Enzyme variants and uses thereof

Abstract

The present disclosure relates generally to polypeptides capable of hydrolysing an amide bond in a polyamide, and more specifically to polypeptides comprising an amino acid sequence of amino acid residues 2-398 of SEQ ID NO: 2 or an amino acid sequence that has at least 70% sequence identity thereto; or polypeptides comprising an amino acid sequence of amino acid residues 2-394 of SEQ ID NO:88 or an amino acid sequence that has at least 75% sequence identity thereto. The present disclosure also extends to methods of using said polypeptides. In another embodiment, the polypeptides disclosed are capable of hydrolysing the nylon 6,6 oligomer to produce adipic acid and hexamethylenediamine.

Description

ENZYME VARIANTS AND USES THEREOF

[0001] The present invention relates to novel synthetic enzymes, more particularly to enzymes that catalyse the hydrolysis of nylon polymers, and uses thereof.BACKGROUND

[0002] All references, including any patent or patent application cited in this specification are hereby incorporated by reference to enable full understanding of the invention. Nevertheless, such references are not to be read as constituting an admission that any of these documents forms part of the common general knowledge in the art, in Australia or in any other country.

[0003] Global industrialization has had significant environmental impact, not least of which is an increase in the manufacture and reliance on plastic and plastic products. These plastics include a range of synthetic or semi-synthetic polymers, such as polyethylene terephthalate, PVC, polypropylene, polystyrene and polyamides.

[0004] Polyamide (also known generically as nylon) is a silk-like thermoplastic that can be melt-processed into fibres, films or shapes, including in combination with a variety of additives. Polyamides have many significant commercial applications, such as in clothing, the production of resins and molded shapes, in food packaging and industrial fibre and rope.

[0005] Whilst there is a growing effort to find suitable and environmentally sustainable alternatives to plastics, including nylon, including their manufacture and disposal, such products remain a significant problem and contribute to a vast majority of environmental pollutants. The environmental significance of this problem is attributed, at least in part, to the chemical nature of plastics, as they do not readily decompose in nature. Approaches to deal with the problem of plastic waste products have typically included incineration, disposal in landfill and mechanical disintegration. However, these approaches also have significant environmental impact. For instance, incineration of plastics produces potentially harmful byproducts that are released into the atmosphere; the decomposition rate of plastics in landfill is typically very slow and there is a risk that toxic materials will leach into groundwater; and mechanical disintegration is relatively expensive and inefficient and there is often limited use for its byproducts.

[0006] Nylon can take many decades to decompose, and while there are ways to recycle nylon, these methods require energy-intensive grinding and re-melting of the polymer. Depending on the nylon and the intended applications, the recycled nylon polymer may need to be mixed with virgin nylon, limiting its flexibility and sustainability.

[0007] More recently, biological (enzymatic) degradation of plastics has been considered as an alternative approach to reducing plastic waste accumulation. In the case of nylon, a strain of bacterium found in the waste water of a nylon factory was discovered to have unique enzymes (NylA, NylB and NylC) that could digest by-products of nylon 6 (a nylon homopolymer of caprolactam) manufacture.

[0008] Whilst enzymatic degradation of plastics such as nylon is an attractive alternative to alleviating the environmental impact of plastic waste products and their disposal, it has not yet seen widespread adoption, including because of its relative inefficiency, slow rate of enzymatic degradation and low levels of enzyme expression in common industrial host organisms. Hence, there remains an urgent need for improved methods and reagents for the enzymatic degradation of plastics, including nylon.SUMMARY OF THE INVENTION

[0009] In one aspect disclosed herein, there is provided a polypeptide capable of hydrolysing an amide bond in a polyamide, wherein the polypeptide comprises an amino acid sequence of amino acid residues 2-398 of SEQ ID NO: 2 or an amino acid sequence that has at least 70% sequence identity thereto. In one embodiment, the polypeptide comprises an amino acid sequence that has at least 75% sequence identity of amino acid residues 2-398 of SEQ ID NO: 2. In one embodiment, the polypeptide comprises an amino acid sequence that has at least 80% sequence identity of amino acid residues 2-398 of SEQ ID NO: 2. In one embodiment, the polypeptide comprises an amino acid sequence that has at least 85% sequence identity of amino acid residues 2-398 of SEQ ID NO: 2. In one embodiment, the polypeptide comprises an amino acid sequence that has at least 90% sequence identity of amino acid residues 2-398 of SEQ ID NO: 2. In one embodiment, the polypeptide comprises an amino acid sequence that has at least 95% sequence identity of amino acid residues 2-398 of SEQ ID NO: 2.

[0010] In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 125, or an amino acid sequence that has at least 80% sequence identity thereto

[0011] In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO:96, or an amino acid sequence that has at least 80% sequence identity thereto.

[0012] In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO:1, or an amino acid sequence that has at least 80% sequence identity thereto.

[0013] In one embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 125, or an amino acid sequence that has at least 80% sequence identity thereto comprises an amino acid sequence wherein i. the amino acid at position 3 is Q or T; ii. the amino acid at position 5 is N or D; iii. the amino acid at position 7 is M or F; iv. the amino acid at position 9 is G or T; v. the amino acid at position 10 is S or F; vi. the amino acid at position 14 is P or A; vii. the amino acid at position 16 is Q or G viii. the amino acid at position 17 is Q or E; ix. the amino acid at position 25 is Q or T; x. the amino acid at position 26 is A or P; xi. the amino acid at position 29 is N or S; xii. the amino acid at position 32 is A or S; xiii. the amino acid at position 39 is I or L; xiv. the amino acid at position 40 is I or V; xv. the amino acid at position 44 is N or Q; xvi. the amino acid at position 48 is G or A xvii. the amino acid at position 51 is A or P; xviii. the amino acid at position 52 is A or V; xix. the amino acid at position 53 is S or W;xx. the amino acid at position 57 is A or R; xxi. the amino acid at position 58 is A or S; xxii. the amino acid at position 60 is R or A; xxiii. the amino acid at position 61 is D, A or G; xxiv. the amino acid at position 63 is D or G; xxv. the amino acid at position 64 is G, E or R; xxvi. the amino acid at position 67 is F or L xxvii. the amino acid at position 70 is P or deleted; xxviii. the amino acid at position 72 is G or A; xxix. the amino acid at position 73 is R or K; xxx. the amino acid at position 74 is S or E; xxxi. the amino acid at position 75 is T, M, G or W; xxxii. the amino acid at position 78 is A or G; xxxiii. the amino acid at position 82 is A or E; xxxiv. the amino acid at position 84 is S or T; xxxv. the amino acid at position 88 is A or G; xxxvi. the amino acid at position 92 is L or M; xxxvii. the amino acid at position 99 is A or S; xxxviii. the amino acid at position 101 is W or H; xxxix. the amino acid at position 110 is S or T; xl. the amino acid at position 114 is L or V; xli. the amino acid at position 123 is G or A; xlii. the amino acid at position 124 is I or T; xliii. the amino acid at position 131 is D or E; xliv. the amino acid at position 132 is R or D; xlv. the amino acid at position 138 is D or A; xlvi. the amino acid at position 143 is H, R or D; xlvii. the amino acid at position 148 is A or V; xlviii. the amino acid at position 149 is A or K; xlix. the amino acid at position 155 is D or G;1. the amino acid at position 167 is S or G; li. the amino acid at position 171 is E or D; lii. the amino acid at position 173 is D or A; liii. the amino acid at position 178 is D or A;liv. the amino acid at position 179 is G or S;Iv. the amino acid at position 180 is D or A;Ivi. the amino acid at position 186 is R, E or Q;Ivii. the amino acid at position 188 is T or M;Iviii. the amino acid at position 193 is A or P; lix. the amino acid at position 194 is S or P; lx. the amino acid at position 195 is D or P;Ixi. the amino acid at position 197 is A or E;Ixii. the amino acid at position 203 is R or Y;Ixiii. the amino acid at position 206 is L or I;Ixiv. the amino acid at position 207 is A or V;Ixv. the amino acid at position 210 is R or K;Ixvi. the amino acid at position 211 is R, K or deleted;Ixvii. the amino acid at position 212 is S, D or G;Ixviii. the amino acid at position 213 is D or A;Ixix. the amino acid at position 215 is P or E;Ixx. the amino acid at position 223 is A or V;Ixxi. the amino acid at position 234 is L or I;Ixxii. the amino acid at position 244 is D or E;Ixxiii. the amino acid at position 249 is H or R;Ixxiv. the amino acid at position 252 is Q or R;Ixxv. the amino acid at position 258 is H or A;Ixxvi. the amino acid at position 262 is I or V;Ixxvii. the amino acid at position 276 is L or I;Ixxviii. the amino acid at position 278 is A or C;Ixxix. the amino acid at position 280 is L, A or P;Ixxx. the amino acid at position 288 is Q or E;Ixxxi. the amino acid at position 291 is L or R;Ixxxii. the amino acid at position 301 is I or V;Ixxxiii. the amino acid at position 304 is E, A or G;Ixxxiv. the amino acid at position 305 is A or W;Ixxxv. the amino acid at position 308 is A or D;Ixxxvi. the amino acid at position 311 is L or R;Ixxxvii. the amino acid at position 312 is Q or T;Ixxxviii. the amino acid at position 313 is N or G;Ixxxix. the amino acid at position 316 is R, K or P; xc. the amino acid at position 321 is R or Q; xci. the amino acid at position 326 is K or H; xcii. the amino acid at position 327 is F or L; xciii. the amino acid at position 328 is F or L; xciv. the amino acid at position 330 is N or G; xcv. the amino acid at position 335 is S or N; xcvi. the amino acid at position 349 is A or G; xcvii. the amino acid at position 359 is I or V; xcviii. the amino acid at position 368 is A or V; xcix. the amino acid at position 370 is L or F; c. the amino acid at position 375 is L or E; ci. the amino acid at position 379 is D or A; cii. the amino acid at position 384 is L or E; ciii. the amino acid at position 385 is C, M or N; civ. the amino acid at position 390 is D, E or R; cv. the amino acid at position 394 is R or A; cvi. the amino acid at position 397 is G or A; and / or cvii. the amino acid at position 398 is G or deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO: 1 , SEQ ID NO:96 or SEQ ID NO: 125.

[0014] In one embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO:96, or an amino acid sequence that has at least 80% sequence identity thereto comprises an amino acid sequence wherein i. the amino acid at position 3 is Q or T; ii. the amino acid at position 5 is N or D; iii. the amino acid at position 10 is S or F; iv. the amino acid at position 17 is Q or E; v. the amino acid at position 25 is Q or T; vi. the amino acid at position 26 is A or P; vii. the amino acid at position 29 is N or S; viii. the amino acid at position 32 is A or S; ix. the amino acid at position 39 is I or L;x. the amino acid at position 40 is I or V; xi. the amino acid at position 44 is N or Q; xii. the amino acid at position 51 is A or P; xiii. the amino acid at position 52 is A or V; xiv. the amino acid at position 53 is S or W; xv. the amino acid at position 57 is A or R; xvi. the amino acid at position 58 is A or S; xvii. the amino acid at position 63 is D or G; xviii. the amino acid at position 64 is G, E or R; xix. the amino acid at position 67 is F or L xx. the amino acid at position 70 is P or deleted; xxi. the amino acid at position 73 is R or K; xxii. the amino acid at position 74 is S or E; xxiii. the amino acid at position 75 is T, M, G or W; xxiv. the amino acid at position 78 is A or G; xxv. the amino acid at position 82 is A or E; xxvi. the amino acid at position 84 is S or T; xxvii. the amino acid at position 88 is A or G; xxviii. the amino acid at position 92 is L or M; xxix. the amino acid at position 99 is A or S; xxx. the amino acid at position 101 is W or H; xxxi. the amino acid at position 110 is S or T; xxxii. the amino acid at position 114 is L or V; xxxiii. the amino acid at position 124 is I or T; xxxiv. the amino acid at position 131 is D or E; xxxv. the amino acid at position 143 is H, R or D; xxxvi. the amino acid at position 148 is A or V; xxxvii. the amino acid at position 149 is A or K; xxxviii. the amino acid at position 167 is S or G; xxxix. the amino acid at position 171 is E or D; xl. the amino acid at position 186 is R, E or Q; xli. the amino acid at position 188 is T or M; xlii. the amino acid at position 193 is A or P; xliii. the amino acid at position 194 is S or P;xliv. the amino acid at position 197 is A or E; xlv. the amino acid at position 207 is A or V; xlvi. the amino acid at position 210 is R or K; xlvii. the amino acid at position 211 is R, K or deleted; xlviii. the amino acid at position 212 is S, D or G; xlix. the amino acid at position 215 is P or E; 1. the amino acid at position 223 is A or V; li. the amino acid at position 234 is L or I; lii. the amino acid at position 244 is D or E; liii. the amino acid at position 249 is H or R; liv. the amino acid at position 252 is Q or R; Iv. the amino acid at position 258 is H or A; Ivi. the amino acid at position 276 is L or I; Ivii. the amino acid at position 280 is L or A; Iviii. the amino acid at position 288 is Q or E; lix. the amino acid at position 291 is L or R; lx. the amino acid at position 301 is I or V; Ixi. the amino acid at position 304 is E, A or G; Ixii. the amino acid at position 305 is A or W; Ixiii. the amino acid at position 311 is L or R; Ixiv. the amino acid at position 312 is Q or T; Ixv. the amino acid at position 313 is N or G; Ixvi. the amino acid at position 316 is R, K or P; Ixvii. the amino acid at position 326 is K or H; Ixviii. the amino acid at position 327 is F or L; Ixix. the amino acid at position 328 is F or L; Ixx. the amino acid at position 330 is N or G; Ixxi. the amino acid at position 349 is A or G; Ixxii. the amino acid at position 359 is I or V; Ixxiii. the amino acid at position 368 is A or V; Ixxiv. the amino acid at position 370 is L or F; Ixxv. the amino acid at position 375 is L or E; Ixxvi. the amino acid at position 385 is C, M or N; Ixxvii. the amino acid at position 390 is D, E or R;Ixxviii. the amino acid at position 397 is G or A; and / orIxxix. the amino acid at position 398 is G or deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96.

[0015] In one embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO:96; amino acid residues 2-398 of SEQ ID NO: 1; or an amino acid sequence that has at least 80% sequence identity thereto, wherein i. the amino acid at position 3 is Q or T; ii. the amino acid at position 5 is N or D; iii. the amino acid at position 25 is Q or T; iv. the amino acid at position 26 is A or P; v. the amino acid at position 32 is A or S; vi. the amino acid at position 40 is I or V; vii. the amino acid at position 51 is A or P; viii. the amino acid at position 52 is A or V; ix. the amino acid at position 53 is S or W; x. the amino acid at position 57 is A or R; xi. the amino acid at position 58 is A or S; xii. the amino acid at position 63 is D or G; xiii. the amino acid at position 64 is G, E or R; xiv. the amino acid at position 70 is P or deleted; xv. the amino acid at position 73 is R or K; xvi. the amino acid at position 74 is S or E; xvii. the amino acid at position 75 is T, M or W; xviii. the amino acid at position 78 is A or G; xix. the amino acid at position 82 is A or E; xx. the amino acid at position 84 is S or T; xxi. the amino acid at position 88 is A or G; xxii. the amino acid at position 92 is L or M; xxiii. the amino acid at position 99 is A or S; xxiv. the amino acid at position 101 is W or H; xxv. the amino acid at position 110 is S or T; xxvi. the amino acid at position 114 is L or V; xxvii. the amino acid at position 124 is I or T;xxviii. the amino acid at position 131 is D or E; xxix. the amino acid at position 143 is H, R or D; xxx. the amino acid at position 148 is A or V; xxxi. the amino acid at position 149 is A or K; xxxii. the amino acid at position 167 is S or G; xxxiii. the amino acid at position 171 is E or D; xxxiv. the amino acid at position 186 is R, E or Q; xxxv. the amino acid at position 193 is A or P; xxxvi. the amino acid at position 194 is S or P; xxxvii. the amino acid at position 197 is A or E; xxxviii. the amino acid at position 207 is A or V; xxxix. the amino acid at position 210 is R or K; xl. the amino acid at position 211 is R, K or deleted; xli. the amino acid at position 212 is S, D or G; xlii. the amino acid at position 215 is P or E; xliii. the amino acid at position 223 is A or V; xliv. the amino acid at position 234 is L or I; xlv. the amino acid at position 244 is D or E; xlvi. the amino acid at position 276 is L or I; xlvii. the amino acid at position 280 is L or A; xlviii. the amino acid at position 288 is Q or E; xlix. the amino acid at position 291 is L or R; 1. the amino acid at position 301 is I or V; li. the amino acid at position 304 is E, A or G; lii. the amino acid at position 305 is A or W; liii. the amino acid at position 311 is L or R; liv. the amino acid at position 312 is Q or T; Iv. the amino acid at position 313 is N or G; Ivi. the amino acid at position 316 is R, K or P; Ivii. the amino acid at position 328 is F or L; Iviii. the amino acid at position 330 is N or G; lix. the amino acid at position 349 is A or G; lx. the amino acid at position 368 is A or V; Ixi. the amino acid at position 370 is L or F;Ixii. the amino acid at position 375 is L or E;Ixiii. the amino acid at position 385 is C or N;Ixiv. the amino acid at position 390 is D, E or R; and / orIxv. the amino acid at position 397 is G or A, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96.

[0016] In one embodiment, the polypeptide comprises or consists of the amino acid sequence having amino acid residues 2-397 of SEQ ID NO:2; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 15 ; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO:5; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 10; the amino acid sequence having amino acid residues 2-397 of SEQ ID NO:8; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO:4; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 12; the amino acid sequence having amino acid residues 2-397 of SEQ ID NO:91; the amino acid sequence having amino acid residues 2-397 of SEQ ID NO:92; the amino acid sequence having amino acid residues 2-397 of SEQ ID NO:93; the amino acid sequence having amino acid residues 2-397 of SEQ ID NO:94; the amino acid sequence having amino acid residues 2-397 of SEQ ID NO:95; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 14; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO:89; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO:6; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO:90; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO:9; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO:7; the amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 11 or the amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 13.

[0017] In another embodiment; the polypeptide consists of the amino acid sequence of SEQ ID NO:2; SEQ ID NO:15; SEQ ID NO:5; SEQ ID NO:10; SEQ ID NO:8; SEQ ID NO:4; SEQ ID NO: 12; SEQ ID NO:91; SEQ ID NO:92; SEQ ID NO:93; SEQ ID NO:94; SEQ ID NO:95; SEQ ID NO: 14; SEQ ID NO:89; SEQ ID NO:6; SEQ ID NO:90; SEQ ID NO:9; SEQ ID NO:7; SEQ ID NO: 11; or SEQ ID NO: 13.

[0018] In one aspect disclosed herein, there is provided a polypeptide capable of hydrolysing an amide bond in a polyamide, wherein the polypeptide comprises or consists of an amino acid sequence of amino acid residues 2-394 of SEQ ID NO:88 or an amino acidsequence that has more than 75% sequence identity thereto. In one embodiment, the polypeptide comprises at least 80% sequence identity to SEQ ID NO:88. In one embodiment, the polypeptide comprises at least 85% sequence identity to SEQ ID NO:88. In one embodiment, the polypeptide comprises at least 90% sequence identity to SEQ ID NO:88. . In one embodiment, the polypeptide comprises at least 95% sequence identity to SEQ ID NO:88.

[0019] In one embodiment, the polypeptide comprises the amino acid sequence having amino acid residues 2-394 of SEQ ID NO: 123. In one embodiment, the polypeptide consists of the amino acid sequence of SEQ ID NO: 123.

[0020] In another embodiment, the polypeptide comprises or consists of the amino acid sequence having amino acid residues 2-392 of SEQ ID NO:3; the amino acid sequence having amino acid residues 2-394 of SEQ ID NO:63; or the amino acid sequence having amino acid residues 2-398 of SEQ ID NO:48. In one embodiment, the polypeptide comprises the amino acid sequence of amino acid residues 2-394 of the amino acid sequence of SEQ ID NO:63.

