envelope glycoprotein GP350 (BLLF1), envelope glycoprotein H (gH, BXLF2), envelope glycoprotein L (gL, BKRF2), envelope glycoprotein M (gM, BBRF3), envelope glycoprotein N (gN, BLRF1), glycoprotein 42 (BZLF2), glycoprotein BDLF3 (BDLF3), glycoprotein BILF2 (BILF2), G-protein coupled receptor BILF1 (BILF1), latent membrane protein 1 (LMP1, BNLF1), latent membrane protein 2 (LMP2), major capsid protein (MCP, BcLF1), major tegument protein (BNRF1), mRNA export factor ICP27 homolog (BMLF1, BSLF2), protein BDLF2 (BDLF2), protein BMRF2 (BMRF2), protein BNLF2a (BNLF2a), protein LF2 (LF2), protein UL24 homolog (BXRF1), putative BARFO protein (BARFO), secreted protein BARF1 (BARF1), triplex capsid protein 1 (TRX1, BORF1), and triplex capsid protein 2 (TRX2, BDLF1), or a fragment or derivative of any of the foregoing.

[0118] Embodiment 24. The affinity ligand of Embodiment 23, wherein the antigen is a polypeptide or protein selected from the group consisting of glycoprotein gp42, glycoprotein gp350, envelope glycoprotein H (gH), envelope glycoprotein L (gL), and latent membrane protein 2 (LMP2), or a fragment or derivative of any of the foregoing.

[0119] Embodiment 25. An affinity chromatography resin comprising a solid support and a plurality of the affinity ligands of any one of Embodiments 1 to 24 covalently attached to the solid support.