[0021] In one embodiment, the polypeptide consists of the amino acid sequence of SEQ ID NO:3, SEQ ID NO:63 or SEQ ID NO:48. In one embodiment, the polypeptide consists of the amino acid sequence of SEQ ID NO:63. In one embodiment, the polypeptide consists of the amino acid sequence of SEQ ID NO: 3.

[0022] In one aspect disclosed herein, there is provided a polypeptide capable of hydrolysing an amide bond in a polyamide, wherein the polypeptide comprises or consists of an amino acid sequence of amino acid residues 2-391 of SEQ ID NO:45 or an amino acid sequence that has at least 61% sequence identity thereto. In one embodiment, the polypeptide comprises at least 70% sequence identity to SEQ ID NO:45. In one embodiment, the polypeptide comprises at least 75% sequence identity to SEQ ID NO:45. In one embodiment, the polypeptide comprises at least 80% sequence identity to SEQ ID NO:45. In one embodiment, the polypeptide comprises at least 85% sequence identity to SEQ ID NO:45 In one embodiment, the polypeptide comprises at least 90% sequence identity to SEQ ID NO:45. In one embodiment, the polypeptide comprises at least 95% sequence identity to SEQ ID NO:45.

[0023] In one embodiment, the polypeptide comprises or consists of the amino acid sequence having amino acid residues 2-391 of SEQ ID NO:25 or the amino acid sequence having amino acid residues 2-391 of SEQ ID NO:22. In one embodiment, the polypeptide consists of the amino acid sequence of SEQ ID NO:25, SEQ ID NO:63 or SEQ ID NO:22.

[0024] In one embodiment, the polypeptides disclosed herein are capable of hydrolysing a nylon polymer. In another embodiment, the polypeptides disclosed herein has adipic acid mono- and di-N-alkyl amide hydrolase activity.

[0025] In another embodiment, the polypeptides disclosed herein are capable of hydrolysing a nylon polymer, wherein the nylon polymer is a nylon oligomer. In one embodiment, the nylon polymer or nylon oligomer is a nylon 6 polymer, a nylon 6 oligomer, a nylon 6,6 polymer or a nylon 6,6 oligomer. In another embodiment, the nylon polymer or the nylon oligomer is a nylon 6,6 polymer or a nylon 6,6 oligomer. In one embodiment, the nylon oligomer is selected from the group consisting of a dimer, a trimer, a tetramer, a pentamer and hexamer of nylon 6,6. In one embodiment, the nylon oligomer is a dimer. In one embodiment, the nylon oligomer is a trimer. In one embodiment, the nylon oligomer is a tetramer. In a further embodiment, the nylon 6,6 oligomer is an aqueous soluble nylon 6,6 oligomer.

[0026] In another embodiment, the polypeptides disclosed herein is capable of hydrolysing the nylon 6,6 oligomer to produce adipic acid and hexamethylenediamine.

[0027] In an embodiment, the polypeptides disclosed herein demonstrate increased recombinant expression in a host cell system; increased enzyme activity; and / or increased thermostability, when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86.

[0028] The present disclosure also extends to a composition comprising the polypeptide as described herein.

[0029] The present disclosure also extends to a nucleic acid sequence encoding the polypeptide described herein.

[0030] The present disclosure also extends to an expression vector comprising the nucleic acid sequence described herein.

[0031] The present disclosure also extends to a host cell comprising the nucleic acid sequence or the expression vector described herein.

[0032] In another aspect, the present disclosure provides a method of producing a polypeptide capable of hydrolysing an amide bond in a polyamide, the method comprising: i) providing the polynucleotide described herein; ii) expressing the polynucleotide in a host cell under conditions sufficient to allow the host cell to produce the polypeptide; and iii) collecting the polypeptide produced by the host cell in ii).

[0033] In another aspect, the present disclosure provides a method of hydrolysing nylon polymer or nylon oligomer, the method comprising exposing the nylon polymer or nylon oligomer to the polypeptide, the composition or the host cell disclosed herein under conditions sufficient to convert the nylon-6, 6 polymer to adipic acid and I or hexamethylenediamine .

[0034] In another aspect, the present disclosure provides a method of degrading a nylon- containing product, the method comprising exposing the nylon-containing product to the polypeptide, the composition or the host cell as disclosed herein.

[0035] In an embodiment, the methods disclosed herein comprises i) chemical processing of nylon polymer, nylon oligomer or nylon-containing product to generate nylon oligomers, ii) exposing the nylon oligomers generated in step (i) to the polypeptide, the composition or the host cell as disclosed herein, under conditions sufficient to produce adipic acid and I or hexamethylenediamine. In one embodiment, the nylon oligomers are nylon dimers, nylon trimers, nylon tetramers, nylon pentamers and nylon hexamers. In another embodiment, the nylon oligomers are aqueous soluble nylon oligomers.

[0036] In an embodiment, the nylon in the nylon polymer, nylon oligomer or the nylon- containing product is a nylon 6 or a nylon 6,6. In another embodiment, the nylon is a nylon 6,6.

[0037] In another aspect, the present disclosure provides a method of hydrolysing a nylon 6,6 polymer, a nylon 6,6 oligomer or a nylon 6,6-comprising product, the method comprising exposing the nylon 6,6 polymer, nylon 6,6 oligomer or a nylon 6,6 comprising product to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73,74 and 87, or an amino acid sequence that has at least 70% sequence identity to the amino acid sequenceof any one of SEQ ID NOs: 73,74 and 87. In an embodiment, the polypeptide comprises the amino acid sequence of any one of SEQ ID NOs: 73, 74 and 87.

[0038] In another embodiment, the methods described herein further comprise recovering the adipic acid and I or the hexamethylenediamine produced in step (ii).

[0039] In another aspect, the present disclosure provides a composition comprising the adipic acid and I or the hexamethylenediamine recovered by the methods disclosed herein.

[0040] In another aspect, the present disclosure provides a method of producing a nylon polymer using the composition of adipic acid and I or the hexamethylenediamine recovered by the methods disclosed herein.BRIEF DESCRIPTION OF THE FIGURES

[0041] Figure 1 shows the enzymatic hydrolysis of nylon 6,6 oligomers (A). The red dashed lines in (B) indicate the amide bonds that are hydrolysed by the polypeptides disclosed herein.

[0042] Figure 2 shows the amino acid sequences of nylonase NylB, P07061 and P07062.

[0043] Figure 3 shows whole-cell activity assays demonstrating the activity of variant polypeptides having amino acid sequence of SEQ ID NOs:2-15 and 88 in hydrolysing an amide bond in a polyamide, in comparison to a selection of extant I ancestral NylB polypeptides. A colorimetric assay is used to detect free amines in solution (absorbance at 335 nm). Increased concentration of free amine groups in the reaction solution corresponds to hydrolysis of Nylon-6, 6 trimer substrate to hexamethylenediamine and adipic acid.

[0044] Figure 4 shows the thermostability of variants A6 (SEQ ID NO:4) and C2 (SEQ ID NO:2) as determined by Circular Dichroism, in comparison to extant variants A10 (SEQ ID NO:75), All (SEQ ID NO:76), Bl l (SEQ ID NO:78), CIO ((SEQ ID NO:79), Cl l (SEQ ID NO:80), H10 (SEQ ID NO:86) as determined by Differential Scanning Fluorimetry.

[0045] Figure 5 shows an alignment of polypeptides of SEQ ID NOs:2, 4-15.

[0046] Figure 6 shows the activity of immobilised enzyme of SEQ ID NO:4 on the nylon 6,6 dimer, trimer and tetramer when incubated with the immobilised enzyme for 1 hour at40 °C. Conversion of the substrate was quantified by LC-MS and compared to a control (no enzyme at time 0 hour).

[0047] Figure 7 shows activity of variant polypeptides generated from further engineering (Phase II). High-throughput (crude / whole-cell activity) screening of candidate nylon 6,6 enzymes (compared to Genl Top (SEQ ID NO: 4) and no enzyme control) against nylon 6,6 oligomers. The candidates appearing to have the highest whole-cell activity is highlighted (A). Purified candidate enzymes were analysed for activity against nylon 6,6, dimers, trimers and tetramers using LC-MS (B). Y-axis values correspond to % normalised conversion of nylon 6,6 oligomers in comparison to no-enzyme control.

[0048] Figure 8 shows refined LC-MS assay comparing the whole-cell activity of top candidates from Phase 1 (SEQ ID NOs 4, 48, 88, 63 and 4) and Phase II (SEQ ID NOs: 89, 90, 91, 92, 93, 94) in comparison to extant sequences (SEQ ID NOs 75, 76, 79, 80, 81, 82, 83, 84, 86) on nylon 6,6, dimers (A) trimers (B) and tetramers (C). Y-axis values correspond to nylon 6,6 oligomer peak area as measured using LC-MS.

[0049] Figure 9 shows the kinetic stability of top candidates from Gen 2 (SEQ ID NOs: 89, 90, 91, 92, 93, 95) on nylon 6,6, trimer at 40-80°C. Y-axis values correspond to % normalised conversion of nylon 6,6 trimer in comparison to no-enzyme control.

[0050] Figure 10 shows an alignment of polypeptides of SEQ ID NOs:2, 4-15, 89, and 91- 95, with consensus II (SEQ ID NO:96).

[0051] Figure 11 shows the activity of SEQ ID NOs: 100-115 and 121-123 in comparison to extant sequence of SEQ ID NO: 75 on nylon 6,6, trimers. A no-enzyme control is included.

[0052] Figure 12 shows an alignment of SEQ ID NOs: 2, 4-15, 89, 91-95, and 97-122 in generating consensus sequence of consensus IV (SEQ ID NO: 125).DETAILED DESCRIPTION OF THE INVENTION

[0053] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, preferredmethods and materials are described. For the purposes of the present invention, the following terms are defined below.

[0054] The articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article, unless explicitly stated otherwise. By way of example, “an element” means one element or more than one element.

[0055] As used herein, the term "about" refers to a quantity, level, value, dimension, size, or amount that varies by as much as 10% (e.g, by 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% or 1%) to a reference quantity, level, value, dimension, size, or amount.

[0056] Throughout this specification, unless the context requires otherwise, the words "has", "have", "having" "comprise", "comprises" and "comprising" will be understood to imply the inclusion of a stated step or element or group of steps or elements but not the exclusion of any other step or element or group of steps or elements.

[0057] The present disclosure relates generally to the design and production of engineered polypeptides, including those with improved hydrolase activity, such as high activity, or broader activity on nylon polymers and nylon oligomers, particularly nylon dimers, trimers, tetramers, and pentamers of nylon. The present disclosure is predicated, at least in part, on the inventors' surprising finding that said engineered polypeptides, from the ancestral sequence reconstruction of extant and ancestral variants of the NylB family, have one or more increased or enhanced properties relative to one or more of the extant enzymes. For example, in certain embodiments, the engineered polypeptides disclosed herein have increased activity in hydrolysing an amide bond in a polyamide; as shown in an example provided herein, an improved ability to hydrolyse amide bonds in nylon polyamide. In certain embodiments, the engineered polypeptides disclosed herein have increased thermal stability. This is a highly surprising discovery, including because the temperature conditions to which a hypothetical ancestral enzyme may have been exposed would not be too dissimilar to the temperature or conditions to which one or more corresponding extant enzymes are exposed. In certain embodiments, the engineered polypeptides disclosed herein are associated with increased recombinant expression in a host cell system, where such host cells are modified by insertion of a polynucleotide sequence encoding said enzymes. This is particularly advantageous, as the presently known nylonase enzymes only demonstrate low levels of enzyme expression in common industrial host organisms.

[0058] In one aspect disclosed herein, there is provided a polypeptide capable of hydrolysing an amide bond in a polyamide, wherein the polypeptide comprises an amino acid sequence of amino acid residues 2-398 of SEQ ID NO: 2 or an amino acid sequence that has at least 70% sequence identity thereto.

[0059] In an embodiment, the polypeptide comprises at least one amino acid modification at a position selected from the group consisting of amino acid positions 3, 5, 7, 9, 10, 14, 16, 17, 25, 26, 29, 32, 39, 40, 44, 48, 51, 52, 53, 57, 58, 60, 61, 63, 64, 67, 70, 72, 73, 74, 75, 78, 82, 84, 88, 92, 99, 101, 110, 114, 123, 124, 131, 132, 138,143, 148, 149, 155, 167, 171,173, 178, 179, 180, 186, 188, 193, 194, 195, 197, 203, 206, 207, 210, 211, 212, 213, 215,223, 234, 244, 249, 252, 258, 262, 276, 278, 280, 288, 291, 301, 304, 305, 308, 311, 312,313, 316, 321, 326, 327, 328, 330, 335, 349, 359, 368, 370, 375, 379, 384, 385, 390, 394,397 and 398, with numbering relative to SEQ ID NO: 125.

[0060] In an embodiment, the polypeptide comprises at least one amino acid modification at a position selected from the group consisting of amino acid positions 3, 5, 10, 17, 25, 26, 29, 32, 39, 40, 44, 51, 52, 53, 57, 58, 63, 64, 67, 70, 73, 74, 75, 78, 82, 84, 88, 92, 99, 101, 110, 114, 124, 131, 143, 148, 149, 167, 171, 186, 188, 193, 194, 197, 207, 210, 211, 212, 215, 223, 234, 244, 249, 252, 258, 276, 280, 288, 291, 301, 304, 305, 311, 312, 313, 316, 326, 327, 328, 330, 349, 359, 368, 370, 375, 385, 390, 397 and 398, with numbering relative to SEQ ID NO: 96.

[0061] In another embodiment, the at least one amino acid modification is selected from the group consisting of Q3 or T3; N5 or D5; M7 or F7; G9 or T9; S10 or F10; P14 or A14; Q16 or G16; Q17 or E17; Q25 or T25; A26 or P26; N29 or S29; A32 or S32; 139 or L39; 140 or V40; N44 or Q44; G48 or A48; A51 or P51; A52 or V52; S53 or W53; A57 or R57; A58 or S58; R60 or A60; D61, A61 or G61; D63 or G63; G64, E64 or R64; F67 or L67; P70 or 70del; G72 or A72; R73 or K73; S74 or E74; T75, M75, G75 or W75; A78 or G78; A82 or E82; S84 of T84; A88 or G88; L92 or M92; A99 or S99; W101 or H101; SI 10 or T110; LI 14 or VI 14; G123 or A123; 1124 or T124; D131 of E 131 ; R132 or D132; D138 or A138; H143, D143 or R143; A148 or V148; A149 or K149; D155 or G155; S167 or G167; E171 or D171; D173 or A173; D178 or A178; G179 or S179; D180 or A180; R186, Q186 or E186; T188 or M188; A193 or P193; S194 or P194; D195 or P195; A197 or E197; R203 or Y203; L206 or 1206; A207 or V207; R210 or K210; R211, K211 or 21 Idel; S212, G212 or D212; D213 or A213; P215 or E215; A223 or V223; L234 or 1234; D244 or E244; H249 orR249; Q252 or R252; H258 or A258; 1262 or V262; 1276 or L276; A278 or C278; L280 or A280; E288 or Q288; R291 or L291; 1301 or V301; G304, E304 or A304; A305 or W305; D308 or A308; L311 or R311; Q312 or T312; N313 or G313; R316, P316 or K316; R321 or Q321; K326 or H326; F327 or L327; F328 or L328; N330 or G330; S335 or N335; A349 or G349; 1359 or V359; V368 or A368; L370 or F370; L375 or E375; D379 or A379; C385, L384 or E384; N385 or M385; D390, E390 or R390; R394 or A394; G397 or A397 and G398 or 398del, with numbering relative to SEQ ID NO: 96 or SEQ ID NO: 125.

[0062] In an embodiment, the polypeptide comprises at least one amino acid modification at a position selected from the group consisting of amino acid positions 3, 5, 25, 26, 32, 40, 51, 52, 53, 57, 58, 63, 64, 70, 73, 74, 75, 78, 82, 84, 88, 92, 99, 101, 110, 114, 124, 131, 143, 148, 149, 167, 171, 186, 193, 194, 197, 207, 210, 211, 212, 215, 223, 234, 244, 276, 280, 288, 291, 301, 304, 305, 311, 312, 313, 316, 328, 330, 349, 368, 370, 375, 385, 390 and 397, wherein the numbering is relative to the amino acid positions of SEQ ID NO: 1.

[0063] In another embodiment, the at least one amino acid modification is selected from the group consisting of Q3 or T3; N5 or D5; Q25 or T25; A26 or P26; A32 or S32; 140 or V40; A51 or P51; A52 or V52; S53 or W53; A57 or R57; A58 or S58; D63 or G63; G64, E64 or R64; P70 or 70del; R73 or K73; S74 or E74; T75, M75 or W75; A78 or G78; A82 or E82; S84 of T84; A88 or G88; L92 or M92; A99 or S99; W101 or H101; SI 10 or T110; LI 14 or V114; 1124 or T124; D131 of E131; H143, D143 or R143; A148 or V148; A149 or K149; S167 or G167; E171 or D171; R186, Q186 or E186; A193 or P193; S194 or P194; A197 or E197; A207 or V207; R210 or K210; R211, K211 or 21 Idel; S212, G212 or D212; P215 or E215; A223 or V223; L234 or 1234; D244 or E244; 1276 or L276; L280 or A280; E288 or Q288; R291 or L291; 1301 or V301; G304, E304 or A304; A305 or W305; L311 or R311; Q312 or T312; N313 or G313; R316, P316 or K316; F328 or L328; N330 or G330; A349 or G349; V368 or A368; L370 or F370; L375 or E375; C385 orN385; D390, E390 orR390 and G397 or A397, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1.

[0064] There is provided a polypeptide capable of hydrolysing an amide bond in a polyamide, having at least 80% sequence identity to the amino acid sequence of SEQ ID NO:2. There is also provided a polypeptide capable of hydrolysing an amide bond in a polyamide, having at least 90% sequence identity to the amino acid sequence of SEQ ID NO:2. There is also provided a polypeptide capable of hydrolysing an amide bond in apolyamide, having at least 95% sequence identity to the amino acid sequence of SEQ ID NO:2.

[0065] In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO:96 or an amino acid sequence that has at least 80% sequence identity thereto.