[0120] Embodiment 26. The affinity chromatography resin of Embodiment 25, wherein the solid support is in the form of discrete polymeric particles, ceramic or glass beads, a woven or non-woven membrane, or a polymeric monolith.Informal Sequence ListingResidues SEQ ID MAQGTVDAKFDKEVWTAYWEIRHLPNLNVKQKEAFILSLVDDP SQSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 1 NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFEAQKIEWHEHHHHHHCDocket No. 1580.00230WOResidues SEQ ID MAQGTVDAKFDKELHDAYWEIRKLPNLNVQQKSAFILSLVDDPS QSANLLAEAKKLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 2 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDARFDREFWEAQDEIWDLPNLNWEQQHAFITSLRDDP SQSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLPSEQ ID NO: 3 NLTEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIF EAQKIEWHEHHHHHHC MAQGTVDARFDREFEDALYEINSLPNLNYRQWYAFIQSLIDDPS QSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 4 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDARFDREFFEALEEIQQLPNLNYHQWWAFIQSLYDDPS QSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 5 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDARFDREWEDALEEIHQLPNLNYAQWHAFIYSLIDDPS QSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 6 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPS QRWRLLDEARTLNWAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 7 NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLND AQAPKGLNDIF EAQKIEWHEHHHHHHC MAQGTVDAKQDQWLEFARNRIEWLPNLTEEQRRAFIESLRDDPS QKRTLLYEATVLNRAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 8 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKQDIWLEFARHRIERLPNLTEEQRRAFIESLRDDPSQ KRRLLYEASTLNNAQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 9 TEEQRNKFIQ SLKDDP S Q S ANLL AE AKKLND AQ APKGLNDIFE A QKIEWHEHHHHHHC MAQGTVDAKRDYWLEKARVKIEFLPNLTEEQRRAFIESLRDDPS QRHKLLWEAHWLNKAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 10 NLTEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIF EAQKIEWHEHHHHHHC MAQGTVDARQDFRLERARHHIESLPNLTEEQRRAFIESLRDDPSQ RWHLLAEARRLNFAQAPKAAAVDAKFDKEQQAARAEILHLPNLSEQ ID NO: 11 TEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTVDARHDHYLEWAREYIEELPNLTEEQRRAFIESLRDDPS QWVDLLNEAWFLNWAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 12 NLTEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIF EAQKIEWHEHHHHHHC MAQGTVDAKKDASLEFARGVIEWLPNLTEEQRRAFIESLRDDPS SEQ ID NO: 13QEYWLLHEALILNHAQAPKAAAVDAKFDKEQQAARAEILHLPNDocket No. 1580.00230WOResidues SEQ ID LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKTDQSLEWARAIIEWLPNLTEEQRRAFIESLRDDPS QEYYLLNEAYYLNNAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 14 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKEDADLEYARSIIEWLPNLTEEQRRAFIESLRDDPSQ EYLLLLEAHDLNNAQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 15 TEEQRNKFIQ SLKDDP S Q S ANLL AE AKKLND AQ APKGLNDIFE A QKIEWHEHHHHHHC MAQGTVDARTDRELEFARTVIEWLPNLTEEQRRAFIESLRDDPSQ EYFLLLEAQRLNEAQAPKAAAVDAKFDKEQQAARAEILHLPNLT SEQ ID NO: 16 EEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIFEAQ KIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTERQRRYFIRKLRWDPS QSAILLAYAKRSNDLQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 17 TEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEDQRRFFIRKLRVDPSR SARLLATAKHYNDNQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 18 TEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEYQRRYFIRKLRWDPS QSANLLAYAKRWNDSQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 19 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQ APKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRWFISKLRWDPS RSAGLLAQAKAYNDDQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 20 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEKQRRRFISHLRWDPS NSAWLLAKAKLYNDAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 21 NLTEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIF EAQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEHQRRYFIRKLRWDPS RSAGLLAIAKHWNDDQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 22 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPS RSASLLAYAKLYNDEQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 23 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTERQRRYFIRKLRWDPS QSATLLAYAKTFNDRQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 24 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFEAQKIEWHEHHHHHHCDocket No. 1580.00230WOResidues SEQ ID MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRYFIRKLRWDPS H SARLL AY AKFFNDEQAPK AAA VDAKFDKEQ Q AARAEILHLPN SEQ ID NO: 25 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEKQRRYFIHWLRYDPS RSAKLLAHAKSKNDRQAPKAAAVDAKFDKEQQAARAEILHLPNSEQ ID NO: 26 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTESQRRYFIWALRLDPS KSAHLLARAKWRNDVQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 27 NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIF EAQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRRFIWSLRQDPS ESAHLLASAKRKNDWQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 28 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTESQRRDFIYSLRQDPSH SAHLLAFAKWHNDKQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 29 TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEAQRRDFIWALRSDPS ESAKLLAFAKWHNDRQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 30 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRDFIYSLRFDPSS SALLLALAKWHNDEQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 31 TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRDFIWSLRLDPS ASAQLLAFAKWHNDNQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 32 NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIF EAQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEYQRRDFIHALRVDPS SSAHLLAFAKWHNDRQAPKAAAVDAKFDKEQQ AARAEILHLPN SEQ ID NO: 33 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEDQRRGFIYALRIDPSA SAALLAAAKHDNDRQAPKAAAVDAKFDKEQQAARAEILHLPNLSEQ ID NO: 34 TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEDQRRGFIHSLRVDPS HSADLLAFARWLNDDQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 35 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELAHAYWEIRHLPNLNLKQRGAFIRSLADDP SEQ ID NO: 36SQSANLLAEAKKLNDAQAPKAAAVDAKFDKEQQAARAEILHLPDocket No. 1580.00230WOResidues SEQ ID NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIF EAQKIEWHEHHHHHHC MAQGTVDAKFDKEIFQAYWEIRHLPNLNNHQKTAFIHSLVDDPS QSANLLAEAKKLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 37 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQAAAGDEAGRWQWLGEAYLQRGDYDRAIEYYQRALELDPN NATAWIALGDAYLYRGDYDRAIEYYQRALELDPNNAWAWGAL GEAYAARGDYDRAIEYYQRALELDPNNERARNVLEWARRSRRA SEQ ID NO: 38 AAVDAKFDKEQQAARAEILHLPNLTEEQRNKFIQSLKDDPSQSA NLLAEAKKLNDAQAPKGLNDIFEAQKIEWHEHHHHHHC MADEAARWDDLGFAYVSRGDYDRAIEYYERALELDPNNAAAW VLLGEAYAVRGDYDRAIEYYERALELDPDDAHAWLHLGSAYTI RGDYDRAIEYYERALELDPEDEAARIYLEDARKARKAAAVDAK SEQ ID NO: 39 FDKEQQAARAEILHLPNLTEEQRNKFIQSLKDDPSQSANLLAEAK KLNDAQAPKGLNDIFEAQKIEWHEHHHHHHC MADEAARWDWLGWAYAARGDYDRAIEYYERALELDPNNALA WSILGRAYAARGDYDRAIEYYERALELDPDDAFAWIDLGVAYA ARGDYDRAIEYYERGLELDPEDEDARFRLEAARKARKAAAVDA SEQ ID NO: 40 KFDKEQQAARAEILHLPNLTEEQRNKFIQSLKDDPSQSANLLAEA KKLNDAQAPKGLNDIFEAQKIEWHEHHHHHHC MADEAARWWYLGTAYIVRGDYDRAIEYYERALELDPNNAYAW RILGDAYDSRGDYDRAIEYYERALELDPDDATAWWLLGRAYTS RGDYDRAIEYYERALELDPEDEDARWVLEAARKARKAAAVDA SEQ ID NO: 41 KFDKEQQAARAEILHLPNLTEEQRNKFIQSLKDDPSQSANLLAEA KKLNDAQAPKGLNDIFEAQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPS RSASLLAYAKLYNDEQAPKQPRGPTIKPDEAARWWYLGTAYIV RGDYDRAIEYYERALELDPNNAYAWRILGDAYDSRGDYDRAIE YYERALELDPDDATAWWLLGRAYTSRGDYDRAIEYYERALELD SEQ ID NO: 42 PEDEDARWVLEAARKARKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTDEAARWWYLGTAYIVRGDYDRAIEYYERALELDPNNA YAWRILGDAYDSRGDYDRATEYYERALELDPDDATAWWLLGR AYTSRGDYDRAIEYYERALELDPEDEDARWVLEAARKARKQPR GPTIKPVDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPS SEQ ID NO: 43 RSASLLAYAKLYNDEQAPKAAAVDAKFDKEQQAARAEILHLPN LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFE AQKIEWHEHHHHHHC MAQGTVDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPS RSASLLAYAKLYNDEQAPKQPRGPTIKPDEAARWDWLGWAYA ARGDYDRAIEYYERALELDPNNALAWSILGRAYAARGDYDRAIE YYERALELDPDDAFAWIDLGVAYAARGDYDRAIEYYERGLELD SEQ ID NO: 44 PEDEDARFRLEAARKARKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKGLNDIFEAQKIEWHEHHHHHHCDocket No. 1580.00230WOResidues SEQ ID MAQGTVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPS QRWRLLDEARTLNWAQAPKQPRGPTIKPDEAARWWYLGTAYIV RGDYDRAIEYYERALELDPNNAYAWRILGDAYDSRGDYDRAIE YYERALELDPDDATAWWLLGRAYTSRGDYDRAIEYYERALELD SEQ ID NO: 45 PEDEDARWVLEAARKARKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQ SLKDDP S Q S ANLL AE AKKLND AQ APKGLNDIFE A QKIEWHEHHHHHHC MAQGTDEAARWWYLGTAYIVRGDYDRAIEYYERALELDPNNA YAWRILGDAYDSRGDYDRAIEYYERALELDPDDATAWWLLGR AYTSRGDYDRAIEYYERALELDPEDEDARWVLEAARKARKQPR GPTIKPVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPSQ SEQ ID NO: 46 RWRLLDEARTLNWAQAPKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAE AKKLNDAQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPS QRWRLLDEARTLNWAQAPKQPRGPTIKPDEAARWDWLGWAYA ARGDYDRAIEYYERALELDPNNALAWSILGRAYAARGDYDRAIE YYERALELDPDDAFAWIDLGVAYAARGDYDRAIEYYERGLELD SEQ ID NO: 47 PEDEDARFRLEAARKARKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTDEAARWDWLGWAYAARGDYDRAIEYYERALELDPNN ALAWSILGRAYAARGDYDRAIEYYERALELDPDDAFAWIDLGV AYAARGDYDRAIEYYERGLELDPEDEDARFRLEAARKARKQPR GPTIKPVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPSQ SEQ ID NO: 48 RWRLLDEARTLNWAQAPKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQAPKGLNDIFEA QKIEWHEHHHHHHC MAQGTVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPS SEQ ID NO: 49 QRWRLLDEARTLNWAQAPKC VDAX1X2DX3X4X5X6X7AX8X9X10IX11X12LPNLX13X14X15QX16X17X1 8FIX19X20LX21X22DPSX23X24X25X26LLX27X28AX29X30X31NX32X33QA SEQ ID NO: 50 PK VDAX1FDX3EX5X6X7AX8X9EIX11X12LPNLNX14X15QX16X17AFIX19SSEQ ID NO: 51 LX21DDPSQSANLLAEAX29X30LNDAQAPK VDAX1X2DX3X4LEX7ARX9X10IEX12LPNLTEEQRRAFIESLRDDPSSEQ ID NO: 52 QX24X25X26LLX27EAX29X30LNX32AQAPK VDAKFDKELEEARAEIERLPNLTEX15QRRX18FIX19X20LRX22DPSXSEQ ID NO: 53 23SAX26LLAX28AX29X30X31NDX33QAPK DEAX1RWX2X3LGX4AYX5X6RGDYDRAIEYYX7RALELDPNNAX8 AWX9X10LGX11AYX12X13RGDYDRAIEYYX14RALELDPX15X16AX1SEQ ID NO: 54 7AWX18X19LGX20AYX21X22RGDYDRAIEYYX23RX24LELDPX25X26E X27ARX28X29LEX30 ARX31X32RX33 VDAKFDKEVWTAYWEIRHLPNLNVKQKEAFILSLVDDPSQSANL SEQ ID NO: 55 LAEARRLNDAQAPK VDAKFDKELHDAYWEIRKLPNLNVQQKSAFILSLVDDPSQSANL SEQ ID NO: 56LAE AKKLND AQAPKDocket No. 1580.00230WOResidues SEQ ID VDARFDREFWEAQDEIWDLPNLNWEQQHAFITSLRDDPSQSANL SEQ ID NO: 57 LAEARRLNDAQAPK VDARFDREFEDALYEINSLPNLNYRQWYAFIQSLIDDPSQSANLL SEQ ID NO: 58 AEARRLNDAQAPK VDARFDREFFEALEEIQQLPNLNYHQWWAFIQSLYDDPSQSANL SEQ ID NO: 59 LAEARRLNDAQAPK VDARFDREWEDALEEIHQLPNLNYAQWHAFIYSLIDDPSQSANL SEQ ID NO: 60 LAEARRLNDAQAPK VD AKRD QRLEYARKRIE SLPNLTEEQRRAFIE SLRDDP S QRWRLL SEQ ID NO: 61 DEARTLNWAQAPK VDAKQDQWLEFARNRIEWLPNLTEEQRRAFIESLRDDPSQKRTL SEQ ID NO: 62 LYEATVLNRAQAPK VDAKQDIWLEFARHRIERLPNLTEEQRRAFIESLRDDPSQKRRLL SEQ ID NO: 63 YEASTLNNAQAPK VDAKRDYWLEKARVKIEFLPNLTEEQRRAFIESLRDDPSQRHKL SEQ ID NO: 64 LWEAHWLNKAQAPK VDARQDFRLERARHHIESLPNLTEEQRRAFIESLRDDPSQRWHLL SEQ ID NO: 65 AEARRLNFAQAPK VDARHDHYLEWAREYIEELPNLTEEQRRAFIESLRDDPSQWVDL SEQ ID NO: 66 LNEAWFLNWAQAPK VDAKKDASLEFARGVIEWLPNLTEEQRRAFIESLRDDPSQEYWL SEQ ID NO: 67 LHEALILNHAQAPK VDAKTDQSLEWARAIIEWLPNLTEEQRRAFIESLRDDPSQEYYLL SEQ ID NO: 68 NEAYYLNNAQAPK VDAKEDADLEYARSIIEWLPNLTEEQRRAFIESLRDDPSQEYLLL SEQ ID NO: 69 LEAHDLNNAQAPK VDARTDRELEF ART VIEWLPNLTEEQRRAF IE SLRDDP S QEYFLL SEQ ID NO: 70 LEAQRLNEAQAPK VDAKFDKELEEARAEIERLPNLTERQRRYFIRKLRWDPSQSAILL SEQ ID NO: 71 AYAKRSNDLQAPK VDAKFDKELEEARAEIERLPNLTEDQRRFFIRKLRVDPSRSARLL SEQ ID NO: 72 ATAKHYNDNQAPK VD AKFDKELEEARAEIERLPNLTEYQRRYFIRKLRWDP SQ S ANLL SEQ ID NO: 73 AYAKRWNDSQAPK VDAKFDKELEEARAEIERLPNLTEWQRRWFISKLRWDPSRSAGL SEQ ID NO: 74 LAQAKAYNDDQAPK VDAKFDKELEEARAEIERLPNLTEKQRRRFISHLRWDPSNSAWLL SEQ ID NO: 75 AKAKLYNDAQAPK VDAKFDKELEEARAEIERLPNLTEHQRRYFTRKLRWDPSRSAGLL SEQ ID NO: 76 AIAKHWNDDQAPK VDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPSRSASLL SEQ ID NO: 77 AYAKLYNDEQAPK VDAKFDKELEEARAEIERLPNLTERQRRYFIRKLRWDPSQSATLL SEQ ID NO: 78 AYAKTFNDRQAPK VDAKFDKELEEARAEIERLPNLTEWQRRYFIRKLRWDPSHSARL SEQ ID NO: 79LAYAKFFNDEQAPKDocket No. 1580.00230WOResidues SEQ ID VDAKFDKELEEARAEIERLPNLTEKQRRYFIHWLRYDPSRSAKLL SEQ ID NO: 80 AHAKSKNDRQAPK VD AKFDKELEE ARAEIERLPNLTE S QRRYFIW ALRLDP SK S AHLL SEQ ID NO: 81 ARAKWRNDVQAPK VDAKFDKELEEARAEIERLPNLTEWQRRRFIWSLRQDPSESAHLL SEQ ID NO: 82 ASAKRKNDWQAPK VD AKFDKELEE ARAEIERLPNLTE S QRRDFIY SLRQDP SH S AHLL SEQ ID NO: 83 AF AKWHNDK Q APK VDAKFDKELEEARAEIERLPNLTEAQRRDFIWALRSDPSESAKLL SEQ ID NO: 84 AFAKWHNDRQAPK VDAKFDKELEEARAEIERLPNLTEWQRRDFIYSLRFDPSSSALLL SEQ ID NO: 85 ALAKWHNDEQAPK VDAKFDKELEEARAEIERLPNLTEWQRRDFIWSLRLDPSASAQLL SEQ ID NO: 86 AFAKWHNDNQAPK VD AKFDKELEEARAEIERLPNLTEYQRRDFIHALRVDP S S S AHLL SEQ ID NO: 87 AFAKWHNDRQAPK VD AKFDKELEE ARAEIERLPNLTEDQRRGFIYALRIDPS AS AALL SEQ ID NO: 88 AAAKHDNDRQAPK VDAKFDKELEEARAEIERLPNLTEDQRRGFIHSLRVDPSHSADLL SEQ ID NO: 89 AFARWLNDDQAPK VDAKFDKELAHAYWEIRHLPNLNLKQRGAFIRSLADDPSQSANL SEQ ID NO: 90 LAEAKKLNDAQAPK VDAKFDKEIFQAYWEIRHLPNLNNHQKTAFIHSLVDDPSQSANL SEQ ID NO: 91 LAEAKKLNDAQAPK DEAGRWQWLGEAYLQRGD YDRAIE YYQRALELDPNNATAWIA LGDAYLYRGDYDRAIEYYQRALELDPNNAWAWGALGEAYAAR SEQ ID NO: 92 GDYDRAIEYYQRALELDPNNERARNVLEWARRSRR DEAAR WDDLGFAYVSRGDYDRAIEYYERALELDPNNAAAWVL LGEAYAVRGDYDRAIEYYERALELDPDDAHAWLHLGSAYTIRG SEQ ID NO: 93 DYDRAIEYYERALELDPEDEAARIYLEDARKARK DEAARWDWLGWAYAARGDYDRAIEYYERALELDPNNALAWSI LGRAYAARGDYDRAIEYYERALELDPDDAFAWIDLGVAYAARG SEQ ID NO: 94 DYDRAIEYYERGLELDPEDEDARFRLEAARKARK DEAARWWYLGTAY1VRGDYDRA1EYYERALELDPNNAYAWR1LGD A YD SRGD YDRAIE YYERALELDPDD AT AWWLLGRA YT SRG SEQ ID NO: 95 DYDRAIEYYERALELDPEDEDARWVLEAARKARK VDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPSRSASLL AYAKLYNDEQAPKQPRGPTIKPDEAARWWYLGTAYIVRGDYDR AIE YYERALELDPNNA YAWRILGD A YD SRGD YDRAIE YYERALE SEQ ID NO: 96 LDPDDATAWWLLGRAYTSRGD YDRAIE YYERALELDPEDED AR WVLEAARKARK DEAARWWYLGTAYIVRGD YDRAIE YYERALELDPNNA YAWRIL GDAYD SRGD YDRAIE YYERALELDPDD AT AWWLLGRA YT SRG DYDRAIE YYERALELDPEDED ARWVLEAARKARKQPRGPTIKPV SEQ ID NO: 97 DAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPSRSASLLAYAKLYNDEQAPKDocket No. 1580.00230WOResidues SEQ ID VDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPSRSASLL AYAKLYNDEQAPKQPRGPTIKPDEAARWDWLGWAYAARGDYD RAIEYYERALELDPNNALAWSILGRAYAARGDYDRAIEYYERAL SEQ ID NO: 98 ELDPDDAFAWIDLGV AY AARGD YDRAIE YYERGLELDPEDEDA RFRLEAARKARK VD AKRD QRLE Y ARKRIE SLPNLTEEQRRAFIE SLRDDP S QRWRLL DEARTLNWAQAPKQPRGPTIKPDEAARWWYLGTAYIVRGDYDR AIEYYERALELDPNNAYAWRILGDAYDSRGDYDRAIEYYERALE SEQ ID NO: 99 LDPDDATAWWLLGRAYTSRGDYDRAIEYYERALELDPEDEDAR WVLEAARKARK DEAARWWYLGTAYIVRGDYDRAIEYYERALELDPNNAYAWRIL GD A YD SRGD YDRAIE YYERALELDPDD AT AWWLLGRA YT SRG DYDRAIEYYERALELDPEDEDARWVLEAARKARKQPRGPTIKPV SEQ ID NO: 100 DAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPSQRWRLLD EARTLNWAQAPK VD AKRD QRLEYARKRIE SLPNLTEEQRRAFIE SLRDDP S QRWRLL DEARTLNWAQAPKQPRGPTIKPDEAARWDWLGWAYAARGDYD RAIEYYERALELDPNNALAWSILGRAY AAR GD YDRAIE YYERAL SEQ ID NO: 101 ELDPDDAFAWIDLGV AY AARGD YDRAIEYYERGLELDPEDEDA RFRLEAARKARK DEAARWDWLGW AY AARGD YDRAIE YYERALELDPNN AL AWSI LGRAYAARGD YDRAIE YYERALELDPDD AFAWIDLGV A YAARG DYDRAIEYYERGLELDPEDEDARFRLEAARKARKQPRGPTIKPV SEQ ID NO: 102 DAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPSQRWRLLD EARTLNWAQAPK MAQGTVDAKFDKEVWTAYWEIRHLPNLNVKQKEAFILSLVDDP SQSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 103 NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELHDAYWEIRKLPNLNVQQKSAFILSLVDDPS QSANLLAEAKKLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 104 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDARFDREFWEAQDEIWDLPNLNWEQQHAFITSLRDDP SQSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 105 NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDARFDREFEDALYEINSLPNLNYRQWYAFIQSLIDDPS QSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 106 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDARFDREFFEALEEIQQLPNLNYHQWWAFIQSLYDDPS QSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 107 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDARFDREWEDALEEIHQLPNLNYAQWHAFIYSLIDDPS QSANLLAEARRLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 108 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPS QRWRLLDEARTLNWAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 109NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKCDocket No. 1580.00230WOResidues SEQ ID MAQGTVDAKQDQWLEFARNRIEWLPNLTEEQRRAFIESLRDDPS QKRTLLYEATVLNRAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 110 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKQDIWLEFARHRIERLPNLTEEQRRAFIESLRDDPSQ KRRLLYEASTLNNAQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 111 TEEQRNKFIQSLKDDPSQSANLLAE AKKLNDAQAPKC MAQGTVDAKRDYWLEKARVKIEFLPNLTEEQRRAFIESLRDDPS QRHKLLWEAHWLNKAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 112 NLTEEQRNKFIQSLKDDPSQSANLLAE AKKLNDAQAPKC MAQGTVDARQDFRLERARHHIESLPNLTEEQRRAFIESLRDDPSQ RWHLLAEARRLNFAQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 113 TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDARHDHYLEWAREYIEELPNLTEEQRRAFIESLRDDPS QWVDLLNEAWFLNWAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 114 NLTEEQRNKFIQSLKDDPSQSANLLAE AKKLNDAQAPKC MAQGTVDAKKDASLEFARGVIEWLPNLTEEQRRAFIESLRDDPS QEYWLLHEALILNHAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 115 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKTDQSLEWARAIIEWLPNLTEEQRRAFIESLRDDPS QEYYLLNEAYYLNNAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 116 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKEDADLEYARSIIEWLPNLTEEQRRAFIESLRDDPSQ EYLLLLEAHDLNNAQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 117 TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDARTDRELEFARTVIEWLPNLTEEQRRAFIESLRDDPSQ EYFLLLEAQRLNEAQAPKAAAVDAKFDKEQQAARAEILHLPNLT SEQ ID NO: 118 EEQRNKFIQ SLKDDP SQ S ANLL AE AKKLND AQ APKC MAQGTVDAKFDKELEEARAEIERLPNLTERQRRYFIRKLRWDPS QSAILLAYAKRSNDLQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 119 TEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQ APKC MAQGTVDAKFDKELEEARAEIERLPNLTEDQRRFFIRKLRVDPSR SARLLATAKHYNDNQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 120 TEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQ APKC MAQGTVDAKFDKELEEARAEIERLPNLTEYQRRYFIRKLRWDPS QSANLLAYAKRWNDSQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 121 LTEEQRNKFIQSLKDDPSQSANLLAE AKKLND AQ APKC MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRWFISKLRWDPS RSAGLLAQAKAYNDDQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 122 LTEEQRNKFIQSLKDDPSQSANLLAE AKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEKQRRRFISHLRWDPS NSAWLLAKAKLYNDAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 123 NLTEEQRNKFIQSLKDDPSQSANLLAE AKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEHQRRYFIRKLRWDPS RSAGLLAIAKHWNDDQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 124LTEEQRNKFIQSLKDDPSQSANLLAE AKKLNDAQAPKCDocket No. 1580.00230WOResidues SEQ ID MAQGTVDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPS RSASLLAYAKLYNDEQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 125 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTERQRRYFIRKLRWDPS QSATLLAYAKTFNDRQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 126 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRYFIRKLRWDPS HSARLLAYAKFFNDEQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 127 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEKQRRYFIHWLRYDPS RSAKLLAHAKSKNDRQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 128 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTESQRRYFIWALRLDPS KSAHLLARAKWRNDVQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 129 NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRRFIWSLRQDPS ESAHLLASAKRKNDWQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 130 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTESQRRDFIYSLRQDPSH SAHLLAFAKWHNDKQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 131 TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEAQRRDFIWALRSDPS ESAKLLAFAKWHNDRQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 132 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRDFIYSLRFDPSS SALLLALAKWHNDEQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 133 TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEWQRRDFIWSLRLDPS ASAQLLAFAKWHNDNQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 134 NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEYQRRDFIHALRVDPS SSAHLLAFAKWHNDRQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 135 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEDQRRGFIYALRIDPSA SAALLAAAKHDNDRQAPKAAAVDAKFDKEQQAARAEILHLPNL SEQ ID NO: 136 TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEDQRRGFIHSLRVDPS HSADLLAFARWLNDDQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 137 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELAHAYWEIRHLPNLNLKQRGAFIRSLADDP SQSANLLAEAKKLNDAQAPKAAAVDAKFDKEQQAARAEILHLP SEQ ID NO: 138 NLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKEIFQAYWEIRHLPNLNNHQKTAFIHSLVDDPS QSANLLAEAKKLNDAQAPKAAAVDAKFDKEQQAARAEILHLPN SEQ ID NO: 139 LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQAAAGDEAGRWQWLGEAYLQRGDYDRAIEYYQRALELDPN SEQ ID NO: 140NATAWIALGDAYLYRGDYDRAIEYYQRALELDPNNAWAWGALDocket No. 1580.00230WOResidues SEQ ID GEAYAARGDYDRAIEYYQRALELDPNNERARNVLEWARRSRRA AAVDAKFDKEQQAARAEILHLPNLTEEQRNKFIQSLKDDPSQSA NLLAEAKKLNDAQAPKC MADEAARWDDLGFAYVSRGDYDRAIEYYERALELDPNNAAAW VLLGEAYAVRGDYDRAIEYYERALELDPDDAHAWLHLGSAYTI RGDYDRAIEYYERALELDPEDEAARIYLEDARKARKAAAVDAK SEQ ID NO: 141 FDKEQQAARAEILHLPNLTEEQRNKFIQSLKDDPSQSANLLAEAK KLNDAQAPKC MADEAARWDWLGWAYAARGDYDRAIEYYERALELDPNNALA WSIL GRAY AARGDYDRAIEYYERALELDPDDAFAWIDLGV AYA ARGDYDRAIEYYERGLELDPEDEDARFRLEAARKARKAAAVDA SEQ ID NO: 142 KFDKEQQAARAEILHLPNLTEEQRNKFIQSLKDDPSQSANLLAEA KKLNDAQAPKC MADEAARWWYLGTAYIVRGDYDRAIEYYERALELDPNNAYAW RILGDAYDSRGDYDRAIEYYERALELDPDDATAWWLLGRAYTS RGDYDRAIEYYERALELDPEDEDARWVLEAARKARKAAAVDA SEQ ID NO: 143 KFDKEQQAARAEILHLPNLTEEQRNKFIQSLKDDPSQSANLLAEA KKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPS RSASLLAYAKLYNDEQAPKQPRGPTIKPDEAARWWYLGTAYIV RGDYDRAIEYYERALELDPNNAYAWRILGDAYDSRGDYDRAIE SEQ ID NO: 144 YYERALELDPDDATAWWLLGRAYTSRGDYDRAIEYYERALELD PEDEDARWVLEAARKARKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTDEAARWWYLGTAYIVRGDYDRAIEYYERALELDPNNA YAWRILGDAYDSRGDYDRAIEYYERALELDPDDATAWWLLGR AYTSRGDYDRAIEYYERALELDPEDEDARWVLEAARKARKQPR SEQ ID NO: 145 GPTIKPVDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPS RSASLLAYAKLYNDEQAPKAAAVDAKFDKEQQAARAEILHLPN LTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKFDKELEEARAEIERLPNLTEQQRRWFIRKLRWDPS RSASLLAYAKLYNDEQAPKQPRGPTIKPDEAARWDWLGWAYA ARGDYDRAIEYYERALELDPNNALAWSILGRAYAARGDYDRAIE SEQ ID NO: 146 YYERALELDPDDAFAWIDLGVAYAARGDYDRAIEYYERGLELD PEDEDARFRLEAARKARKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPS QRWRLLDEARTLNWAQAPKQPRGPTIKPDEAARWWYLGTAYIV RGDYDRAIEYYERALELDPNNAYAWRILGDAYDSRGDYDRAIE SEQ ID NO: 147 YYERALELDPDDATAWWLLGRAYTSRGDYDRAIEYYERALELD PEDEDARWVLEAARKARKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTDEAARWWYLGTAYIVRGDYDRAIEYYERALELDPNNA YAWRILGDAYDSRGDYDRAIEYYERALELDPDDATAWWLLGR SEQ ID NO: 148 AYTSRGDYDRAIEYYERALELDPEDEDARWVLEAARKARKQPRGPTIKPVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPSQDocket No. 1580.00230WOResidues SEQ ID RWRLLDEARTLNWAQAPKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPS QRWRLLDEARTLNWAQAPKQPRGPTIKPDEAARWDWLGWAYA ARGDYDRAIEYYERALELDPNNALAWSILGRAYAARGDYDRAIE SEQ ID NO: 149 YYERALELDPDDAFAWIDLGVAYAARGDYDRAIEYYERGLELD PEDEDARFRLEAARKARKAAAVDAKFDKEQQAARAEILHLPNL TEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC MAQGTDEAARWDWLGWAYAARGDYDRAIEYYERALELDPNN ALAWSILGRAYAARGDYDRAIEYYERALELDPDDAFAWIDLGV AYAARGDYDRAIEYYERGLELDPEDEDARFRLEAARKARKQPR SEQ ID NO: 150 GPTIKPVDAKRDQRLEYARKRIESLPNLTEEQRRAFIESLRDDPSQ RWRLLDEARTLNWAQAPKAAAVDAKFDKEQQAARAEILHLPNLTEEQRNKFIQSLKDDPSQSANLLAEAKKLNDAQAPKC

Claims

Docket No. 1580.00230WOCLAIMSWhat is claimed is:

1. A human herpesvirus affinity ligand of Formula IL1-D1-L2-D2-L3-DE-DT-L4 (Formula I)whereinL1, L2, L3, and L4 are each independently absent or a linker;Di and D2 are each independently an antigen-binding domain and D2 is optional;DE is an optional structural domain; andDT is an optional C-terminal tag domain,wherein at least one of D1 or D2 comprises an amino acid sequence of Formula 1:VDAX1X2DX3X4X5X6X7AX8X9X10IX11X12LPNLX13X14X15QX16X17X18FIX19X20LX21X2 2DPSX23X24X25X26LLX27X28AX29X30X31NX32X33QAPK (SEQ ID NO: 50)whereinX1 is R or K; X18 is A, G, R, Y, D, F, W;X2is F, R, Q, T, H, K, E; Xi9is R, S, H, Y, W, E, L, Q, T;X3is R, K, H, A, Q, Y, I, F; X20 is S, A, K, H, W;X4is E, Y, D, S, W, R; X21 is R, Y, A, 1, V;X5is L, V, I, F, W; X22is W, V, Y, I, F, L, Q, S, D;X6 is W, F, E, A, H; X23 is Q, A, S, H, K, E, R, N;X7 is E, Y, R, F, K, W, T, D, Q, H; X24 is S, R, K, E, W;Xsis Q, L, Y, R; X25 is A, V, Y, H, R, W;X9is A, K, H, N, V, G, T, S, E, W, Y, D; X26 is N, D, L, Y, F, W, K, R, T, H, Q, A, G, S, I;Xiois E, Y, I, V, K, R, H; X27is A, N, L, H, W, Y, D;Xn is E, R, N, H, Q, W; X28 is E, F, S, R, L, A, H, K, T, I, Q, Y; Xi2is D, Q, S, H, K, E, W, F, R; X29 is R, K, W, H, Y, Q, L, S, T;X13 is N or T; X30 is R, K, F, D, Y, I, W, T, V, H, S, L, A;X14 is E, V, N, L, Y, W; X3iis L, K, R, H, D, Y, W, F, S;X15 is R, Y, W, Q, H, D, K, S, A, E; X32 is D, W, N, E, H, K, R, F; and X16 is R, K, W, Q; X33 is A, D, W, V, R, K, N, E, S, L.Docket No. 1580.00230WOX17 is H, W, Y, G, S, T, E, R;.