[0066] In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 125; wherein i. the amino acid at position 3 is Q or T; ii. the amino acid at position 5 is N or D; iii. the amino acid at position 7 is M or F; iv. the amino acid at position 9 is G or T; v. the amino acid at position 10 is S or F; vi. the amino acid at position 14 is P or A; vii. the amino acid at position 16 is Q or G viii. the amino acid at position 17 is Q or E; ix. the amino acid at position 25 is Q or T; x. the amino acid at position 26 is A or P; xi. the amino acid at position 29 is N or S; xii. the amino acid at position 32 is A or S; xiii. the amino acid at position 39 is I or L; xiv. the amino acid at position 40 is I or V; xv. the amino acid at position 44 is N or Q; xvi. the amino acid at position 48 is G or A xvii. the amino acid at position 51 is A or P; xviii. the amino acid at position 52 is A or V; xix. the amino acid at position 53 is S or W; xx. the amino acid at position 57 is A or R; xxi. the amino acid at position 58 is A or S; xxii. the amino acid at position 60 is R or A; xxiii. the amino acid at position 61 is D, A or G; xxiv. the amino acid at position 63 is D or G; xxv. the amino acid at position 64 is G, E or R;xxvi. the amino acid at position 67 is F or L xxvii. the amino acid at position 70 is P or deleted; xxviii. the amino acid at position 72 is G or A; xxix. the amino acid at position 73 is R or K; xxx. the amino acid at position 74 is S or E; xxxi. the amino acid at position 75 is T, M, G or W; xxxii. the amino acid at position 78 is A or G; xxxiii. the amino acid at position 82 is A or E; xxxiv. the amino acid at position 84 is S or T; xxxv. the amino acid at position 88 is A or G; xxxvi. the amino acid at position 92 is L or M; xxxvii. the amino acid at position 99 is A or S; xxxviii. the amino acid at position 101 is W or H; xxxix. the amino acid at position 110 is S or T; xl. the amino acid at position 114 is L or V; xli. the amino acid at position 123 is G or A; xlii. the amino acid at position 124 is I or T; xliii. the amino acid at position 131 is D or E; xliv. the amino acid at position 132 is R or D; xlv. the amino acid at position 138 is D or A; xlvi. the amino acid at position 143 is H, R or D; xlvii. the amino acid at position 148 is A or V; xlviii. the amino acid at position 149 is A or K; xlix. the amino acid at position 155 is D or G;1. the amino acid at position 167 is S or G; li. the amino acid at position 171 is E or D; lii. the amino acid at position 173 is D or A; liii. the amino acid at position 178 is D or A; liv. the amino acid at position 179 is G or S;Iv. the amino acid at position 180 is D or A;Ivi. the amino acid at position 186 is R, E or Q;Ivii. the amino acid at position 188 is T or M;Iviii. the amino acid at position 193 is A or P; lix. the amino acid at position 194 is S or P;lx. the amino acid at position 195 is D or P;Ixi. the amino acid at position 197 is A or E;Ixii. the amino acid at position 203 is R or Y;Ixiii. the amino acid at position 206 is L or I;Ixiv. the amino acid at position 207 is A or V;Ixv. the amino acid at position 210 is R or K;Ixvi. the amino acid at position 211 is R, K or deleted;Ixvii. the amino acid at position 212 is S, D or G;Ixviii. the amino acid at position 213 is D or A;Ixix. the amino acid at position 215 is P or E;Ixx. the amino acid at position 223 is A or V;Ixxi. the amino acid at position 234 is L or I;Ixxii. the amino acid at position 244 is D or E;Ixxiii. the amino acid at position 249 is H or R;Ixxiv. the amino acid at position 252 is Q or R;Ixxv. the amino acid at position 258 is H or A;Ixxvi. the amino acid at position 262 is I or V;Ixxvii. the amino acid at position 276 is L or I;Ixxviii. the amino acid at position 278 is A or C;Ixxix. the amino acid at position 280 is L, A or P;Ixxx. the amino acid at position 288 is Q or E;Ixxxi. the amino acid at position 291 is L or R;Ixxxii. the amino acid at position 301 is I or V;Ixxxiii. the amino acid at position 304 is E, A or G;Ixxxiv. the amino acid at position 305 is A or W;Ixxxv. the amino acid at position 308 is A or D;Ixxxvi. the amino acid at position 311 is L or R;Ixxxvii. the amino acid at position 312 is Q or T;Ixxxviii. the amino acid at position 313 is N or G;Ixxxix. the amino acid at position 316 is R, K or P; xc. the amino acid at position 321 is R or Q; xci. the amino acid at position 326 is K or H; xcii. the amino acid at position 327 is F or L; xciii. the amino acid at position 328 is F or L;xciv. the amino acid at position 330 is N or G; xcv. the amino acid at position 335 is S or N; xcvi. the amino acid at position 349 is A or G; xcvii. the amino acid at position 359 is I or V; xcviii. the amino acid at position 368 is A or V; xcix. the amino acid at position 370 is L or F; c. the amino acid at position 375 is L or E; ci. the amino acid at position 379 is D or A; cii. the amino acid at position 384 is L or E; ciii. the amino acid at position 385 is C, M or N; civ. the amino acid at position 390 is D, E or R; cv. the amino acid at position 394 is R or A; cvi. the amino acid at position 397 is G or A; and / or cvii. the amino acid at position 398 is G or deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1, SEQ ID NO:96 or SEQ ID NO: 125.

[0067] In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO:96, wherein i. the amino acid at position XI is Q or T; ii. the amino acid at position X2 is N or D; iii. the amino acid at position X3 is S or F; iv. the amino acid at position X4 is Q or E; v. the amino acid at position X5 is Q or T; vi. the amino acid at position X6 is A or P; vii. the amino acid at position X7 is N or S; viii. the amino acid at position X8 is A or S; ix. the amino acid at position X9 is I or L; x. the amino acid at position X10 is I or V; xi. the amino acid at position XI 1 is N or Q; xii. the amino acid at position X12 is A or P; xiii. the amino acid at position X 13 is A or V; xiv. the amino acid at position X14 is S or W; xv. the amino acid at position XI 5 is A or R;xvi. the amino acid at position X 16 is A or S; xvii. the amino acid at position X17 is D or G; xviii. the amino acid at position X18 is G, E or R; xix. the amino acid at position X19 is F or L xx. the amino acid at position X20 is P or deleted; xxi. the amino acid at position X21 is R or K; xxii. the amino acid at position X22 is S or E; xxiii. the amino acid at position X23 is T, M, G or W; xxiv. the amino acid at position X24 is A or G; xxv. the amino acid at position X25 is A or E; xxvi. the amino acid at position X26 is S or T; xxvii. the amino acid at position X27 is A or G; xxviii. the amino acid at position X28 is L or M; xxix. the amino acid at position X29 is A or S; xxx. the amino acid at position X30 is W or H; xxxi. the amino acid at position X31 is S or T; xxxii. the amino acid at position X32 is L or V; xxxiii. the amino acid at position X33 is I or T; xxxiv. the amino acid at position X34 is D or E; xxxv. the amino acid at position X35 is H, R or D; xxxvi. the amino acid at position X36 is A or V; xxxvii. the amino acid at position X37 is A or K; xxxviii. the amino acid at position X38 is S or G; xxxix. the amino acid at position X39 is E or D; xl. the amino acid at position X40 is R, E or Q; xli. the amino acid at position X41 is T or M; xlii. the amino acid at position X42 is A or P; xliii. the amino acid at position X43 is S or P; xliv. the amino acid at position X44 is A or E; xlv. the amino acid at position X45 is A or V; xlvi. the amino acid at position X46 is R or K; xlvii. the amino acid at position X47 is R, K or deleted; xlviii. the amino acid at position X48 is S, D or G; xlix. the amino acid at position X49 is P or E;1. the amino acid at position X50 is A or V; li. the amino acid at position X51 is L or I; lii. the amino acid at position X52 is D or E; liii. the amino acid at position X53 is H or R; liv. the amino acid at position X54 is Q or R;Iv. the amino acid at position X55 is H or A;Ivi. the amino acid at position X56 is L or I;Ivii. the amino acid at position X57 is L or A;Iviii. the amino acid at position X58 is Q or E; lix. the amino acid at position X59 is L or R; lx. the amino acid at position X60 is I or V;Ixi. the amino acid at position X61 is E, A or G;Ixii. the amino acid at position X62 is A or W;Ixiii. the amino acid at position X63 is L or R;Ixiv. the amino acid at position X64 is Q or T;Ixv. the amino acid at position X65 is N or G;Ixvi. the amino acid at position X66 is R, K or P;Ixvii. the amino acid at position X67 is K or H;Ixviii. the amino acid at position X68 is F or L;Ixix. the amino acid at position X69 is F or L;Ixx. the amino acid at position X70 is N or G;Ixxi. the amino acid at position X71 is A or G;Ixxii. the amino acid at position X72 is I or V;Ixxiii. the amino acid at position X73 is A or V;Ixxiv. the amino acid at position X74 is L or F;Ixxv. the amino acid at position X75 is L or E;Ixxvi. the amino acid at position X76 is C, M or N;Ixxvii. the amino acid at position X77 is D, E or R;Ixxviii. the amino acid at position X78 is G or A; and / orIxxix. the amino acid at position X79 is G or deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96.

[0068] In an embodiment, the polypeptide comprises an amino acid sequence whereini. the amino acid at position 3 is Q or T; ii. the amino acid at position 5 is N or D; iii. the amino acid at position 10 is S or F; iv. the amino acid at position 17 is Q or E; v. the amino acid at position 25 is Q or T; vi. the amino acid at position 26 is A or P; vii. the amino acid at position 29 is N or S; viii. the amino acid at position 32 is A or S; ix. the amino acid at position 39 is I or L; x. the amino acid at position 40 is I or V; xi. the amino acid at position 44 is N or Q; xii. the amino acid at position 51 is A or P; xiii. the amino acid at position 52 is A or V; xiv. the amino acid at position 53 is S or W; xv. the amino acid at position 57 is A or R; xvi. the amino acid at position 58 is A or S; xvii. the amino acid at position 63 is D or G; xviii. the amino acid at position 64 is G, E or R; xix. the amino acid at position 67 is F or L xx. the amino acid at position 70 is P or deleted; xxi. the amino acid at position 73 is R or K; xxii. the amino acid at position 74 is S or E; xxiii. the amino acid at position 75 is T, M, G or W; xxiv. the amino acid at position 78 is A or G; xxv. the amino acid at position 82 is A or E; xxvi. the amino acid at position 84 is S or T; xxvii. the amino acid at position 88 is A or G; xxviii. the amino acid at position 92 is L or M; xxix. the amino acid at position 99 is A or S; xxx. the amino acid at position 101 is W or H; xxxi. the amino acid at position 110 is S or T; xxxii. the amino acid at position 114 is L or V; xxxiii. the amino acid at position 124 is I or T; xxxiv. the amino acid at position 131 is D or E;xxxv. the amino acid at position 143 is H, R or D; xxxvi. the amino acid at position 148 is A or V; xxxvii. the amino acid at position 149 is A or K; xxxviii. the amino acid at position 167 is S or G; xxxix. the amino acid at position 171 is E or D; xl. the amino acid at position 186 is R, E or Q; xli. the amino acid at position 188 is T or M; xlii. the amino acid at position 193 is A or P; xliii. the amino acid at position 194 is S or P; xliv. the amino acid at position 197 is A or E; xlv. the amino acid at position 207 is A or V; xlvi. the amino acid at position 210 is R or K; xlvii. the amino acid at position 211 is R, K or deleted; xlviii. the amino acid at position 212 is S, D or G; xlix. the amino acid at position 215 is P or E; 1. the amino acid at position 223 is A or V; li. the amino acid at position 234 is L or I; lii. the amino acid at position 244 is D or E; liii. the amino acid at position 249 is H or R; liv. the amino acid at position 252 is Q or R; Iv. the amino acid at position 258 is H or A; Ivi. the amino acid at position 276 is L or I; Ivii. the amino acid at position 280 is L or A; Iviii. the amino acid at position 288 is Q or E; lix. the amino acid at position 291 is L or R; lx. the amino acid at position 301 is I or V; Ixi. the amino acid at position 304 is E, A or G; Ixii. the amino acid at position 305 is A or W; Ixiii. the amino acid at position 311 is L or R; Ixiv. the amino acid at position 312 is Q or T; Ixv. the amino acid at position 313 is N or G; Ixvi. the amino acid at position 316 is R, K or P; Ixvii. the amino acid at position 326 is K or H; Ixviii. the amino acid at position 327 is F or L;Ixix. the amino acid at position 328 is F or L;Ixx. the amino acid at position 330 is N or G;Ixxi. the amino acid at position 349 is A or G;Ixxii. the amino acid at position 359 is I or V;Ixxiii. the amino acid at position 368 is A or V;Ixxiv. the amino acid at position 370 is L or F;Ixxv. the amino acid at position 375 is L or E;Ixxvi. the amino acid at position 385 is C, M or N;Ixxvii. the amino acid at position 390 is D, E or R;Ixxviii. the amino acid at position 397 is G or A; and / orIxxix. the amino acid at position 398 is G or deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96.

[0069] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N; iii. the amino acid at position 10 is F; iv. the amino acid at position 17 is Q; v. the amino acid at position 25 is Q; vi. the amino acid at position 26 is A; vii. the amino acid at position 29 is N; viii. the amino acid at position 32 is S; ix. the amino acid at position 39 is I; x. the amino acid at position 40 is V; xi. the amino acid at position 44 is N; xii. the amino acid at position 51 is P; xiii. the amino acid at position 52 is V; xiv. the amino acid at position 53 is S; xv. the amino acid at position 57 is R; xvi. the amino acid at position 58 is A; xvii. the amino acid at position 63 is G; xviii. the amino acid at position 64 is E; xix. the amino acid at position 67 is Fxx. the amino acid at position 70 is P xxi. the amino acid at position 73 is K; xxii. the amino acid at position 74 is E; xxiii. the amino acid at position 75 is T; xxiv. the amino acid at position 78 is G; xxv. the amino acid at position 82 is E; xxvi. the amino acid at position 84 is T; xxvii. the amino acid at position 88 is G; xxviii. the amino acid at position 92 is L; xxix. the amino acid at position 99 is A; xxx. the amino acid at position 101 is H; xxxi. the amino acid at position 110 is S; xxxii. the amino acid at position 114 is V; xxxiii. the amino acid at position 124 is T; xxxiv. the amino acid at position 131 is E; xxxv. the amino acid at position 143 is H; xxxvi. the amino acid at position 148 is A; xxxvii. the amino acid at position 149 is A; xxxviii. the amino acid at position 167 is S; xxxix. the amino acid at position 171 is E; xl. the amino acid at position 186 is R; xli. the amino acid at position 188 is T; xlii. the amino acid at position 193 is P; xliii. the amino acid at position 194 is S; xliv. the amino acid at position 197 is A; xlv. the amino acid at position 207 is A; xlvi. the amino acid at position 210 is K; xlvii. the amino acid at position 211 is K; xlviii. the amino acid at position 212 is G; xlix. the amino acid at position 215 is P;1. the amino acid at position 223 is V; li. the amino acid at position 234 is L; lii. the amino acid at position 244 is D; liii. the amino acid at position 249 is H;liv. the amino acid at position 252 is Q;Iv. the amino acid at position 258 is H;Ivi. the amino acid at position 276 is I;Ivii. the amino acid at position 280 is L;Iviii. the amino acid at position 288 is E; lix. the amino acid at position 291 is R; lx. the amino acid at position 301 is I;Ixi. the amino acid at position 304 is E;Ixii. the amino acid at position 305 is A;Ixiii. the amino acid at position 311 is L;Ixiv. the amino acid at position 312 is T;Ixv. the amino acid at position 313 is N;Ixvi. the amino acid at position 316 is K;Ixvii. the amino acid at position 326 is K;Ixviii. the amino acid at position 327 is F;Ixix. the amino acid at position 328 is L;Ixx. the amino acid at position 330 is N;Ixxi. the amino acid at position 349 is A;Ixxii. the amino acid at position 359 is I;Ixxiii. the amino acid at position 368 is V;Ixxiv. the amino acid at position 370 is L;Ixxv. the amino acid at position 375 is L;Ixxvi. the amino acid at position 385 is C;Ixxvii. the amino acid at position 390 is R;Ixxviii. the amino acid at position 397 is G; andIxxix. the amino acid at position 398 is G, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 89. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 89.

[0070] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is N;iii. the amino acid at position 10 is F; iv. the amino acid at position 17 is Q; v. the amino acid at position 25 is Q; vi. the amino acid at position 26 is P; vii. the amino acid at position 29 is N; viii. the amino acid at position 32 is A; ix. the amino acid at position 39 is I; x. the amino acid at position 40 is I; xi. the amino acid at position 44 is N; xii. the amino acid at position 51 is P; xiii. the amino acid at position 52 is V; xiv. the amino acid at position 53 is S; xv. the amino acid at position 57 is R; xvi. the amino acid at position 58 is S; xvii. the amino acid at position 63 is G; xviii. the amino acid at position 64 is E; xix. the amino acid at position 67 is F; xx. the amino acid at position 70 is P; xxi. the amino acid at position 73 is K; xxii. the amino acid at position 74 is E; xxiii. the amino acid at position 75 is W; xxiv. the amino acid at position 78 is G; xxv. the amino acid at position 82 is A; xxvi. the amino acid at position 84 is S; xxvii. the amino acid at position 88 is A; xxviii. the amino acid at position 92 is M; xxix. the amino acid at position 99 is A; xxx. the amino acid at position 101 is W; xxxi. the amino acid at position 110 is S; xxxii. the amino acid at position 114 is V; xxxiii. the amino acid at position 124 is I; xxxiv. the amino acid at position 131 is E; xxxv. the amino acid at position 143 is R; xxxvi. the amino acid at position 148 is A;xxxvii. the amino acid at position 149 is A; xxxviii. the amino acid at position 167 is S; xxxix. the amino acid at position 171 is E; xl. the amino acid at position 186 is R; xli. the amino acid at position 188 is T; xlii. the amino acid at position 193 is P; xliii. the amino acid at position 194 is P; xliv. the amino acid at position 197 is A; xlv. the amino acid at position 207 is V; xlvi. the amino acid at position 210 is K; xlvii. the amino acid at position 211 is K; xlviii. the amino acid at position 212 is G; xlix. the amino acid at position 215 is P;1. the amino acid at position 223 is A; li. the amino acid at position 234 is L; lii. the amino acid at position 244 is D; liii. the amino acid at position 249 is H; liv. the amino acid at position 252 is Q;Iv. the amino acid at position 258 is H;Ivi. the amino acid at position 276 is I;Ivii. the amino acid at position 280 is A;Iviii. the amino acid at position 288 is E; lix. the amino acid at position 291 is R; lx. the amino acid at position 301 is I;Ixi. the amino acid at position 304 is E;Ixii. the amino acid at position 305 is W;Ixiii. the amino acid at position 311 is L;Ixiv. the amino acid at position 312 is Q;Ixv. the amino acid at position 313 is G;Ixvi. the amino acid at position 316 is R;Ixvii. the amino acid at position 326 is K;Ixviii. the amino acid at position 327 is F;Ixix. the amino acid at position 328 is F;Ixx. the amino acid at position 330 is N;Ixxi. the amino acid at position 349 is A;Ixxii. the amino acid at position 359 is I;Ixxiii. the amino acid at position 368 is V;Ixxiv. the amino acid at position 370 is L;Ixxv. the amino acid at position 375 is E;Ixxvi. the amino acid at position 385 is N;Ixxvii. the amino acid at position 390 is E;Ixxviii. the amino acid at position 397 is A; andIxxix. the amino acid at position 398 is G, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 90. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 90.

[0071] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N; iii. the amino acid at position 10 is S; iv. the amino acid at position 17 is E; v. the amino acid at position 25 is Q; vi. the amino acid at position 26 is A; vii. the amino acid at position 29 is S; viii. the amino acid at position 32 is S; ix. the amino acid at position 39 is L; x. the amino acid at position 40 is V; xi. the amino acid at position 44 is Q; xii. the amino acid at position 51 is A; xiii. the amino acid at position 52 is A; xiv. the amino acid at position 53 is S; xv. the amino acid at position 57 is A; xvi. the amino acid at position 58 is A; xvii. the amino acid at position 63 is G; xviii. the amino acid at position 64 is E; xix. the amino acid at position 67 is L;xx. the amino acid at position 70 is P; xxi. the amino acid at position 73 is K; xxii. the amino acid at position 74 is E; xxiii. the amino acid at position 75 is G; xxiv. the amino acid at position 78 is A; xxv. the amino acid at position 82 is E; xxvi. the amino acid at position 84 is S; xxvii. the amino acid at position 88 is G; xxviii. the amino acid at position 92 is L; xxix. the amino acid at position 99 is A; xxx. the amino acid at position 101 is H; xxxi. the amino acid at position 110 is S; xxxii. the amino acid at position 114 is V; xxxiii. the amino acid at position 124 is T; xxxiv. the amino acid at position 131 is E; xxxv. the amino acid at position 143 is D; xxxvi. the amino acid at position 148 is V; xxxvii. the amino acid at position 149 is A; xxxviii. the amino acid at position 167 is S; xxxix. the amino acid at position 171 is E; xl. the amino acid at position 186 is R; xli. the amino acid at position 188 is M; xlii. the amino acid at position 193 is P; xliii. the amino acid at position 194 is S; xliv. the amino acid at position 197 is E; xlv. the amino acid at position 207 is A; xlvi. the amino acid at position 210 is K; xlvii. the amino acid at position 211 is K; xlviii. the amino acid at position 212 is G; xlix. the amino acid at position 215 is P;1. the amino acid at position 223 is V; li. the amino acid at position 234 is L; lii. the amino acid at position 244 is D; liii. the amino acid at position 249 is R;liv. the amino acid at position 252 is R;Iv. the amino acid at position 258 is H;Ivi. the amino acid at position 276 is I;Ivii. the amino acid at position 280 is L;Iviii. the amino acid at position 288 is E; lix. the amino acid at position 291 is R; lx. the amino acid at position 301 is I;Ixi. the amino acid at position 304 is E;Ixii. the amino acid at position 305 is A;Ixiii. the amino acid at position 311 is L;Ixiv. the amino acid at position 312 is T;Ixv. the amino acid at position 313 is N;Ixvi. the amino acid at position 316 is R;Ixvii. the amino acid at position 326 is H;Ixviii. the amino acid at position 327 is L;Ixix. the amino acid at position 328 is L;Ixx. the amino acid at position 330 is N;Ixxi. the amino acid at position 349 is A;Ixxii. the amino acid at position 359 is V;Ixxiii. the amino acid at position 368 is V;Ixxiv. the amino acid at position 370 is L;Ixxv. the amino acid at position 375 is L;Ixxvi. the amino acid at position 385 is C;Ixxvii. the amino acid at position 390 is R;Ixxviii. the amino acid at position 397 is G; andIxxix. the amino acid at position 398 is deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 91. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 91.

[0072] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N;iii. the amino acid at position 10 is S; iv. the amino acid at position 17 is E; v. the amino acid at position 25 is Q; vi. the amino acid at position 26 is A; vii. the amino acid at position 29 is S; viii. the amino acid at position 32 is S; ix. the amino acid at position 39 is L; x. the amino acid at position 40 is V; xi. the amino acid at position 44 is Q; xii. the amino acid at position 51 is P; xiii. the amino acid at position 52 is A; xiv. the amino acid at position 53 is S; xv. the amino acid at position 57 is A; xvi. the amino acid at position 58 is A; xvii. the amino acid at position 63 is G; xviii. the amino acid at position 64 is E; xix. the amino acid at position 67 is L; xx. the amino acid at position 70 is P; xxi. the amino acid at position 73 is K; xxii. the amino acid at position 74 is E; xxiii. the amino acid at position 75 is G; xxiv. the amino acid at position 78 is A; xxv. the amino acid at position 82 is E; xxvi. the amino acid at position 84 is S; xxvii. the amino acid at position 88 is G; xxviii. the amino acid at position 92 is L; xxix. the amino acid at position 99 is A; xxx. the amino acid at position 101 is H; xxxi. the amino acid at position 110 is S; xxxii. the amino acid at position 114 is V; xxxiii. the amino acid at position 124 is T; xxxiv. the amino acid at position 131 is E; xxxv. the amino acid at position 143 is D; xxxvi. the amino acid at position 148 is V;xxxvii. the amino acid at position 149 is A; xxxviii. the amino acid at position 167 is S; xxxix. the amino acid at position 171 is E; xl. the amino acid at position 186 is R; xli. the amino acid at position 188 is M; xlii. the amino acid at position 193 is P; xliii. the amino acid at position 194 is S; xliv. the amino acid at position 197 is E; xlv. the amino acid at position 207 is A; xlvi. the amino acid at position 210 is K; xlvii. the amino acid at position 211 is K; xlviii. the amino acid at position 212 is G; xlix. the amino acid at position 215 is P;1. the amino acid at position 223 is V; li. the amino acid at position 234 is L; lii. the amino acid at position 244 is D; liii. the amino acid at position 249 is R; liv. the amino acid at position 252 is R;Iv. the amino acid at position 258 is A;Ivi. the amino acid at position 276 is I;Ivii. the amino acid at position 280 is L;Iviii. the amino acid at position 288 is E; lix. the amino acid at position 291 is R; lx. the amino acid at position 301 is I;Ixi. the amino acid at position 304 is E;Ixii. the amino acid at position 305 is A;Ixiii. the amino acid at position 311 is L;Ixiv. the amino acid at position 312 is T;Ixv. the amino acid at position 313 is N;Ixvi. the amino acid at position 316 is R;Ixvii. the amino acid at position 326 is H;Ixviii. the amino acid at position 327 is L;Ixix. the amino acid at position 328 is F;Ixx. the amino acid at position 330 is N;Ixxi. the amino acid at position 349 is A;Ixxii. the amino acid at position 359 is V;Ixxiii. the amino acid at position 368 is V;Ixxiv. the amino acid at position 370 is L;Ixxv. the amino acid at position 375 is L;Ixxvi. the amino acid at position 385 is M;Ixxvii. the amino acid at position 390 is R;Ixxviii. the amino acid at position 397 is G; andIxxix. the amino acid at position 398 is deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 92. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 92.

[0073] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N; iii. the amino acid at position 10 is S; iv. the amino acid at position 17 is E; v. the amino acid at position 25 is Q; vi. the amino acid at position 26 is A; vii. the amino acid at position 29 is S; viii. the amino acid at position 32 is S; ix. the amino acid at position 39 is L; x. the amino acid at position 40 is V; xi. the amino acid at position 44 is Q; xii. the amino acid at position 51 is P; xiii. the amino acid at position 52 is A; xiv. the amino acid at position 53 is S; xv. the amino acid at position 57 is A; xvi. the amino acid at position 58 is A; xvii. the amino acid at position 63 is G; xviii. the amino acid at position 64 is E; xix. the amino acid at position 67 is L;xx. the amino acid at position 70 is P; xxi. the amino acid at position 73 is K; xxii. the amino acid at position 74 is E; xxiii. the amino acid at position 75 is G; xxiv. the amino acid at position 78 is A; xxv. the amino acid at position 82 is E; xxvi. the amino acid at position 84 is S; xxvii. the amino acid at position 88 is G; xxviii. the amino acid at position 92 is L; xxix. the amino acid at position 99 is A; xxx. the amino acid at position 101 is H; xxxi. the amino acid at position 110 is S; xxxii. the amino acid at position 114 is V; xxxiii. the amino acid at position 124 is T; xxxiv. the amino acid at position 131 is E; xxxv. the amino acid at position 143 is D; xxxvi. the amino acid at position 148 is V; xxxvii. the amino acid at position 149 is A; xxxviii. the amino acid at position 167 is S; xxxix. the amino acid at position 171 is E; xl. the amino acid at position 186 is R; xli. the amino acid at position 188 is M; xlii. the amino acid at position 193 is P; xliii. the amino acid at position 194 is S; xliv. the amino acid at position 197 is E; xlv. the amino acid at position 207 is A; xlvi. the amino acid at position 210 is K; xlvii. the amino acid at position 211 is K; xlviii. the amino acid at position 212 is G; xlix. the amino acid at position 215 is P;1. the amino acid at position 223 is V; li. the amino acid at position 234 is L; lii. the amino acid at position 244 is D; liii. the amino acid at position 249 is R;liv. the amino acid at position 252 is R;Iv. the amino acid at position 258 is A;Ivi. the amino acid at position 276 is I;Ivii. the amino acid at position 280 is L;Iviii. the amino acid at position 288 is E; lix. the amino acid at position 291 is R; lx. the amino acid at position 301 is I;Ixi. the amino acid at position 304 is E;Ixii. the amino acid at position 305 is A;Ixiii. the amino acid at position 311 is L;Ixiv. the amino acid at position 312 is T;Ixv. the amino acid at position 313 is N;Ixvi. the amino acid at position 316 is R;Ixvii. the amino acid at position 326 is H;Ixviii. the amino acid at position 327 is L;Ixix. the amino acid at position 328 is L;Ixx. the amino acid at position 330 is N;Ixxi. the amino acid at position 349 is A;Ixxii. the amino acid at position 359 is V;Ixxiii. the amino acid at position 368 is V;Ixxiv. the amino acid at position 370 is L;Ixxv. the amino acid at position 375 is L;Ixxvi. the amino acid at position 385 is M;Ixxvii. the amino acid at position 390 is R;Ixxviii. the amino acid at position 397 is G; andIxxix. the amino acid at position 398 is deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 93. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 93.

[0074] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N;iii. the amino acid at position 10 is S; iv. the amino acid at position 17 is E; v. the amino acid at position 25 is Q; vi. the amino acid at position 26 is A; vii. the amino acid at position 29 is S; viii. the amino acid at position 32 is S; ix. the amino acid at position 39 is L; x. the amino acid at position 40 is V; xi. the amino acid at position 44 is Q; xii. the amino acid at position 51 is A; xiii. the amino acid at position 52 is A; xiv. the amino acid at position 53 is S; xv. the amino acid at position 57 is A; xvi. the amino acid at position 58 is A; xvii. the amino acid at position 63 is G; xviii. the amino acid at position 64 is E; xix. the amino acid at position 67 is L; xx. the amino acid at position 70 is P; xxi. the amino acid at position 73 is K; xxii. the amino acid at position 74 is E; xxiii. the amino acid at position 75 is G; xxiv. the amino acid at position 78 is A; xxv. the amino acid at position 82 is E; xxvi. the amino acid at position 84 is S; xxvii. the amino acid at position 88 is G; xxviii. the amino acid at position 92 is L; xxix. the amino acid at position 99 is A; xxx. the amino acid at position 101 is H; xxxi. the amino acid at position 110 is S; xxxii. the amino acid at position 114 is V; xxxiii. the amino acid at position 124 is T; xxxiv. the amino acid at position 131 is E; xxxv. the amino acid at position 143 is D; xxxvi. the amino acid at position 148 is V;xxxvii. the amino acid at position 149 is A; xxxviii. the amino acid at position 167 is S; xxxix. the amino acid at position 171 is E; xl. the amino acid at position 186 is R; xli. the amino acid at position 188 is M; xlii. the amino acid at position 193 is P; xliii. the amino acid at position 194 is S; xliv. the amino acid at position 197 is E; xlv. the amino acid at position 207 is A; xlvi. the amino acid at position 210 is K; xlvii. the amino acid at position 211 is K; xlviii. the amino acid at position 212 is G; xlix. the amino acid at position 215 is P;1. the amino acid at position 223 is V; li. the amino acid at position 234 is L; lii. the amino acid at position 244 is D; liii. the amino acid at position 249 is R; liv. the amino acid at position 252 is R;Iv. the amino acid at position 258 is A;Ivi. the amino acid at position 276 is I;Ivii. the amino acid at position 280 is L;Iviii. the amino acid at position 288 is E; lix. the amino acid at position 291 is R; lx. the amino acid at position 301 is I;Ixi. the amino acid at position 304 is E;Ixii. the amino acid at position 305 is A;Ixiii. the amino acid at position 311 is L;Ixiv. the amino acid at position 312 is T;Ixv. the amino acid at position 313 is N;Ixvi. the amino acid at position 316 is R;Ixvii. the amino acid at position 326 is H;Ixviii. the amino acid at position 327 is L;Ixix. the amino acid at position 328 is L;Ixx. the amino acid at position 330 is N;Ixxi. the amino acid at position 349 is A;Ixxii. the amino acid at position 359 is V;Ixxiii. the amino acid at position 368 is V;Ixxiv. the amino acid at position 370 is L;Ixxv. the amino acid at position 375 is L;Ixxvi. the amino acid at position 385 is M;Ixxvii. the amino acid at position 390 is R;Ixxviii. the amino acid at position 397 is G; andIxxix. the amino acid at position 398 is deleted wherein the numbering is relative to the amino acid positions of SEQ ID NO: 1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 94. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 94.

[0075] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N; iii. the amino acid at position 10 is S; iv. the amino acid at position 17 is E; v. the amino acid at position 25 is Q; vi. the amino acid at position 26 is A; vii. the amino acid at position 29 is S; viii. the amino acid at position 32 is S; ix. the amino acid at position 39 is L; x. the amino acid at position 40 is V; xi. the amino acid at position 44 is Q; xii. the amino acid at position 51 is P; xiii. the amino acid at position 52 is A; xiv. the amino acid at position 53 is S; xv. the amino acid at position 57 is A; xvi. the amino acid at position 58 is A; xvii. the amino acid at position 63 is G; xviii. the amino acid at position 64 is R; xix. the amino acid at position 67 is Lxx. the amino acid at position 70 is P; xxi. the amino acid at position 73 is K; xxii. the amino acid at position 74 is E; xxiii. the amino acid at position 75 is G; xxiv. the amino acid at position 78 is A; xxv. the amino acid at position 82 is A; xxvi. the amino acid at position 84 is S; xxvii. the amino acid at position 88 is G; xxviii. the amino acid at position 92 is L; xxix. the amino acid at position 99 is A; xxx. the amino acid at position 101 is H; xxxi. the amino acid at position 110 is S; xxxii. the amino acid at position 114 is V; xxxiii. the amino acid at position 124 is T; xxxiv. the amino acid at position 131 is E; xxxv. the amino acid at position 143 is D; xxxvi. the amino acid at position 148 is V; xxxvii. the amino acid at position 149 is A; xxxviii. the amino acid at position 167 is S; xxxix. the amino acid at position 171 is E; xl. the amino acid at position 186 is R; xli. the amino acid at position 188 is M; xlii. the amino acid at position 193 is P; xliii. the amino acid at position 194 is S; xliv. the amino acid at position 197 is E; xlv. the amino acid at position 207 is A; xlvi. the amino acid at position 210 is K; xlvii. the amino acid at position 211 is K; xlviii. the amino acid at position 212 is G; xlix. the amino acid at position 215 is P;1. the amino acid at position 223 is V; li. the amino acid at position 234 is L; lii. the amino acid at position 244 is D; liii. the amino acid at position 249 is R;liv. the amino acid at position 252 is R;Iv. the amino acid at position 258 is H;Ivi. the amino acid at position 276 is I;Ivii. the amino acid at position 280 is L;Iviii. the amino acid at position 288 is E; lix. the amino acid at position 291 is R; lx. the amino acid at position 301 is I;Ixi. the amino acid at position 304 is E;Ixii. the amino acid at position 305 is A;Ixiii. the amino acid at position 311 is L;Ixiv. the amino acid at position 312 is T;Ixv. the amino acid at position 313 is N;Ixvi. the amino acid at position 316 is R;Ixvii. the amino acid at position 326 is H;Ixviii. the amino acid at position 327 is L;Ixix. the amino acid at position 328 is L;Ixx. the amino acid at position 330 is N;Ixxi. the amino acid at position 349 is A;Ixxii. the amino acid at position 359 is V;Ixxiii. the amino acid at position 368 is V;Ixxiv. the amino acid at position 370 is L;Ixxv. the amino acid at position 375 is L;Ixxvi. the amino acid at position 385 is C;Ixxvii. the amino acid at position 390 is R;Ixxviii. the amino acid at position 397 is G; andIxxix. the amino acid at position 398 is deleted.

[0076] wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 95. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 95.1n an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 1 or an amino acid sequence that has at least 80% sequence identity thereto.

[0077] In another embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO:1, wherein i. the amino acid at position XI is Q or T; ii. the amino acid at position X2 is N or D; iii. the amino acid at position X3 is Q or T; iv. the amino acid at position X4 is A or P; v. the amino acid at position X5 is A or S; vi. the amino acid at position X6 is I or V; vii. the amino acid at position X7 is A or P; viii. the amino acid at position X8 is A or V; ix. the amino acid at position X9 is S or W; x. the amino acid at position X 10 is A or R; xi. the amino acid at position XI 1 is A or S; xii. the amino acid at position X12 is D or G; xiii. the amino acid at position X13 is G, E or R; xiv. the amino acid at position X14 is P or deleted; xv. the amino acid at position X15 is R or K; xvi. the amino acid at position X16 is S or E; xvii. the amino acid at position X17 is T, M or W; xviii. the amino acid at position X18 is A or G; xix. the amino acid at position X19 is A or E; xx. the amino acid at position X20 is S or T; xxi. the amino acid at position X21 is A or G; xxii. the amino acid at position X22 is L or M; xxiii. the amino acid at position X23 is A or S; xxiv. the amino acid at position X24 is W or H; xxv. the amino acid at position X25 is S or T; xxvi. the amino acid at position X26 is L or V; xxvii. the amino acid at position X27 is I or T; xxviii. the amino acid at position X28 is D or E; xxix. the amino acid at position X29 is H, R or D; xxx. the amino acid at position X30 is A or V; xxxi. the amino acid at position X31 is A or K; xxxii. the amino acid at position X32 is S or G;xxxiii. the amino acid at position X33 is E or D; xxxiv. the amino acid at position X34 is R, E or Q; xxxv. the amino acid at position X35 is A or P; xxxvi. the amino acid at position X36 is S or P; xxxvii. the amino acid at position X37 is A or E; xxxviii. the amino acid at position X38 is A or V; xxxix. the amino acid at position X39 is R or K; xl. the amino acid at position X40 is R, K or deleted; xli. the amino acid at position X41 is S, D or G; xlii. the amino acid at position X42 is P or E; xliii. the amino acid at position X43 is A or V; xliv. the amino acid at position X44 is L or I; xlv. the amino acid at position X45 is D or E; xlvi. the amino acid at position X46 is L or I; xlvii. the amino acid at position X47 is L or A; xlviii. the amino acid at position X48 is Q or E; xlix. the amino acid at position X49 is L or R;1. the amino acid at position X50 is I or V; li. the amino acid at position X51 is E, A or G; lii. the amino acid at position X52 is A or W; liii. the amino acid at position X53 is L or R; liv. the amino acid at position X54 is Q or T;Iv. the amino acid at position X55 is N or G;Ivi. the amino acid at position X56 is R, K or P;Ivii. the amino acid at position X57 is F or L;Iviii. the amino acid at position X58 is N or G; lix. the amino acid at position X59 is A or G; lx. the amino acid at position X60 is A or V;Ixi. the amino acid at position X61 is L or F;Ixii. the amino acid at position X62 is L or E;Ixiii. the amino acid at position X63 is C or N;Ixiv. the amino acid at position X64 is D, E or R; and / orIxv. the amino acid at position X65 is G or A.

[0078] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is D; iii. the amino acid at position 25 is Q; iv. the amino acid at position 26 is P; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is I; vii. the amino acid at position 51 is P; viii. the amino acid at position 52 is V; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is R; xi. the amino acid at position 58 is A; xii. the amino acid at position 63 is D; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is K; xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is M; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is E; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is S; xxiv. the amino acid at position 101 is H; xxv. the amino acid at position 110 is T; xxvi. the amino acid at position 114 is L; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 is R; xxx. the amino acid at position 148 is V; xxxi. the amino acid at position 149 is K; xxxii. the amino acid at position 167 is G; xxxiii. the amino acid at position 171 is E;xxxiv. the amino acid at position 186 is Q; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is S; xxxvii. the amino acid at position 197 is E; xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is deleted; xli. the amino acid at position 212 is S; xlii. the amino acid at position 215 is E; xliii. the amino acid at position 223 is V; xliv. the amino acid at position 234 is I; xlv. the amino acid at position 244 is E; xlvi. the amino acid at position 276 is L; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is Q; xlix. the amino acid at position 291 is L;1. the amino acid at position 301 is I; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is A; liii. the amino acid at position 311 is R; liv. the amino acid at position 312 is Q;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is K;Ivii. the amino acid at position 328 is F;Iviii. the amino acid at position 330 is N; lix. the amino acid at position 349 is G; lx. the amino acid at position 368 is A;Ixi. the amino acid at position 370 is F;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is E; andIxv. the amino acid at position 397 is A, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having aminoacid residues 2-397 of SEQ ID NO: 2. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 2.