2. The affinity ligand of claim 1, wherein at least one of D1 or D2 comprises an amino acid sequence of Formula la:VDAX1FDX3EX5X6X7AX8X9EIX11X12LPNLNX14X15QX16X17AFIX19SLX21DDPSQSA NLLAEAX29X30LNDAQAPK (SEQ ID NO: 51)whereinXi is R or K;X3 is R or K;X5 is L, V, I, F, W;X6is W, F, E, A, H;X7is E, T, D, Q, H;X8is Q, L, Y;X$>is E, W, Y, D;Xn is R, N, H, Q, W;Xi2is D, Q, S, H, K;X14 is V, N, L, Y, W;X15 is R, Q, H, K, S, A, E;Xi6 is R, K, W, Q;X17 is H, W, Y, G, S, T, E;Xi9is R, H, Y, L, Q, T;X21 is R, Y, A, I, V;X29 is R or K; andX30 is R or K.

3. The affinity ligand of claim 2, wherein at least one of D1 or D2 comprises the amino acid sequence of any one of SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60, SEQ ID NO: 90, or SEQ ID NO: 91.

4. The affinity ligand of claim 3, wherein the affinity ligand comprises or consists of an amino acid sequence of SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 138 or SEQ ID NO: 139, or an amino acid sequence having at least 94% identity thereto.Docket No. 1580.00230WO5. The affinity ligand of any one of claims 2 to 4, whereinXi is R or K;X3 is R or K;X5 is L or V;Xe is A;X7is H;Xs is Y;X9is W;Xu is R;Xi2is H;X14 is V or L;X15 is K;Xi6 is R;X17 is G or S;X19 is R;X21 is A or V;X29 is R or K; andX30 is R or K.

6. The affinity ligand of claim 1, wherein at least one of D1 or D2 comprises an amino acid sequence of Formula lb:VDAXiX2DX3X4LEX7ARX9XioIEXi2LPNLTEEQRRAFIESLRDDPSQX24X25X26LLX27EAX29X30LNX32AQAPK (SEQ ID NO: 52)whereinXi is R or K;X2is R, Q, T, H, K, E;X3 is R, H, A, Q, Y, I, F;X4is E, Y, D, S, W, R;X7is Y, R, F, K, W;X9is A, K, H, N, V, G, T, S, E;Xiois Y, I, V, K, R, H;Xi2is S, E, W, F, R;X24is R, K, E, W;Docket No. 1580.00230WOX25 is V, Y, H, R, W;X26 is D, L, Y, F, W, K, R, T, H;X27is A, N, L, H, W, Y, D;X29is R, W, H, Y, Q, L, S, T;X30 is R, F, D, Y, R, I, W, T, V; andX32 is W, N, E, H, K, R, F.

7. The affinity ligand of claim 6, wherein at least one of D1 or D2 comprises an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, or SEQ ID NO: 70, or an amino acid sequence having at least 94% identity thereto.

8. The affinity ligand of claim 6, wherein the affinity ligand comprises or consists of an amino acid sequence of SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, or an amino acid sequence having at least 94% identity thereto.

9. The affinity ligand of any one of claims 6 to 8, whereinXi is R or K;X2 is R or K;X3is Q;X4is R;X7is Y;X9is K;X10is K or R;Xi2is S;X24 is K or R;X25is W;X26 is R;X27is D;X29 is R;X30 is T; andX32 is W.Docket No. 1580.00230WO10. The affinity ligand of claim 1, wherein at least one of D1 or D2 comprises an amino acid sequence of Formula 2:DEAX1RWX2X3LGX4AYX5X6RGDYDRAIEYYX7RALELDPNNAX8AWX9X10LGX11 AYX12X13RGDYDRAIEYYX14RALELDPX15X16AX17AWX18X19LGX20AYX21X22RGDYDRA IEYYX23RX24LELDPX25X26EX27ARX28X29LEX30ARX31X32RX33 (SEQ ID NO: 54) whereinXi is A or G; X18is I, W, L, or G;X2is D, W, or Q; X19 is D, L, H, or A;X3 is D, W, or Y; X20 is V, R, S, or E;X4 is W, T, F, or E; X21 is A or T;Xs is A, I, V, or L; X22 is A, S, or I;X6 is A, V, S, or Q; X23 is E or Q;X7 is E or Q; X24 is A or G;X8 is L, Y, A, or T; X25 is E or N;X9is S, R, V, or I; X26 is D or N;X10 is I, L, or A; X27 is D, A, or R;X11is R, D, or E; X28is F, W, I, orN;X12 is A, D, or L; X29 is R, V, or Y;X13 is A, S, V or Y; X30 is A, D, or W;X14 is Q or E; X31 is K or R;X15 is N or D; X32 is A or S; andX16 is N or D; X33 is K or R.X17 is F, T, H, or W;11. The affinity ligand of claim 10, whereinXi is A or G; X18is I, W;X2is D, W; Xi9is D, L;X3is W, or Y; X2ois V, R;X4 is W, T,; X21 is A or T;Xs is A, I,; X22 is A, S;X6 is A, V; X23 is E;X7is E; X24 is A or G;X8is L, Y; X25 is E;Docket No. 1580.00230WOX9 is S, R; X26 is D;X10is I; X27 is D;X11 is R, D; X28 is F, W;X12 is A, D; X29 is R, V;X13 is A, S; X30 is A;X14 is E; X31 is K;X15 is D; X32 is A; andXi6 is D; X33 is K.X17 is F, T;12. The affinity ligand of claim 10, wherein at least one of D1 or D2 comprises an amino acid sequence of any one of SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94 or SEQ ID NO: 95, or an amino acid sequence having at least 94% identity thereto.

13. The affinity ligand of claim 10 or 12, wherein the affinity ligand comprises or consists of an amino acid sequence of any one of SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142 or SEQ ID NO: 143, or an amino acid sequence having at least 94% identity thereto.