[0079] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is D; iii. the amino acid at position 25 is T; iv. the amino acid at position 26 is P; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is V; vii. the amino acid at position 51 is A; viii. the amino acid at position 52 is A; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is A; xi. the amino acid at position 58 is S; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is R; xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is M; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is A; xx. the amino acid at position 84 is S; xxi. the amino acid at position 88 is A; xxii. the amino acid at position 92 is M; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is W; xxv. the amino acid at position 110 is S; xxvi. the amino acid at position 114 is V; xxvii. the amino acid at position 124 is I; xxviii. the amino acid at position 131 is D; xxix. the amino acid at position 143 Is H; xxx. the amino acid at position 148 is A;xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is S; xxxiii. the amino acid at position 171 is D; xxxiv. the amino acid at position 186 is R; xxxv. the amino acid at position 193 is A; xxxvi. the amino acid at position 194 is P; xxxvii. the amino acid at position 197 is A; xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is R; xl. the amino acid at position 211 is R; xli. the amino acid at position 212 is G; xlii. the amino acid at position 215 is P; xliii. the amino acid at position 223 is A; xliv. the amino acid at position 234 is I; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is I; xlvii. the amino acid at position 280 is A; xlviii. the amino acid at position 288 is E; xlix. the amino acid at position 291 is R; 1. the amino acid at position 301 is I; li. the amino acid at position 304 is G; lii. the amino acid at position 305 is W; liii. the amino acid at position 311 is L; liv. the amino acid at position 312 is T; Iv. the amino acid at position 313 is G; Ivi. the amino acid at position 316 is R; Ivii. the amino acid at position 328 is F; Iviii. the amino acid at position 330 is G; lix. the amino acid at position 349 is A; lx. the amino acid at position 368 is V; Ixi. the amino acid at position 370 is L; Ixii. the amino acid at position 375 is E; Ixiii. the amino acid at position 385 is N; Ixiv. the amino acid at position 390 is D; andIxv. the amino acid at position 397 is A wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 4. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 4.

[0080] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is D; iii. the amino acid at position 25 is T; iv. the amino acid at position 26 is P; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is V; vii. the amino acid at position 51 is A; viii. the amino acid at position 52 is A; ix. the amino acid at position 53 is W; x. the amino acid at position 57 is A; xi. the amino acid at position 58 is A; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is R; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is R; xvi. the amino acid at position 74 is S; xvii. the amino acid at position 75 is M; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is A; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is H; xxv. the amino acid at position 110 is T; xxvi. the amino acid at position 114 is L; xxvii. the amino acid at position 124 is T;xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is H; xxx. the amino acid at position 148 is V; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is G; xxxiii. the amino acid at position 171 is D; xxxiv. the amino acid at position 186 is E; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is S; xxxvii. the amino acid at position 197 is A; xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is deleted; xli. the amino acid at position 212 is S; xlii. the amino acid at position 215 is E; xliii. the amino acid at position 223 is V; xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is L; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is Q; xlix. the amino acid at position 291 is L;1. the amino acid at position 301 is I; li. the amino acid at position 304 is A; lii. the amino acid at position 305 is A; liii. the amino acid at position 311 is R; liv. the amino acid at position 312 is T;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is R;Ivii. the amino acid at position 328 is F;Iviii. the amino acid at position 330 is G; lix. the amino acid at position 349 is G; lx. the amino acid at position 368 is A;Ixi. the amino acid at position 370 is F;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is D; andIxv. the amino acid at position 397 is A, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-397 of SEQ ID NO: 5. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 5.

[0081] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N; iii. the amino acid at position 25 is Q; iv. the amino acid at position 26 is A; v. the amino acid at position 32 is S; vi. the amino acid at position 40 is V; vii. the amino acid at position 51 is P; viii. the amino acid at position 52 is V; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is R; xi. the amino acid at position 58 is A; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is K; xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is T; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is E; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is H;xxv. the amino acid at position 110 is S; xxvi. the amino acid at position 114 is V; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is H; xxx. the amino acid at position 148 is V; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is S; xxxiii. the amino acid at position 171 is E; xxxiv. the amino acid at position 186 is R; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is S; xxxvii. the amino acid at position 197 is A; xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is K; xli. the amino acid at position 212 is G; xlii. the amino acid at position 215 is P; xliii. the amino acid at position 223 is V; xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is I; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is E; xlix. the amino acid at position 291 is R;1. the amino acid at position 301 is I; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is A; liii. the amino acid at position 311 is L; liv. the amino acid at position 312 is T;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is K;Ivii. the amino acid at position 328 is L;Iviii. the amino acid at position 330 is N;lix. the amino acid at position 349 is A; lx. the amino acid at position 368 is V;Ixi. the amino acid at position 370 is L;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is R; andIxv. the amino acid at position 397 is G, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 6. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 6.

[0082] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is N; iii. the amino acid at position 25 is Q; iv. the amino acid at position 26 is P; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is I; vii. the amino acid at position 51 is P; viii. the amino acid at position 52 is V; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is R; xi. the amino acid at position 58 is S; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is K; xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is W; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is E; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G;xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is W; xxv. the amino acid at position 110 is T; xxvi. the amino acid at position 114 is V; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is R; xxx. the amino acid at position 148 is A; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is S; xxxiii. the amino acid at position 171 is E; xxxiv. the amino acid at position 186 is R; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is P; xxxvii. the amino acid at position 197 is A; xxxviii. the amino acid at position 207 is V ; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is K; xli. the amino acid at position 212 is G; xlii. the amino acid at position 215 is P; xliii. the amino acid at position 223 is V; xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is I; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is E; xlix. the amino acid at position 291 is R;1. the amino acid at position 301 is I; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is W; liii. the amino acid at position 311 is L; liv. the amino acid at position 312 is Q;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is R;Ivii. the amino acid at position 328 is F;Iviii. the amino acid at position 330 is N; lix. the amino acid at position 349 is A; lx. the amino acid at position 368 is V;Ixi. the amino acid at position 370 is L;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is D; andIxv. the amino acid at position 397 is A, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 7. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 7.

[0083] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is D; iii. the amino acid at position 25 is T; iv. the amino acid at position 26 is A; v. the amino acid at position 32 is S; vi. the amino acid at position 40 is V; vii. the amino acid at position 51 is A; viii. the amino acid at position 52 is A; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is A; xi. the amino acid at position 58 is A; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is G; xiv. the amino acid at position 70 is deleted; xv. the amino acid at position 73 is R; xvi. the amino acid at position 74 is S; xvii. the amino acid at position 75 is T; xviii. the amino acid at position 78 is A;xix. the amino acid at position 82 is A; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is H; xxv. the amino acid at position 110 is S; xxvi. the amino acid at position 114 is V; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is H; xxx. the amino acid at position 148 is A; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is S; xxxiii. the amino acid at position 171 is E; xxxiv. the amino acid at position 186 is R; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is S; xxxvii. the amino acid at position 197 is A; xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is R; xl. the amino acid at position 211 is R; xli. the amino acid at position 212 is G; xlii. the amino acid at position 215 is P; xliii. the amino acid at position 223 is V; xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is I; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is E; xlix. the amino acid at position 291 is R;1. the amino acid at position 301 is V; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is A;liii. the amino acid at position 311 is L; liv. the amino acid at position 312 is T;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is R;Ivii. the amino acid at position 328 is L;Iviii. the amino acid at position 330 is G; lix. the amino acid at position 349 is A; lx. the amino acid at position 368 is V;Ixi. the amino acid at position 370 is L;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is R; andIxv. the amino acid at position 397 is G, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-397 of SEQ ID NO: 8. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 8.

[0084] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is N; iii. the amino acid at position 25 is Q; iv. the amino acid at position 26 is P; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is I; vii. the amino acid at position 51 is P; viii. the amino acid at position 52 is V; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is R; xi. the amino acid at position 58 is S; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is K;xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is W; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is E; xx. the amino acid at position 84 is S; xxi. the amino acid at position 88 is A; xxii. the amino acid at position 92 is M; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is W; xxv. the amino acid at position 110 is S; xxvi. the amino acid at position 114 is V; xxvii. the amino acid at position 124 is I; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is R; xxx. the amino acid at position 148 is A; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is S; xxxiii. the amino acid at position 171 is E; xxxiv. the amino acid at position 186 is R; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is P; xxxvii. the amino acid at position 197 is A; xxxviii. the amino acid at position 207 is V ; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is K; xli. the amino acid at position 212 is G; xlii. the amino acid at position 215 is P; xliii. the amino acid at position 223 is A; xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is I; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is E; xlix. the amino acid at position 291 is R;1. the amino acid at position 301 is I; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is W; liii. the amino acid at position 311 is L; liv. the amino acid at position 312 is Q;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is R;Ivii. the amino acid at position 328 is F;Iviii. the amino acid at position 330 is N; lix. the amino acid at position 349 is A; lx. the amino acid at position 368 is V;Ixi. the amino acid at position 370 is L;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is D; andIxv. the amino acid at position 397 is A, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 9. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 9.

[0085] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is D; iii. the amino acid at position 25 is T; iv. the amino acid at position 26 is A; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is V; vii. the amino acid at position 51 is A; viii. the amino acid at position 52 is A; ix. the amino acid at position 53 is W; x. the amino acid at position 57 is R; xi. the amino acid at position 58 is A; xii. the amino acid at position 63 is G;xiii. the amino acid at position 64 is G; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is R; xvi. the amino acid at position 74 is S; xvii. the amino acid at position 75 is W; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is A; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is H; xxv. the amino acid at position 110 is T; xxvi. the amino acid at position 114 is L; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is D; xxix. the amino acid at position 143 Is D; xxx. the amino acid at position 148 is V; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is G; xxxiii. the amino acid at position 171 is D; xxxiv. the amino acid at position 186 is Q; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is S; xxxvii. the amino acid at position 197 is A; xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is R; xli. the amino acid at position 212 is S; xlii. the amino acid at position 215 is E; xliii. the amino acid at position 223 is V; xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is L;xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is Q; xlix. the amino acid at position 291 is L;1. the amino acid at position 301 is I; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is A; liii. the amino acid at position 311 is R; liv. the amino acid at position 312 is T;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is P;Ivii. the amino acid at position 328 is F;Iviii. the amino acid at position 330 is G; lix. the amino acid at position 349 is G; lx. the amino acid at position 368 is A;Ixi. the amino acid at position 370 is F;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is D; andIxv. the amino acid at position 397 is G, wherein the numbering is relative to the amino acid positions of SEQ ID NO: 1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 10. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 10.

[0086] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N; iii. the amino acid at position 25 is Q; iv. the amino acid at position 26 is P; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is I; vii. the amino acid at position 51 is P; viii. the amino acid at position 52 is V; ix. the amino acid at position 53 is S;x. the amino acid at position 57 is R; xi. the amino acid at position 58 is S; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is K; xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is W; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is E; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is W; xxv. the amino acid at position 110 is T; xxvi. the amino acid at position 114 is V; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is R; xxx. the amino acid at position 148 is A; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is S; xxxiii. the amino acid at position 171 is E; xxxiv. the amino acid at position 186 is R; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is P; xxxvii. the amino acid at position 197 is A; xxxviii. the amino acid at position 207 is V; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is K; xli. the amino acid at position 212 is D; xlii. the amino acid at position 215 is P; xliii. the amino acid at position 223 is V;xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is I; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is E; xlix. the amino acid at position 291 is R;1. the amino acid at position 301 is I; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is W; liii. the amino acid at position 311 is L; liv. the amino acid at position 312 is Q;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is R;Ivii. the amino acid at position 328 is F;Iviii. the amino acid at position 330 is N; lix. the amino acid at position 349 is A; lx. the amino acid at position 368 is V;Ixi. the amino acid at position 370 is L;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is D; andIxv. the amino acid at position 397 is A, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 11. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 11.

[0087] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is D; iii. the amino acid at position 25 is T; iv. the amino acid at position 26 is P; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is V;vii. the amino acid at position 51 is A; viii. the amino acid at position 52 is A; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is A; xi. the amino acid at position 58 is S; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is R; xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is W; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is A; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is W; xxv. the amino acid at position 110 is S; xxvi. the amino acid at position 114 is V; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is H; xxx. the amino acid at position 148 is A; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is S; xxxiii. the amino acid at position 171 is E; xxxiv. the amino acid at position 186 is R; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is P; xxxvii. the amino acid at position 197 is A; xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is R; xl. the amino acid at position 211 is R;xli. the amino acid at position 212 is G; xlii. the amino acid at position 215 is P; xliii. the amino acid at position 223 is V ; xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is I; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is E; xlix. the amino acid at position 291 is R;1. the amino acid at position 301 is I; li. the amino acid at position 304 is G; lii. the amino acid at position 305 is W; liii. the amino acid at position 311 is L; liv. the amino acid at position 312 is T;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is R;Ivii. the amino acid at position 328 is F;Iviii. the amino acid at position 330 is G; lix. the amino acid at position 349 is A; lx. the amino acid at position 368 is V;Ixi. the amino acid at position 370 is L;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is D; andIxv. the amino acid at position 397 is A, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 12. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 12.

[0088] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N; iii. the amino acid at position 25 is Q;iv. the amino acid at position 26 is P; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is I; vii. the amino acid at position 51 is P; viii. the amino acid at position 52 is V; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is R; xi. the amino acid at position 58 is S; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is K; xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is W; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is E; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is W; xxv. the amino acid at position 110 is T; xxvi. the amino acid at position 114 is V; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is R; xxx. the amino acid at position 148 is A; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is S; xxxiii. the amino acid at position 171 is E; xxxiv. the amino acid at position 186 is R; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is P; xxxvii. the amino acid at position 197 is A;xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is K; xli. the amino acid at position 212 is D; xlii. the amino acid at position 215 is P; xliii. the amino acid at position 223 is V ; xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is I; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is E; xlix. the amino acid at position 291 is R;1. the amino acid at position 301 is I; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is W; liii. the amino acid at position 311 is L; liv. the amino acid at position 312 is Q;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is R;Ivii. the amino acid at position 328 is F;Iviii. the amino acid at position 330 is N; lix. the amino acid at position 349 is A; lx. the amino acid at position 368 is V;Ixi. the amino acid at position 370 is L;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is D; andIxv. the amino acid at position 397 is A, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 13. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 13.

[0089] In an embodiment, the polypeptide comprises an amino acid sequence whereini. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N; iii. the amino acid at position 25 is Q; iv. the amino acid at position 26 is A; v. the amino acid at position 32 is S; vi. the amino acid at position 40 is V; vii. the amino acid at position 51 is P; viii. the amino acid at position 52 is V; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is A; xi. the amino acid at position 58 is A; xii. the amino acid at position 63 is G; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is K; xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is T; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is E; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is A; xxiv. the amino acid at position 101 is H; xxv. the amino acid at position 110 is S; xxvi. the amino acid at position 114 is L; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is H; xxx. the amino acid at position 148 is A; xxxi. the amino acid at position 149 is A; xxxii. the amino acid at position 167 is S; xxxiii. the amino acid at position 171 is E; xxxiv. the amino acid at position 186 is R;xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is S; xxxvii. the amino acid at position 197 is E; xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is K; xli. the amino acid at position 212 is G; xlii. the amino acid at position 215 is P; xliii. the amino acid at position 223 is V; xliv. the amino acid at position 234 is L; xlv. the amino acid at position 244 is D; xlvi. the amino acid at position 276 is I; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is E; xlix. the amino acid at position 291 is R;1. the amino acid at position 301 is I; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is A; liii. the amino acid at position 311 is L; liv. the amino acid at position 312 is T;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is K;Ivii. the amino acid at position 328 is L;Iviii. the amino acid at position 330 is N; lix. the amino acid at position 349 is A; lx. the amino acid at position 368 is V;Ixi. the amino acid at position 370 is L;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is R; andIxv. the amino acid at position 397 is G, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having aminoacid residues 2-398 of SEQ ID NO: 14. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 14.

[0090] In an embodiment, the polypeptide comprises an amino acid sequence wherein i. the amino acid at position 3 is T; ii. the amino acid at position 5 is D; iii. the amino acid at position 25 is Q; iv. the amino acid at position 26 is P; v. the amino acid at position 32 is A; vi. the amino acid at position 40 is V; vii. the amino acid at position 51 is P; viii. the amino acid at position 52 is V; ix. the amino acid at position 53 is S; x. the amino acid at position 57 is R; xi. the amino acid at position 58 is A; xii. the amino acid at position 63 is D; xiii. the amino acid at position 64 is E; xiv. the amino acid at position 70 is P; xv. the amino acid at position 73 is K; xvi. the amino acid at position 74 is E; xvii. the amino acid at position 75 is M; xviii. the amino acid at position 78 is G; xix. the amino acid at position 82 is E; xx. the amino acid at position 84 is T; xxi. the amino acid at position 88 is G; xxii. the amino acid at position 92 is L; xxiii. the amino acid at position 99 is S; xxiv. the amino acid at position 101 is H; xxv. the amino acid at position 110 is T; xxvi. the amino acid at position 114 is L; xxvii. the amino acid at position 124 is T; xxviii. the amino acid at position 131 is E; xxix. the amino acid at position 143 Is H; xxx. the amino acid at position 148 is V;xxxi. the amino acid at position 149 is K; xxxii. the amino acid at position 167 is G; xxxiii. the amino acid at position 171 is E; xxxiv. the amino acid at position 186 is Q; xxxv. the amino acid at position 193 is P; xxxvi. the amino acid at position 194 is S; xxxvii. the amino acid at position 197 is E; xxxviii. the amino acid at position 207 is A; xxxix. the amino acid at position 210 is K; xl. the amino acid at position 211 is K; xli. the amino acid at position 212 is S; xlii. the amino acid at position 215 is E; xliii. the amino acid at position 223 is V; xliv. the amino acid at position 234 is I; xlv. the amino acid at position 244 is E; xlvi. the amino acid at position 276 is L; xlvii. the amino acid at position 280 is L; xlviii. the amino acid at position 288 is Q; xlix. the amino acid at position 291 is L;1. the amino acid at position 301 is I; li. the amino acid at position 304 is E; lii. the amino acid at position 305 is A; liii. the amino acid at position 311 is R; liv. the amino acid at position 312 is T;Iv. the amino acid at position 313 is N;Ivi. the amino acid at position 316 is K;Ivii. the amino acid at position 328 is F;Iviii. the amino acid at position 330 is N; lix. the amino acid at position 349 is G; lx. the amino acid at position 368 is A;Ixi. the amino acid at position 370 is F;Ixii. the amino acid at position 375 is L;Ixiii. the amino acid at position 385 is C;Ixiv. the amino acid at position 390 is E; andIxv. the amino acid at position 397 is G, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96. In an embodiment, the polypeptide comprises an amino acid sequence having amino acid residues 2-398 of SEQ ID NO: 15. In another embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 15.

[0091] In one aspect disclosed herein, there is provided a polypeptide capable of hydrolysing an amide bond in a polyamide, wherein the polypeptide comprises an amino acid sequence of amino acid residues 2-394 of SEQ ID NO: 88 or an amino acid sequence that has more than 75% sequence identity thereto. In one embodiment, the polypeptide has at least 80% sequence identity to SEQ ID NO:88. In another embodiment, the polypeptide has at least 85% sequence identity to SEQ ID NO:88. In another embodiment, the polypeptide has at least 90 % sequence identity to SEQ ID NO: 88. In another embodiment, the polypeptide has at least 95 % sequence identity to SEQ ID NO:88. In an embodiment, the polypeptide comprises amino acid residues 2-392 of SEQ ID NO:3. In an embodiment, the polypeptide comprises or consists of amino acid sequence of SEQ ID NO:3. In an embodiment, the polypeptide comprises the amino acid residues 2-394 of SEQ ID NO:63. In an embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO:63. In an embodiment, the polypeptide comprises amino acid residues 2-395 of SEQ ID NO:48. In an embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 48. In an embodiment, polypeptide comprises the amino acid residues 2-394 of SEQ ID NO: 123. In an embodiment, the polypeptide comprises or consists of amino acid sequence of SEQ ID NO: 123.

[0092] In one aspect disclosed herein, there is provided a polypeptide capable of hydrolysing an amide bond in a polyamide, wherein the polypeptide comprises an amino acid sequence of amino acid residues 2-391 of SEQ ID NO:45 or an amino acid sequence that has at least 61% sequence identity thereto. In one embodiment, the polypeptide comprises the amino acid residues 2-391 of SEQ ID NO:25. In an embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO:25. In one embodiment, the polypeptide comprises the amino acid residues 2-391 of SEQ ID NO:22. In an embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO:22. In another embodiment, the polypeptide comprises an amino acid sequence that has at least 70% sequence identity to SEQ ID NO:45. In another embodiment, thepolypeptide comprises an amino acid sequence that has at least 75% sequence identity to SEQ ID NO:45. In another embodiment, the polypeptide comprises an amino acid sequence that has at least 80% sequence identity to SEQ ID NO:45. In another embodiment, the polypeptide comprises an amino acid sequence that has at least 85% sequence identity to SEQ ID NO:45. In another embodiment, the polypeptide comprises an amino acid sequence that has at least 90% sequence identity to SEQ ID NO:45. In another embodiment, the polypeptide comprises an amino acid sequence that has at least 95% sequence identity to SEQ ID NO:45. In an embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO:45.

[0093] Herein, the terms "peptide", "polypeptide", "protein", "enzyme" are to be understood as referring to a chain of amino acids linked by peptide bonds, irrespective of the number of amino acids forming said chain. The amino acids are typically represented by their one-letter or three-letters code, according to the following nomenclature: A: alanine (Ala); C: cysteine (Cys); D: aspartic acid (Asp); E: glutamic acid (Glu); F: phenylalanine (Phe); G: glycine (Gly); H: histidine (His); I: isoleucine (He); K: lysine (Lys); L: leucine (Leu); M: methionine (Met); N: asparagine (Asn); P: proline (Pro); Q: glutamine (Gin); R: arginine (Arg); S: serine (Ser); T: threonine (Thr); V: valine (Vai); W: tryptophan (Trp) and Y: tyrosine (Tyr).

[0094] The terms "mutant" and "variant" may be used interchangeably herein to refer to a polypeptide comprising an amino acid sequence that is derived from a wildtype or extant enzyme or from the ancestral sequence reconstruction process and may further comprise a modification or alteration (e.g., a substitution, insertion, and / or deletion), at one or more (e.g., several) positions and having enhanced activity in catalysing the hydrolysis of nylon, nylon polymer or nylon oligomers, when compared to extant nylonases. Such variants may be obtained by various techniques well known in the art, illustrative examples of which include site-directed mutagenesis, random mutagenesis and synthetic oligonucleotide construction. The terms "modification", "alteration", "substitution" and the like, as used herein in relation to an amino acid residue or position, typically mean that the amino acid in the particular position has been modified compared to the amino acid of the wild-type or parent polypeptide.

[0095] Suitable substitutions may include the replacement of an amino acid residue by another selected from the naturally-occurring standard 20 amino acid residues, rare naturally occurring amino acid residues (e.g., hydroxyproline, hydroxylysine, allohydroxylysine, 6-N-methylysine, N-ethylglycine, N-methylglycine, N-ethylasparagine, allo-isoleucine, N- methylisoleucine, N-methylvaline, pyroglutamine, aminobutyric acid, ornithine, norleucine, norvaline), and non-naturally occurring amino acid residue, often made synthetically, (e.g., cyclohexyl-alanine). Preferably, the substitution comprises the replacement of an amino acid residue by another selected from the naturally-occurring standard 20 amino acid residues (G, P, A, V, L, I, M, C, F, Y, W, H, K, R, Q, N, E, D, S and T). The modification or alteration may be identified herein using the following terminology: Y197V denotes that amino acid residue Tyrosine (Y) at position 197 of the parent polypeptide sequence is substituted for a Valine (V). Y197V / I / M denotes that amino acid residue Tyrosine (Y) at position 197 of the parent sequence may be substituted for one of the following amino acids: Valine (V), Isoleucine (I), or Methionine (M). The substitution can be a conservative or nonconservative substitution. Examples of conservative substitutions will be familiar to persons skilled in the art, illustrative examples of which include substitutions within the groups of basic amino acids (arginine, lysine and histidine), acidic amino acids (glutamic acid and aspartic acid), polar amino acids (glutamine, asparagine and threonine), hydrophobic amino acids (methionine, leucine, isoleucine, cysteine and valine), aromatic amino acids (phenylalanine, tryptophan and tyrosine), and small amino acids (glycine, alanine and serine).

[0096] Unless otherwise specified, the positions disclosed in the present application are numbered by reference to the amino acid sequence set forth in SEQ ID NO: 1 or SEQ ID NO: 96, both of which are 398 amino acids in length. In this context, the term "corresponding to", when used in reference to an amino acid position, is intended to mean an amino acid position in a polypeptide sequence when that position is aligned with the equivalent or corresponding position in the sequence set forth in SEQ ID NO:1 or SEQ ID NO:96. For example, the amino acid residue designated Xi in consensus sequence of SEQ ID NO:96 corresponds to the amino acid at position 3 of SEQ ID NO: 96, and vice versa; while the amino acid residue designated X2 in consensus sequence of SEQ ID NO:96 corresponds to the amino acid at position 5 of SEQ ID NO:96, and vice versa. In another example, amino acid at position 360 of consensus sequence of SEQ ID NO:96 corresponds to amino acid at position 359 of SEQ ID NO: 2 and SEQ ID NO:5 (see for example, Figure 9), once aligned.

[0097] As used herein, the term "sequence identity" or "identity" refers to the number (or fraction expressed as a percentage %) of matches (identical amino acid residues) betweentwo polypeptide sequences. In a preferred embodiment, the sequence identity is determined by comparing the sequences when aligned so as to maximize overlap and identity while minimizing sequence gaps. Sequence identity may be determined using any of a number of mathematical global or local alignment algorithms known to persons skilled in the art, depending on the length of the two sequences. Sequences of similar lengths may be aligned using a global alignment algorithms (e.g., Needleman and Wunsch algorithm; Needleman and Wunsch, 1970), which aligns the sequences optimally over the entire length, while sequences of substantially different lengths are preferably aligned using a local alignment algorithm (e.g., Smith and Waterman algorithm (Smith and Waterman, 1981) or Altschul algorithm (Altschul et al., 1997; Altschul et al., 2005)). Alignment for the purposes of determining percent amino acid sequence identity can be achieved by any means available to persons skilled in the art, illustrative examples of which include publicly available computer software, such as is available at hhp: / / biastncbi.nlm.nikgov / or Persons skilled in the art can readily determineappropriate parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared. As used herein, % sequence identity typically refers to values generated using pair wise sequence alignment that creates an optimal global alignment of two sequences (e.g., using the Needleman-Wunsch algorithm), where all search parameters are set to default values, e.g., Scoring matrix = BLOSUM62, Gap open = 10, Gap extend = 0.5, End gap penalty = false, End gap open = 10 and End gap extend = 0.5.

[0098] The present disclosure also extends to a composition comprising the polypeptide as described herein. The present disclosure also extends to a nucleic acid sequence encoding the polypeptide described herein. The present disclosure also extends to an expression vector comprising the nucleic acid sequence described herein. The present disclosure also extends to a host cell comprising the nucleic acid sequence or the expression vector described herein.

[0099] The term "recombinant", as used herein, typically refers to a nucleic acid construct, a vector, a polypeptide or a cell produced by genetic engineering.

[0100] The term "expression", as used herein, typically refers to any step involved in the production of a polypeptide, such as by transcription, post-transcriptional modification, translation, post-translational modification, and secretion.

[0101] The term "expression cassette" denotes a nucleic acid construct comprising a coding region, and suitably a regulatory region to which the coding region is operably linked.

[0102] The term "expression vector" typically means a DNA or RNA molecule that comprises an expression cassette. The expression vector may be a linear or circular double stranded DNA molecule.

[0103] As used herein, the term "nucleic acid", "nucleic sequence" "polynucleotide", "oligonucleotide" and "nucleotide sequence" are used interchangeably and refer to a sequence of deoxyribonucleotides and / or ribonucleotides. The nucleic acids can be DNA (cDNA or gDNA), RNA, or a mixture of the two. It can be in single stranded form or in duplex form or a mixture of the two. It can be of recombinant, artificial and / or synthetic origin and it can comprise modified nucleotides, comprising for example a modified bond, a modified purine or pyrimidine base, or a modified sugar. The nucleic acids of the invention can be in isolated or purified form, and made, isolated and / or manipulated by techniques known per se in the art, e.g., cloning and expression of cDNA libraries, amplification, enzymatic synthesis or recombinant technology. The nucleic acids can also be synthesized in vitro by well-known chemical synthesis techniques, as described in, e.g., Belousov (1997) Nucleic Acids Res. 25:3440-3444.

[0104] The nucleic acid sequences disclosed herein may suitably be codon optimized. Suitable methods for codon optimization will be familiar to persons skilled in the art, illustrative examples of which are described in the reference manual Sambrook et al. (Sambrook et al., 2001).

[0105] The nucleic acid sequences described herein maybe suitably deduced from the amino acid sequence of the polypeptides described herein and codon usage may be adapted according to the host cell in which the nucleic acid shall be transcribed.

[0106] In some embodiments, the nucleic acid sequences described herein may suitably comprise additional nucleotide sequences, such as regulatory regions, i.e., promoters, enhancers, silencers, terminators, signal peptides and the like that can be used to cause or regulate expression of the polypeptide in a selected host cell or system. Alternatively, or in addition, the nucleic acid sequences described herein may further comprise additional nucleotide sequences encoding fusion proteins, such as maltose binding protein (MBP) orglutathion S transferase (GST) that can be used to favor polypeptide expression and / or solubility.

[0107] As noted elsewhere herein, the present disclosure also extends to expression vectors and expression cassettes comprising the nucleic acid sequence described herein, optionally operably linked to one or more control sequences that direct the expression of the nucleic acid sequence in a suitable host cell. Typically, the expression vector or cassette comprises the nucleic acid sequence described herein operably linked to a control sequence such as transcriptional promoter and / or transcription terminator. The control sequence may include a promoter that is recognized by a host cell or an in vitro expression system for expression of the nucleic acid encoding the polypeptide described herein. The promoter will typically comprise a transcriptional control sequence that mediates the expression of the polypeptide. The promoter may be any polynucleotide that shows transcriptional activity in a host cell, including mutant, truncated, and hybrid promoters, and may suitably be obtained from genes encoding extracellular or intracellular polypeptides either homologous or heterologous to the host cell. The control sequence may also be a transcription terminator, which is recognized by a host cell to terminate transcription. The terminator is typically operably linked to the 3'-terminus of the nucleic acid encoding the polypeptide. Any terminator that is functional in the host cell may be used in this context. Typically, the expression vector or cassette comprises the nucleic acid sequence described herein operably linked to a transcriptional promoter and a transcription terminator.

[0108] The term "vector" typically refers to a DNA molecule used as a vehicle to transfer recombinant genetic material into a host cell. Suitable vectors include plasmids, bacteriophages, viruses, fosmids, cosmids, and artificial chromosomes. The vector is typically a DNA sequence that comprises an insert (a heterologous nucleic acid sequence, transgene) and a larger sequence that serves as the "backbone" of the vector. The purpose of a vector which transfers genetic information to the host is typically to isolate, multiply, or express the insert in the target cell. Expression vectors (also referred to as expression constructs) are specifically adapted for the expression of the heterologous sequences in the target cell, and generally have a promoter sequence that drives expression of the heterologous sequences encoding a polypeptide.

[0109] Generally, the regulatory elements that are used in an expression vector include a transcriptional promoter, a ribosome binding site, a terminator, and optionally presentoperator. An expression vector may further comprise an origin of replication for autonomous replication in a host cell, a selectable marker, a limited number of useful restriction enzyme sites, and a potential for high copy number. Suitable expression vectors will be familiar to persons skilled in the art, illustrative examples of which include cloning vectors, modified cloning vectors, plasmids and viruses. Expression vectors that are capable of providing suitable levels of polypeptide expression in different hosts are also well known in the art. The choice of the vector will typically depend on the compatibility of the vector with the host cell into which the vector is to be introduced.

[0110] The present disclosure also extends to a host cell comprising the nucleic acid sequence described herein. The host cell may be transformed, transfected or transduced in a transient or stable manner. The nucleic acid, expression cassette or vector is introduced into a host cell so that the nucleic acid, cassette or vector is maintained as a chromosomal integrant or as a self-replicating extra-chromosomal vector. The term "host cell" encompasses any progeny of a parent host cell that is not identical to the parent host cell due to mutations that occur during replication. The host cell may be any cell useful in the production of a variant of the present invention, e.g., a prokaryote or a eukaryote. The prokaryotic host cell may be any Gram-positive or Gram-negative bacterium. The host cell may also be a eukaryotic cell, such as a yeast, fungal, mammalian, insect or plant cell. In a particular embodiment, the host cell is selected from the group of Escherichia coli, Pseudomonas, Bacillus, Streptomyces, Trichoderma, Aspergillus, Saccharomyces, Pichia, Thermus or Yarrowia.

[0111] The nucleic acid, expression cassette or expression vector according to the invention may be introduced into the host cell by any suitable method known to persons skilled in the art, illustrative examples of which include electroporation, conjugation, transduction, competent cell transformation, protoplast transformation, protoplast fusion, biolistic "gene gun" transformation, PEG-mediated transformation, lipid-assisted transformation or transfection, chemically mediated transfection, lithium acetate-mediated transformation and liposome-mediated transformation.

[0112] In an embodiment, the host cell is a genetically modified host cell or microorganism. In this context, a host cell or microorganism may be genetically modified to enhance the expression of the polypeptide in which it is expressed and / or activity of the host cell. For example, the polypeptide described herein may be used to complement a wild type strain ofa fungus or bacteria known to be capable of nylonase activity, in order to improve and / or increase the nylonase activity of that strain.

[0113] The polypeptides as disclosed herein are capable of hydrolysing an amide bond in a polyamide. In an embodiment, the polypeptides are capable of hydrolysing a polyamide. In another embodiment, the polypeptides are capable of hydrolysing nylon, including nylon 6,6, as shown in the illustrative examples. Enzymes capable of digesting, hydrolysing or breaking down nylon, are broadly called nylonases. Nylonases were first discovered in bacteria capable of digesting byproducts of nylon 6 manufacture. The three nylonases were 6-aminohexanoate-cyclic-dimer hydrolase (El, NylA, UniProt: Pl 3398) 6-aminohexanoate- dimer hydrolase (Eli, NylB, UniProt: P07061) and 6-aminohexanoate-oligomer endohydrolase (EIII, NylC, UniProt: Q57326).

[0114] In some embodiments, the polypeptides disclosed herein have adipic acid mono- and di- N-alkyl amide hydrolase activity. In some embodiments, the polypeptides disclosed herein are capable to hydrolysing nylon 6,6 and nylon 6,6 oligomers. Methods for determining and measuring such activities, including adipic acid mono- and di- N-alkyl amide hydrolase activity, will be known to persons skilled in the art, illustrative examples of which are disclosed elsewhere herein.

[0115] The term "nylon" or "nylon polymer" or "nylon oligomers" refers to synthetic polymers composed of polyamides (repeating units linked by amide links). In one embodiment, the nylon or nylon polymer or nylon oligomer is nylon 6 (polycaprolactam). In another embodiment, the nylon or nylon polymer or nylon oligomer is nylon 6,6 (containing diamines and dicarboxylic acids). In one embodiment, the nylon or nylon polymer or nylon oligomer is a copolymer of different nylons, comprising of nylon 6 and / or nylon 6, 6. In another embodiment, the nylon or nylon polymer or nylon oligomer is a nylon blend of different nylons comprising of nylon 6 and / or nylon 6,6.

[0116] The term "polymer", as used herein, typically refers to a chemical compound or a mixture of compounds whose structure is made up of multiple monomers (repeat units) linked by covalent chemical bonds. Within the context of the invention, the term polymer includes natural or synthetic polymers, constituted of a single type of repeat unit (z.e., homopolymers) or of a mixture of different repeat units (z.e., copolymers or heteropolymers).

[0117] Oligomers are low molecular weight polymers containing few repeated units whose physical properties are typically dependent upon the length of the chain. The term "nylon oligomers" as used herein refers to a molecule of nylon containing a discrete number of repeating nylon monomer unit. In an embodiment, the nylon oligomer contains up to 50- mers of nylon. In another embodiment, the nylon oligomer contains up to 40-mers of nylon. In another embodiment, the nylon oligomer contains up to 30-mers of nylon. In another embodiment, the nylon oligomer contains up to 20-mers of nylon. In another embodiment, the nylon oligomer contains 20-mers, 19-mers, 18-mers, 17-mers, 16-mers, 15-mers, 14- mers ,13-mers, 12-mers, 11-mers, 10-mers of nylon. In another embodiment, the nylon oligomer contains 9-mers, 8-mers, 7-mers, or 6-mers, of nylon. In another embodiment, the nylon oligomer are pentamers, tetramers, trimers or dimers of nylon. In an embodiment, the nylon oligomer is a solubilised nylon oligomer. In an embodiment, the nylon oligomer is an aqueous soluble nylon oligomer. A solubilised nylon oligomer is an oligomer that is not in solid form, but is in solution (i.e. fluid or liquid form). That is the oligomer has dissolved in a liquid solution, usually through the addition of a solvent. In some embodiments, the oligomers are solubilised in an aqueous solution. In some embodiments, the oligomers are solubilised in a non-aqueous solution. In an embodiment, the solubilised nylon oligomer comprises 10-mers, 9-mers, 8-mers, 7-mers, or 6-mers, pentamers, tetramers, trimers and / or dimers of nylon. In an embodiment, the solubilised nylon oligomer comprises 10-mers, 9- mers, 8-mers, 7-mers, or 6-mers, pentamers, tetramers, trimers and / or dimers of nylon 6,6. In an embodiment, the solubilised nylon oligomer comprises 10-mers, 9-mers, 8-mers, 7- mers, or 6-mers, pentamers, tetramers, trimers and / or dimers of nylon 6. In an embodiment, the solubilised nylon oligomer contains pentamers, tetramers, trimers, or dimers. In an embodiment, the solubilised nylon oligomer contains pentamers, tetramers, trimers, or dimers of nylon 6,6. In an embodiment, the solubilised nylon oligomer contains pentamers, tetramers, trimers, or dimers of nylon 6. In some embodiments, the solubilised nylon oligomers is a mix of 10-mers, 9-mers, 8-mers, 7-mers, or 6-mers, pentamers, tetramers, trimers and / or dimers of nylon. In some embodiments, the solubilised nylon oligomers is a mix of 10-mers, 9-mers, 8-mers, 7-mers, or 6-mers, pentamers, tetramers, trimers and / or dimers of nylon 6,6. In some embodiments, the solubilised nylon oligomers is a mix of 10- mers, 9-mers, 8-mers, 7-mers, or 6-mers, pentamers, tetramers, trimers and / or dimers of nylon 6. In an embodiment, the solubilised nylon oligomer is a mix of pentamers, tetramers, trimers, or dimers of nylon. In an embodiment, the solubilised nylon oligomer is a mix of pentamers, tetramers, trimers, or dimers of nylon 6. In an embodiment, the solubilised nylonoligomer is a mix of pentamers, tetramers, trimers, or dimers of nylon 6,6. In an embodiment, the nylon oligomer is a tetramer. In an embodiment, the nylon oligomer is a trimer. In an embodiment, the nylon oligomer is a dimer. In another embodiment, the solubilised nylon oligomers contains tetramers, trimers or dimers. In an embodiment, the nylon oligomer is a tetramer of nylon 6. In an embodiment, the nylon oligomer is a trimer of nylon 6. In an embodiment, the nylon oligomer is a dimer of nylon 6. In another embodiment, the solubilised nylon oligomers contains tetramers, trimers or dimers of nylon 6. In an embodiment, the nylon oligomer is a tetramer of nylon 6,6. In an embodiment, the nylon oligomer is a trimer of nylon 6,6. In an embodiment, the nylon oligomer is a dimer of nylon 6,6. In another embodiment, the solubilised nylon oligomers contains tetramers, trimers or dimers of nylon 6,6.