14. The affinity ligand of any one of claims 1 to 13, wherein D2 is absent.

15. The affinity ligand of any one of claims 1 to 13, whereinLi is a linker of from 1 -6 amino acids;L2, D2 and DT are absent;L3 is absent or a linker of from 1-12 amino acids;DE is present; andL4 is absent or a linker comprising from 1-15 amino acids.

16. The affinity ligand of claim 1, wherein the ligand comprises two antigen-binding domains, Di and D2 and each of Di and D2 is independently defined by the amino acid sequence of SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53 or SEQ ID NO: 54.

17. The affinity ligand of claim 16, wherein each of Di and D2 independently comprises an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 90, SEQ ID NO: 94 or SEQ ID NO: 95.Docket No. 1580.00230WO18. The affinity ligand of claim 16, wherein one of Di and D2 comprises or consists of an amino acid sequence of SEQ ID NO: 77 or SEQ ID NO: 90 and the remainder comprises an amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95.

19. The affinity ligand of claim 18, wherein the affinity ligand comprises or consists of an amino acid sequence of SEQ ID NO: 42, SEQ ID NO: 43, or SEQ ID NO: 44.

20. The affinity ligand of claim 16, wherein one of Di and D2 comprises an amino acid sequence of SEQ ID NO: 61 and the remainder comprises an amino acid sequence of SEQ ID NO: 94 or SEQ ID NO: 95.

21. The affinity ligand of claim 20, wherein the affinity ligand comprises or consists of an amino acid sequence of any one of SEQ ID NO: 45, SEQ ID NO: 46, or SEQ ID NO: 47.

22. The affinity ligand of claim 1, wherein at least one of D1 or D2 comprises or consists of an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 90, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97, or an amino acid sequence having at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity thereto.

23. The affinity ligand of claim 1, wherein Di and D2 each independently comprises an amino acid sequence of any one of SEQ ID NO: 61, SEQ ID NO: 77, SEQ ID NO: 90, SEQ ID NO: 94, SEQ ID NO: 95, or SEQ ID NO: 97, or an amino acid sequence having at least 94% identity thereto.

24. The affinity ligand of claim 1, wherein the affinity ligand comprises or consists of an amino acid sequence of any one of SEQ ID NO: 109, SEQ ID NO: 125, SEQ ID NO: 138, SEQ ID NO: 142 or SEQ ID NO: 143, or an amino acid sequence having at least 94% identity thereto.

25. The affinity ligand of any one of claims 1 to 24, wherein DT, when present, comprises one or more peptide, polypeptide or protein tags selected from the group consisting of a bacteriophage T7 epitope (T7-tag), bacteriophage V5 epitope (V5-tag), biotin-carboxy carrier protein (BCCP), polyhistidine (His-tag), polyaspartate (Asp-tag), polycysteine (Cys-tag), polyphenylalanine (Phe-tag), glutathione S-transferase (GST), maltose binding protein (MBP), calmodulin binding peptide (CBP), intein-chitin binding domain (intein-CBD), streptavadin, streptavadin-binding peptide (SBP), Strep-tag, Protein A of Staphylococcus aureus andDocket No. 1580.00230WOderivatives thereof, calmodulin-binding peptide (CBP), FLAG tag, human influenza hemagglutinin (HA), c-myc epitope, Glu-Glu, HSV epitope, and green fluorescent protein (GFP) and derivatives thereof.

26. The affinity ligand of any one of claims 1 to 25, wherein the affinity ligand comprises a C-terminal thiol containing amino acid residue, optionally a cysteine or threonine residue.

27. The affinity ligand of any one of claims 1 to 26, wherein the affinity ligand binds to a human herpesvirus antigen selected from the group consisting of envelope glycoprotein H (gH), gL, gB, gC, gK, gG, gJ, gD, gl, gE, glycoprotein pV (gpV), gpll, gpIII, gpIV, gpl, gp350 / 220, gp35, gp85, gp42, gpl 10, got, lEgp, and gp47 / 52, or a fragment or derivative of any of the foregoing.

28. The affinity ligand of any one of claims 1 to 27, wherein the affinity ligand binds to a human herpesvirus antigen selected from the group consisting of capsid scaffolding protein (BVRF2 / BdRFl), capsid vertex component 1 (CVC1, BGLF1), capsid vertex component 2 (CVC2, BVRF1), envelope glycoprotein B (gB, BALF4), envelope glycoprotein GP350 (BLLF1), envelope glycoprotein H (gH, BXLF2), envelope glycoprotein L (gL, BKRF2), envelope glycoprotein M (gM, BBRF3), envelope glycoprotein N (gN, BLRF1), glycoprotein 42 (BZLF2), glycoprotein BDLF3 (BDLF3), glycoprotein BILF2 (BILF2), G-protein coupled receptor BILF1 (BILF1), latent membrane protein 1 (LMP1, BNLF1), latent membrane protein 2 (LMP2), major capsid protein (MCP, BcLF1), major tegument protein (BNRF1), mRNA export factor ICP27 homolog (BMLF1, BSLF2), protein BDLF2 (BDLF2), protein BMRF2 (BMRF2), protein BNLF2a (BNLF2a), protein LF2 (LF2), protein UL24 homolog (BXRF1), putative BARFO protein (BARFO), secreted protein BARF1 (BARF1), triplex capsid protein 1 (TRX1, BORF1), and triplex capsid protein 2 (TRX2, BDLF1), or a fragment or derivative of any of the foregoing.

29. The affinity ligand of claim 28, wherein the antigen is a polypeptide or protein selected from the group consisting of glycoprotein gp42, glycoprotein gp35O, envelope glycoprotein H (gH), envelope glycoprotein L (gL), and latent membrane protein 2 (LMP2), or a fragment or derivative of any of the foregoing.

30. An affinity chromatography resin comprising a solid support and a plurality of the affinity ligands of any one of claims 1 to 29 covalently attached to the solid support.Docket No. 1580.00230WO31. The affinity chromatography resin of claim 30, wherein the solid support is in the form of discrete polymeric particles, ceramic or glass beads, a woven or non-woven membrane, or a polymeric monolith.

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