[0118] As used herein, the terms "nylon-containing material", "nylon-containing product" and the like are to be understood as refers to a product, such as nylon product, comprising at least one nylon in crystalline, semi-crystalline or totally amorphous form. The nylon- containing material may refer to any item made from at least one nylon, such as nylon sheet, tube, rod, profile, shape, film, massive block, fibre, textiles, etc., which contains at least one nylon, and possibly other substances or additives, such as plasticizers, mineral or organic fillers. In an embodiment, the nylon-containing material is a textile or fabric comprising at least one nylon containing fibre. In another embodiment, the nylon-containing material is a nylon compound, or nylon formulation, in a molten or solid state, suitable for making a nylon product.

[0119] The nylon-containing material or nylon-containing product may comprise of nylon and one or more synthetic and / or natural materials. The nylon-containing material or nylon- containing product may comprise of nylon and another plastics such as polyesters, acrylics, polyurethanes, polyethylene (PE), polypropylene (PP), polystyrene (PS), and polyvinyl chloride (PVC). The nylon-containing material or nylon-containing product may comprise of nylon and polyethylene terephthalate (PET). The nylon-containing material or nylon- containing product may comprise of nylon and natural materials such as cotton, silk, cellulose, linen and wool.

[0120] In another aspect, the present disclosure provides a method of producing a polypeptide capable of hydrolysing an amide bond in a polyamide, the method comprising: i) providing the polynucleotide described herein; ii) expressing the polynucleotide in a hostcell under conditions sufficient to allow the host cell to produce the polypeptide; and iii) collecting the polypeptide produced by the host cell in ii).

[0121] In another aspect, the present disclosure provides a method of hydrolysing nylon polymer or nylon oligomer, the method comprising exposing the nylon polymer or nylon oligomer to the polypeptide, the composition or the host cell disclosed herein under conditions sufficient to convert the nylon-6, 6 polymer to adipic acid and I or hexamethylenediamine .

[0122] In another aspect, the present disclosure provides a method of degrading a nylon- containing product, the method comprising exposing the nylon-containing product to the polypeptide, the composition or the host cell as disclosed herein.

[0123] In an embodiment, the methods disclosed herein comprises i) chemical processing of nylon polymer, nylon oligomer or nylon-containing product to generate nylon dimer, trimer, and tetramer, and ii) exposing the nylon dimer, trimer, and tetramer, and pentamers generated in step (i) to the polypeptide, the composition or the host cell as disclosed herein, under conditions sufficient to produce adipic acid and I or hexamethylenediamine.

[0124] In an embodiment, the nylon in the nylon polymer, nylon oligomer or the nylon- containing product is a nylon 6 or a nylon 6,6. In another embodiment, the nylon is a nylon 6,6.

[0125] In another aspect, the present disclosure provides a method of hydrolysing a nylon 6,6 polymer, a nylon 6,6 oligomer or a nylon 6,6-comprising product, the method comprising exposing the nylon 6,6 polymer, nylon 6,6 oligomer or a nylon 6,6 comprising product to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73,74 and 87, or an amino acid sequence that has at least 70% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 73,74 and 87. In an embodiment, the polypeptide comprises an amino acid sequence that is at least 80% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 73,74 and 87. In an embodiment, the polypeptide comprises an amino acid sequence that is at least 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 73,74 and 87. In an embodiment, the polypeptide comprises an amino acid sequence that is at least 95% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 73,74 and 87. In an embodiment, the polypeptide comprises or consists of the amino acid residues 2-392 of SEQ ID NO: 73. In an embodiment, thepolypeptide comprises or consists of the amino acid residues 2-392 of SEQ ID NO: 74. In an embodiment, the polypeptide comprises the amino acid residues or consists of 2-392 of SEQ ID NO: 87. In an embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 73. In an embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 74. In an embodiment, the polypeptide comprises or consists of the amino acid sequence of SEQ ID NO: 87. In one embodiment, the polypeptide is a wildtype protein having NylB activity.

[0126] In another embodiment, the methods described herein further comprise recovering the adipic acid and I or the hexamethylenediamine produced in step (ii).

[0127] In another aspect, the present disclosure provides a composition comprising the adipic acid and I or the hexamethylenediamine recovered by the methods disclosed herein.

[0128] In another aspect, the present disclosure provides a method of producing a nylon polymer using the composition of adipic acid and I or the hexamethylenediamine recovered by the methods disclosed herein.

[0129] In the context of the present disclosure, reference to increased or enhanced activity hydrolysis of an amide bond in a polyamide, indicates an increased ability of the polypeptides / novel engineered polypeptides and its variants to hydrolyse nylon, nylon polymers or nylon oligomers, when compared to extant or wild-type NylB or nylonase enzymes. In an embodiment, the activity of the polypeptide described herein is increased by at least about 1%. In an embodiment, the activity of the polypeptide described herein is increased by at least about 5%, preferably by at least about 10%, preferably by at least about 20%, preferably by at least about 30%, preferably by at least about 40%, preferably by at least about 50%, preferably by at least about 100%, preferably by at least about 200%, preferably by at least about 300%, preferably by at least about 400%, preferably by at least about 500%, preferably by at least about 600%, preferably by at least about 700%, preferably by at least about 800%, preferably by at least about 900%, or more preferably by at least about 1,000% or more in comparison to extant or wild-type NylB or nylonase enzymes.

[0130] As noted elsewhere herein, the present inventors have engineered polypeptides with improved, increased or enhanced hydrolase activity, such as high activity, or broader activity on nylon polymers and nylon oligomers. These nylon oligomers may be solubilised nylon oligomers. The solubilised nylon oligomers may be dimers, trimers, tetramers, pentamers,hexamers, 7-mers, 8-mers, 9-mers or 1 Omers of nylon. The present disclosure is predicated, at least in part, on the inventors' surprising finding that said engineered polypeptides, from the ancestral sequence reconstruction of extant and ancestral variants of the NylB family, have one or more increased or enhanced properties relative to one or more of the extant enzymes. This increased or enhanced activity is also seen with substrates of nylon 6,6 oligomers. For example, in certain embodiments, the engineered polypeptides disclosed herein have increased activity in hydrolysing an amide bond in a polyamide; as shown in an example provided herein, an improved ability to hydrolyse amide bonds in nylon polyamide. In certain embodiments, the engineered polypeptides disclosed herein have increased thermal stability. This is a highly surprising discovery, including because the temperature conditions to which a hypothetical ancestral enzyme may have been exposed would not be too dissimilar to the temperature or conditions to which one or more corresponding extant enzymes are exposed. In certain embodiments, the engineered polypeptides disclosed herein are associated with increased recombinant expression in a host cell system, where such host cells are modified by insertion of a polynucleotide sequence encoding said enzymes. Presently known nylonase enzymes only demonstrate low levels of enzyme expression in common industrial host organisms.

[0131] The polypeptides disclosed here in have superior properties for use in industrial processes. With the aim of improving the activity of hydrolases, in particular those that can hydrolyse nylon, in conditions (i.e. heat, pH and pressure conditions) at which industrial degradation of nylon polymer and / or nylon-containing products are typically performed. The polypeptides disclosed herein capable of hydrolysing an amide bond in a polyamide (see Figure 1), and are particularly suited for the degradation of nylon 6,6 polymers, nylon 6,6, oligomers and / or nylon 6,6-containing materials or products. The engineered polypeptides as described herein have adipic acid mono- and di- N-alkyl amide hydrolase activity.

[0132] In the context of the present disclosure, reference to increased or enhanced activity may include one or more of the following: increased ability of the polypeptide to hydrolyse an amide bond in a polyamide, including nylons such as nylon 6 and nylon 6,6 when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73- 86; increased recombinant expression in a host cell system when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86; increased whole cellactivity when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86; and increased thermostability when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86.

[0133] In an embodiment, the polypeptide capable of hydrolysing an amide bond in a polyamide as disclosed herein demonstrates increased recombinant expression in a host cell system, when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86.

[0134] In another embodiment, the polypeptide capable of hydrolysing an amide bond in a polyamide as disclosed herein demonstrates increased thermostability compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86.

[0135] In another embodiment, the polypeptide capable of hydrolysing an amide bond in a polyamide as disclosed herein comprises increased whole cell activity when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86. As used herein, the term whole cell activity typically refers to the ability of the polyamide, just as nylon polymer or nylon oligomers when expressed in a host cell system.

[0136] In an embodiment, the activity of the polypeptide in hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers as disclosed herein is similar to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73- 86. In an embodiment, the activity of the polypeptide in hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers as disclosed herein described herein of the polypeptide described herein is increased by at least about 5%, preferably by at least about 10%, preferably by at least about 20%, preferably by at least about 30%, preferably by at least about 40%, preferably by at least about 50%, preferably by at least about 100%, preferably by at least about 200%, preferably by at least about 300%, preferably by at least about 400%, preferably by at least about 500%, preferably by at least about 600%, preferably by at least about 700%, preferably by at least about 800%, preferably by at least about 900%, or more preferably by at least about 1 ,000% or more to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86, or a wildtype NylB enzyme. In an embodiment, the activity of the polypeptide in hydrolysing nylon 6,6 oligomers as disclosed herein is similar to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86. In an embodiment, the activity of the polypeptide inhydrolysing nylon 6,6 oligomers as disclosed herein described herein of the polypeptide described herein is increased by at least about 5%, preferably by at least about 10%, preferably by at least about 20%, preferably by at least about 30%, preferably by at least about 40%, preferably by at least about 50%, preferably by at least about 100%, preferably by at least about 200%, preferably by at least about 300%, preferably by at least about 400%, preferably by at least about 500%, preferably by at least about 600%, preferably by at least about 700%, preferably by at least about 800%, preferably by at least about 900%, or more preferably by at least about 1 ,000% or more to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86, or a wildtype NylB enzyme. Suitable methods of determining or measuring enzyme activity of a polypeptide will be familiar to persons skilled in the art, an illustrative example of which is described elsewhere herein. Other methods of measuring nylon hydrolysis are described in Kiumarsi and Parvinzadeh, 2010 J Appl Polymer Sci, 116:3140 and Gashti et al., 2013 Preparative Biochemistry & Biotechnology, 43:798, the contents of which are incorporated herein by reference in their entirety. In an embodiment, the activity of the polypeptide in hydrolysing nylon 6,6 oligomers as disclosed herein is increased by at least about 5%, preferably by at least about 10%, preferably by at least about 20%, preferably by at least about 30%, preferably by at least about 40%, preferably by at least about 50%, preferably by at least about 100%, preferably by at least about 200%, preferably by at least about 300%, preferably by at least about 400%, preferably by at least about 500%, preferably by at least about 600%, preferably by at least about 700%, preferably by at least about 800%, preferably by at least about 900%, or more preferably by at least about 1,000% or more in comparison to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86, or a wildtype NylB enzyme, when determined by a colorimetric assay or LC-MC methods using solubilised nylon 6,6 oligomers, including dimers, trimers and tetramers as a substrate.

[0137] The activity of the polypeptide in hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers as disclosed herein may be assigned an absolute value or a value relative to the activity of a comparator (e.g., polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86, or a wildtype NylB enzyme. In an embodiment, the activity of the polypeptide is measured as the rate of monomers and / or oligomers is released over time, under suitable conditions of temperature, pH and buffer. In another embodiment, the activity of the polypeptide is measured as the rate of digestion ofthe substrate (i.e. measuring rate of change in the concentration or amounts of substrate) over time under suitable conditions of temperature, pH and buffer.

[0138] The activity of the polypeptide in hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers can be measured or assayed using a purified enzyme. Alternatively, enzyme activity can be measured as a function of the activity of the enzyme when recombinantly expressed in a host cell system (also referred to herein as cellular catalytic activity or whole cell activity).

[0139] Advantageously, the polypeptide described herein exhibits increased or enhanced recombinant expression in a host cell by at least about 5%, preferably by at least about 10%, preferably by at least about 20%, preferably by at least about 30%, preferably by at least about 40%, preferably by at least about 50%, preferably by at least about 100%, preferably by at least about 200%, preferably by at least about 300%, preferably by at least about 400%, preferably by at least about 500%, preferably by at least about 600%, preferably by at least about 700%, preferably by at least about 800%, preferably by at least about 900%, or more preferably by at least about 1,000% in comparison to polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86, or a wildtype NylB enzyme.

[0140] The polypeptide here displays increased thermostability, that is the ability of to resist irreversible change in enzyme function or activity (i.e. denature) when exposed to extreme temperatures. The polypeptide described herein retains its ability to hydrolyse an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers after exposure to a range of temperatures from about 4°C to about 70°C, preferably from about 10°C to about 70°C, preferably from about 20°C to about 60°C, more preferably from about 35°C to about 55°C, even more preferably from about 40°C to about 50°C, even more preferably at about 45°C. In an embodiment, the polypeptide described herein exhibits activity at a temperature from about 10°C to about 60°C, preferably from about 20°C to about 60°C, preferably from about 30°C to about 60°C, more preferably from about 40°C to about 60°C, even more preferably from about 40°C to about 50°C, or even more preferably at about 45°C. Advantageously, the polypeptide described herein exhibits the ability to hydrolyse an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers in a range of temperatures from about 10°C to about 80°C, preferably from about 20°C to about 70°C, preferably from about 30°C to about 60°C, more preferably from about 35°C to about 55°C, even more preferably from about 40°C to about 50°C, even morepreferably at about 45°C. In an embodiment, the polypeptide described herein exhibits activity at a temperature from about 10°C to about 70°C, preferably from about 20°C to about 60°C, preferably from about 30°C to about 60°C, more preferably from about 40°C to about 60°C, even more preferably from about 40°C to about 50°C, or even more preferably at about 45 °C. In an embodiment, the ability to hydrolyse an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers is measurable at a temperature between about 40°C and about 70°C, preferably between about 40°C and about 50°C, or even more preferably at about 45 °C. In another particular embodiment, the ability to hydrolyse an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers is measurable is still measurable at a temperature between about 10°C and about 30°C, preferably between about 15°C and about 28°C, corresponding to the mean temperature in the natural environment (ambient temperature).

[0141] In an embodiment, the polypeptide comprises the ability to hydrolyse an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers is measurable at a temperature from about 10°C to about 60°C, preferably from about 20°C to about 60°C, preferably from about 30°C to about 60°C, more preferably from about 40°C to about 60°C, even more preferably from about 40°C to about 50°C, or even more preferably at about 45°C of at least about 5%, preferably by at least about 10%, preferably by at least about 20%, preferably by at least about 30%, preferably by at least about 40%, preferably by at least about 50%, preferably by at least about 100%, preferably by at least about 200%, preferably by at least about 300%, preferably by at least about 400%, preferably by at least about 500%, preferably by at least about 600%, preferably by at least about 700%, preferably by at least about 800%, preferably by at least about 900%, or more preferably by at least about 1,000% or more in comparison to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86.

[0142] In another particular embodiment, the polypeptide described herein has increased activity in hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers, when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme, at a temperature of between about 10°C and about 70°C, preferably between about 20°C and about 60°C, preferably from about 30°C to about 60°C, preferably between about 40°C and about 60°C, preferably between about 40°C and about 50oC, or more preferably at about 45°C. In an embodiment,the polypeptide described herein is capable of hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers at between about 20°C to about 70°C of at least about 5%, preferably by at least about 10%, preferably by at least about 20%, preferably by at least about 30%, preferably by at least about 40%, preferably by at least about 50%, preferably by at least about 100%, preferably by at least about 200%, preferably by at least about 300%, preferably by at least about 400%, preferably by at least about 500%, preferably by at least about 600%, preferably by at least about 700%, preferably by at least about 800%, preferably by at least about 900%, or more preferably by at least about 1,000% or more in comparison to the activity of a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme.

[0143] In another embodiment, the polypeptide described herein has increased activity in hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers, compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme, at a temperature between about 10°C and about 60°C, preferably between about 20°C and about 50°C, even more preferably between about 20°C and about 40°C, or even more preferably at about 40°C. In an embodiment, the polypeptide described herein has activity in hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers at a temperature between about 10°C and about 40°C of at least about 5%, preferably by at least about 10%, preferably by at least about 20%, preferably by at least about 30%, preferably by at least about 40%, preferably by at least about 50%, preferably by at least about 100%, preferably by at least about 200%, preferably by at least about 300%, preferably by at least about 400%, preferably by at least about 500%, preferably by at least about 600%, preferably by at least about 700%, preferably by at least about 800%, preferably by at least about 900%, or more preferably by at least about 1,000% or more in comparison to the activity a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme at the same temperature.

[0144] In an embodiment, the polypeptide described herein exhibits a measurable activity in hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers at least in a range of pH from 5 to 11, preferably in a range of pH from 6 to 10, more preferably in a range of pH from 6.5 to 9, even more preferably in a range of pH from 7 to 8.

[0145] Advantageously, the thermostability of the polypeptide described herein is not significantly impaired compared to a polypeptide of polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme. In some embodiments, the thermostability of the polypeptide described herein is improved when compared to the thermostability of a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme. The term "improved thermostability" or "increased thermostability", as used herein, indicates an increased ability of the enzyme to resist changes in its chemical and / or physical structure at higher temperatures, more specifically at temperature between 40°C and 70°C, as compared a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme. In an embodiment, the polypeptides described herein have an increased half-life at a temperature between 40°C and 70°C, as compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme. The polypeptides described herein may exhibit a higher or equivalent melting temperature (Tm) as compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme. In some embodiments, the polypeptide described herein shows improved thermostability at a temperature of between 40°C and 70°C as compared to polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme.

[0146] The thermostability of a polypeptide may be evaluated by any suitable means known to persons skilled in the art. For example, thermostability can be assessed by measuring the residual enzyme activity of the polypeptide after incubation at different temperatures. The ability to perform multiple rounds of hydrolysis at different temperatures can also be evaluated. Differential Scanning Fluorimetry (DSF) may also be used to assess the thermostability of the polypeptide. Circular dichroism may also be used to measure thermostability of the polypeptides described herein, including their melting temperatures (Tm). The term "melting temperature (Tm)" is understood to mean a given protein corresponds to the temperature at which 50% of said protein is denatured.

[0147] In an embodiment, the polypeptide described herein exhibits a melting temperature (Tm) of from about 45°C to about 80°C, preferably from about 50°C to about 75°C, preferably from about 52°C to about 75°C. In an embodiment, the polypeptide described herein exhibits a melting temperature (Tm) that is lower than the melting temperature (Tm)exhibited by a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73- 86 or a wildtype NylB enzyme. In one embodiment, the polypeptide described herein exhibits a melting temperature (Tm) that is higher than the melting temperature (Tm) exhibited by a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73- 86 or a wildtype NylB enzyme.

[0148] The present disclosure also extends to a method of producing a polypeptide capable of hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers, the method comprising: a) providing a nucleic acid sequence as described herein; b) expressing the nucleic acid sequence in a host cell culture, thereby producing the polypeptide; and c) recovering the polypeptide produced in (b) from the host cell culture.

[0149] The present invention disclosure also extends to in vitro methods of producing the polypeptide described herein, the method comprising (a) contacting a nucleic acid, cassette or vector of the invention with an in vitro expression system; and (b) recovering the polypeptide produced. In vitro expression systems are well-known by the person skilled in the art and are commercially available.

[0150] Suitable host cells will be familiar to persons skilled in the art, illustrative examples of which include a recombinant Bacillus, recombinant E. coli, recombinant Pseudomonas, recombinant Aspergillus, recombinant Trichoderma, recombinant Streptomyces, recombinant Saccharomyces, recombinant Pichia, recombinant Thermus or recombinant Yarrowia. In an embodiment, the host cell is an E. coli.

[0151] The host cells may be cultivated in a nutrient medium suitable for production of polypeptides, using methods that will be known to persons skilled in the art. Suitable examples include cultivating the host cells by shake flask cultivation, or small-scale or large- scale fermentation (including continuous, batch, fed- batch, or solid state fermentations) in laboratory or industrial fermentors performed in a suitable medium and under conditions allowing the enzyme to be expressed and / or isolated. The cultivation will typically take place in a suitable nutrient medium, from commercial suppliers or prepared according to published compositions (e.g., in catalogues of the American Type Culture Collection) or any other culture medium suitable for cell growth.Where the polypeptide is expressed and / orsecreted into the nutrient medium, the polypeptide can be used in the form of a cellular I supernatant mixture, or in the form of a crude cell lysate. Alternatively, the polypeptide can be recovered directly from the culture supernatant. Conversely, the polypeptide can be recovered from cell lysates or after permeabilisation of the host cell membrane. The polypeptide may be recovered using any suitable method known to persons skilled in the art, illustrative examples of which include collection, centrifugation, filtration, extraction, spray-drying, evaporation, or precipitation. Optionally, the polypeptide may be partially or totally purified by a variety of procedures known in the art including, but not limited to, thermal shock, chromatography (e.g., ion exchange, affinity, hydrophobic, chromatofocusing, and size exclusion), electrophoretic procedures (e.g., preparative isoelectric focusing), differential solubility (e.g., ammonium sulfate precipitation), SDS- PAGE, or extraction to obtain substantially pure polypeptides.

[0152] The polypeptide may be used, in purified form, either alone or in combination with additional enzymes (e.g., PETases. MHETases or nylonases), to catalyze enzymatic reactions involved in the degradation and / or recycling of a nylon containing material. The polypeptides described herein may be in soluble form, or on solid phase. In particular, they may be bound to cell membranes or lipid vesicles, or to synthetic supports such as glass, plastic, polymers, filter membranes, e.g., in the form of beads, columns, plates and the like.

[0153] The present disclosure also extends to compositions comprising the polypeptide, the nucleic acid or the host cell described herein.

[0154] The composition may be liquid or dry, for instance in the form of a powder. In some embodiments, the composition is a lyophilisate. For instance, the composition may comprise the polypeptide, nucleic acid and / or host cells and optionally excipients and / or reagents etc. Suitable excipients may include buffers commonly used in biochemistry, agents for adjusting pH, preservatives such as sodium benzoate, sodium sorbate or sodium ascorbate, conservatives, protective or stabilizing agents such as starch, dextrin, arabic gum, salts, sugars e.g., sorbitol, trehalose or lactose, glycerol, polyethylene glycol, polyethene glycol, polypropylene glycol, propylene glycol, divalent ions such as calcium, sequestering agent such as EDTA, reducing agents (e.g., beta-mercaptoethanol, dithiothreitol, ascorbic acid, tris(2-carboxyethyl)phosphine), amino acids, a carrier such as a solvent or an aqueous solution, and the like.

[0155] In an embodiment, the composition comprises the polypeptide described herein (the polypeptide may be present in the composition in an isolated or at least partially purified form). In an embodiment, the composition comprises the polypeptide described herein in an amount of from about 0.1% to about 99.9%, preferably from about 0.1% to about 50%, preferably from about 0.1% to about 30%, preferably from about 0.1% to about 5% by weight of the total weight of the composition. In a preferred embodiment, the composition comprises the polypeptide described herein in an amount of from about 0.1 to about 5% by weight of the total weight of the composition. In another embodiment, the composition comprises the polypeptide described herein in an amount of from about 0.1 to about 0.2% by weight of the total weight of the composition. The amount of polypeptide in the composition may be suitably adapted by persons skilled in the art, depending e.g., on the nature and I or amount of the polyester containing material to be degraded (hydrolysed) and / or the presence or absence of any additional enzymes / polypeptides in the composition.

[0156] The compositions described herein may further comprise additional polypeptide(s) exhibiting enzymatic activity, not limited to nylonases.

[0157] In an embodiment, the polypeptide described herein is solubilized in an aqueous medium together with one or more excipients, such as excipients that may suitably stabilize or protect the polypeptide from degradation. For example, the polypeptides described herein may be solubilized in water and then admixed with excipients such as glycerol, sorbitol, dextrin, starch, glycol such as propanediol, salt, etc. The resulting admixture may then be dried so as to obtain a powder. Methods for drying such mixture are well known to the one skilled in the art and include, without limitation, lyophilisation, freeze-drying, spray-drying, supercritical drying, down-draught evaporation, thin-layer evaporation, centrifugal evaporation, conveyor drying, fluidized bed drying, drum drying or any combination thereof.

[0158] In an embodiment, the composition comprises at least one host cell expressing the polypeptide described herein, or an extract thereof. By "extract of a cell" is meant any fraction obtained from a cell, such as cell supernatant, cell debris, cell walls, DNA extract, enzymes or enzyme preparation or any preparation derived from cells by chemical, physical and / or enzymatic treatment, which is essentially free of living cells. Preferred extracts are enzymatically-active extracts. The composition may comprise one or several host cells orextract thereof containing the polypeptide described herein, and optionally one or several additional cells.

[0159] As noted elsewhere herein, the present inventors have surprisingly found that the polypeptides described herein have greater activity in hydrolysing an amide bond in a polyamide or in hydrolysing nylon 6 oligomers or nylon 6,6 oligomers when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86 or a wildtype NylB enzyme. Thus, disclosed herein is a method of hydrolysing nylon polymer or nylon oligomer or a nylon-containing product, the method comprising exposing nylon polymer or nylon oligomer or a nylon-containing product to the polypeptide, the composition or the host cell described herein, under conditions sufficient to convert the nylon polymer or nylon oligomer or a nylon-containing product, adipic acid and I or hexamethylenediamine.

[0160] The present disclosure extends to the use the polypeptide, the composition or the host cell described herein for degrading a nylon polymer or nylon oligomer or a nylon-containing product in aerobic or anaerobic conditions and / or recycling nylon containing material, as plastic products made of or containing nylon and / or producing biodegradable plastic products containing nylon. Such methods are particularly useful for degrading a nylon polymer or nylon oligomer or a nylon-containing product comprising nylon 6,6 polymer or nylon 6,6 oligomer or a nylon 6,6-containing product.

[0161] Advantageously, the nylon(s) of the nylon-containing material or nylon-containing product is (are) depolymerized up to monomers and / or oligomers. In an embodiment, at least one nylon polymer or nylon oligomer is degraded to yield re-polymerizable monomers and I or oligomers, which are advantageously retrieved or recovered for further use.

[0162] In an embodiment, nylon(s) of the nylon-containing material or nylon-containing product is (are) fully degraded.

[0163] As noted elsewhere herein, the nylon product may comprise at least one polyester selected from the group consisting of polyethylene terephthalate (PET), polylactic acid (PLA), polytrimethylene terephthalate (PTT), polybutylene terephthalate (PBT), polyethylene isosorbide terephthalate (PEIT), polyethylene terephthalate (PET), polyhydroxyalkanoate (PHA), polybutylene succinate (PBS), polybutylene succinate adipate (PBSA), polybutylene adipate terephthalate (PBAT), polyethylene furanoate (PEF),polycaprolactone (PCL), poly(ethylene adipate) (PEA) and combinations of any of the foregoing.

[0164] The time required for degrading a nylon-containing material or nylon-containing product may vary depending on the nylon-containing material or nylon-containing product itself (i.e., nature and origin of the material / product, its composition, shape etc.), the type and amount of polypeptide used, as well as various process parameters (i.e., temperature, pH, additional agents, etc.). One skilled in the art may easily adapt the process parameters to the polyester containing material.

[0165] Advantageously, the degrading process is implemented at a temperature from about 10°C to about 70°C, preferably from about 20°C to about 60°C, preferably from about 30°C to about 60°C, more preferably from about 40°C to about 60°C, even more preferably from about 40°C to about 50°C, or even more preferably at about 45°C. The temperature is typically be maintained below an inactivating temperature, which corresponds to the temperature at which the polypeptide is inactivated and / or the recombinant microorganism does not synthesize, produce or release the polypeptide described herein.

[0166] In an embodiment, the nylon polymer, a nylon oligomer or a nylon-comprising product or nylon-comprising material may be pretreated or pre-procesed prior to be contacted with the polypeptide in order to physically change its structure or solubility, so as to increase the surface of contact between the nylon and the enzyme. In an embodiment, the nylon polymer, a nylon oligomer or a nylon-comprising product or nylon-comprising material may be pre-treated or pre-processed so that the nylon is solubilised prior to being contacted with the polypeptide.

[0167] Monomers resulting from the depolymerization or degradation process or method may be suitably recovered, sequentially or continuously. A single type of monomers or several different types of monomers may be recovered, depending on the starting nylon polymer, nylon oligomer or nylon-containing material I product.

[0168] The recovered monomers may be further purified, using any suitable purifying method and conditioned in a repolymerizable form. Illustrative examples of suitable purifying methods include stripping process, separation by aqueous solution, steam selective condensation, filtration and concentration of the medium after the bioprocess, separation, distillation, vacuum evaporation, extraction, electrodialysis, adsorption, ion exchange,precipitation, crystallization, concentration and acid addition dehydration and precipitation, nanofiltration, acid catalyst treatment, semi continuous mode distillation or continuous mode distillation, solvent extraction, evaporative concentration, evaporative crystallization, liquid / liquid extraction, hydrogenation, azeotropic distillation process, acid or heat catalysed lactamisation, adsorption, column chromatography, simple vacuum distillation and microfiltration, combined or not.

[0169] The repolymerizable monomers may be used to synthesize new nylon polymers. Advantageously, nylons of the same nature are repolymerized. However, it is possible to mix the recovered monomers with other monomers, for example, in order to synthesize new copolymers of nylon. Alternatively, the recovered monomers may be used as chemical intermediates in order to produce new chemical compounds of interest.

[0170] The present disclosure also extends to a compound comprising the polypeptide, composition and I or host cell expressing said polypeptide or an extract thereof containing said polypeptide.

[0171] The present disclosure also extends to a masterbatch composition comprising the polypeptide, composition and I or host cell expressing said polypeptide or an extract thereof containing said polypeptide.

[0172] Advantageously, such compound or masterbatch composition described herein can be used for the production of a nylon containing material and / or plastic article that will include the polypeptide described herein.

[0173] In an embodiment, the resulting compound, masterbatch composition or nylon article is a biodegradable plastic compound, masterbatch composition or plastic article complying with at least one of the relevant standards and / or labels known by the person skilled in the art, such as standard EN 13432, standard ASTM D6400, OK Biodegradation Soil (Label Vincotte), OK Biodegradation Water (Label Vincotte), OK Compost (Label Vincotte), OK Home Compost (Label Vincotte).

[0174] The polypeptides disclosed herein are suitable for a range of applications, including industrial applications, illustrative examples of which include as additives in detergents, compositions, textiles production, electronics and biomedical applications. For example,the polypeptides disclosed herein can be employed in textile production, where it can be used as an exonuclease to suitably modify the properties of textile fibres.

[0175] The invention will now be described with reference to the following Examples which illustrate some preferred aspects of the present invention. However, it is to be understood that the particularity of the following description of the invention is not to supersede the generality of the preceding description of the inventionEXAMPLESExample 1: Ancestral sequence reconstruction

[0176] NylB sequences, P07061 from Flavobacterium sp. (strain K172) and P07062 from Flavobacterium sp. (strain K172) were used as query sequences for a blast search of the refseq protein database on NCBI, and the swissprot database, using an E-value threshold of 10e-10.

[0177] Redundancy was removed to 90% sequence identity using CD-HIT (Fu et al. 2012) and the non-redundant sequence dataset was aligned using the GINSI protocol of MAFFT with DASH structural homology enabled. 100 independent replicates of maximum likelihood model parameterization and tree-search were conducted in IQ-TREE, with empirical Bayesian ancestral sequence reconstruction. The maximum likelihood model was fitted by Bayesian information criteria as LG+F+R9. For all tree-search replicates, 1000 replicates of ultra-fast bootstrap approximation were performed. Tree-search and bootstrap approximation converged within the default algorithm parameters for all replicates. Empirical Bayesian ancestral sequence reconstruction was additionally performed in CodeML of the PAML software suite, using the LG+G4 sequence evolution model.

[0178] Indel events in ancestral sequences generated by IQ-TREE and CodeML were modelled by a Jukes-Cantor-like equal rates model that assumes the probability of insertion and deletion are equal, in the Ape package for R. Phylogenetic trees were visualized using GGtree in R. 172 ancestral and extant sequences were chosen for synthesis by their genealogical relationship to the extant sequences with NylB activity (Phase I sequences). These sequences (SEQ ID NOs: 2-72; see Table 1) spanned the full evolutionary trajectory from the last common ancestor through to the extant NylBs and were supported by high ultra-fast bootstrap approximations (~95) and mean posterior probabilities (~80%).Example 2: Protein expression and purification

[0179] Plasmids were transformed by heat shock into chemically competent E. cloni® cells (Lucigen) and plated onto Lysogeny broth (LB) agar supplemented with 100 pg / mL kanamycin and incubated at 37 °C overnight. A single colony was used to inoculate 1.5 mL LB media supplemented with 100 pg / mL kanamycin in a 2.2 mL 96- well deep well block and grown at 1050 rpm at 37 °C until OD ~0.8, then induced with 1 mM IPTG and grown at 1050 rpm at room temperature (RT; 25 °C) for an additional 16 hours.

[0180] Cells were harvested by centrifugation at 2000 x g for 15 minutes at RT and resuspended in Lysis Buffer (IX BugBuster® Protein Extraction Reagent (Merck- Millipore), 20 mM Tris, 300 mM NaCl, 1 U / ml Turbonuclease (Sigma) pH 8). The cell suspension was left to incubate at RT for 20 minutes with gentle shaking. The lysate was separated from the insoluble cell debris by centrifugation at 2250 x g for 1 hour at RT. The clarified lysate was then diluted with 100 pl of Equilibration Buffer (20 mM Tris, 300 mM NaCl pH 8) and purified by nickel-charged IMAC using a 96-well HisPur™Ni-NTA Spin Plate (ThermoFisher Scientific) equilibrated in Equilibration Buffer, washing the sample three times with 250 pl of Wash Buffer (20 mM Tris, 300 mM NaCl, 10 mM imidazole pH 8) and eluting with 250 pl of Elution Buffer (20 mM Tris, 300 mM NaCl, 150 mM imidazole pH 8). All centrifugation steps following addition of Wash or Elution Buffer were at 1000 x g for 1 minute at RT. The eluate was stored at 4 °C.Example 3: Colorimetric assay to measure activity and identification of candidate polypeptides having increased ability to hydrolyse an amide bond in a polyamide

[0181] 100 ul of 2 mM nylon 6,6 trimer in Reaction Buffer (45 mM NaHiPCU, 90 mM NaCl, pH 7.5) was pre-equilibrated to 40 °C. The reaction was initiated by the addition of 100 ul of a 1:10 dilution of the eluate from the 96-well Ni-NTA purification in Reaction Buffer, then incubated at 40 °C for 3 hours. The reaction was quenched by filtration through a 10 kDa MWCO centrifugal filter at 13,000 g for 15 minutes to remove the enzyme. 100 ul of the flow-through was added to 50 ul of a 0.01% solution of 2,4,6-trinitrobenzene sulfonic acid (TNBSA) in Reaction Buffer and the absorbance was measured at 335 nm using the Epoch Microplate Spectrophotometer (BioTek).

[0182] Many, if not all of the variant sequences identified by the ancestral sequence reconstruction demonstrated ability to hydrolyse amide bonds in a polyamide. However, a number of variant polypeptides (SEQ ID NOs: 2-16) were surprisingly found to haveincreased ability to hydrolyse amide bonds in a polyamide when compared to extant sequences (Figure 3).

[0183] Polypeptides of SEQ ID NOs: 2-16 were also tested for their recombinant expression levels. The expression yields are shown in Table 3.

[0184] Analysis of the sequences indicated that many of the polypeptides had increased ability to hydrolyse amide bonds in a polyamide when compared to extant sequences showed high sequence identity (see Figure 5) that is represented by the consensus sequence of SEQ ID NO:1:SEQ ID NO: 1:MTX1TX2LMQGFPPAPEQQVTLANWRX3X4PFNRWX5FHHVREIX6PTANIPRGPGX7 X8X9PLPX10X11PRDLX12X13IAFEGX14DGX15X16X17TVX18EMLX19EX20YTDX21FLVX2 2HRGRIVX23EX24YANGMTPHX25PHIX26FSVSKSITGX27LAGILVX28RGQLDPDAPV TX29YIPEX30X31GSAYGDATVRHVLDMTVX32IDFX33EDYLDPDGDFARYRX34ATG WNPX35X36DGX37TPSDLRSFLX38TLX39X40X41DGX42HGETFHYX43SPNSDLLGWIX4 4ERASGQRFAX45LLSEHIWQPMGAEHDAYITVDRLGAPRTAGGX46CATX47RDLAR FGX48MMX49NRGVANGRQX50VPX51X52WIDDIX53X54X55GDX56EAWARGDFAKFX5 7PX58GRYRSKWYVTGNARGAFCX59IGIHGQWIYIDPAAEVVIX60KX61SSQPX62PVD DAMDRLX63LAAFX64AIARALX65G(where X1-X65 is any amino acid)Example 4: Thermal stability

[0185] Thermostability of the polypeptides disclosed herein were measured using differential scanning fluorimetry or circular dichroism (CD) using a Chirascan™ spectropolarimeter (Applied Photophysics). In the differential scanning fluorimetry method, a 20 pl reaction containing lx Protein Thermal Shift™ Dye (ThermoFisher Scientific), lx Protein Thermal Shift™ Buffer (ThermoFisher Scientific) and 10 pM of purified enzyme was prepared and transferred to a MicroAmp™ EnduraPlate™ Optical 96-Well Clear Reaction plate (ThermoFisher Scientific). The QuantStudio 3 Real-Time PCR System (ThermoFisher Scientific) was used to measure fluorescence as the sample was heated from 25 to 90 °C at a rate of 0.05 °C / second. The data were analysed using the Protein Thermal Shift™ Software and a Boltzmann curve was fitted to the data to determine the Tm. In theCD method, proteins were diluted to 0.2 mg / mL in Reaction buffer. The path length of the cuvette was 1 mm. CD at 222 nm was measured at 0.5 °C intervals as the temperature was increased at 1 °C / min from 20 °C to 90 °C.

[0186] The mean Tm measured for extant variants of SEQ ID NOs: 75, 76, 78-80 and 86 were 51.4, 49.2, 48.4, 55.1, 41.8, 71.7, respectively. Polypeptides of SEQ ID NOs: 4 and 2 had mean Tm of 75.0, and 71.1 °C, respectively (Figure 4).Example 5: Engineering polypeptides with increased activity

[0187] To engineer variants with improved ability to hydrolyse amide bonds in a polyamide, a number of high-activity polypeptides from the first round of ancestral reconstruction in Example 1 were selected as a base for further engineering. This second round of engineering identified a number of new sequences (Phase II) that demonstrated improved activity on nylon 6,6, oligomers in whole-cell assays, when compared to extant sequences and SEQ ID NO:4 from the first round of ancestral reconstruction. A number of variants demonstrated high whole-cell activity. In Figure 7A, C4 corresponds to SEQ ID NO:90; D3 corresponds to a polypeptide comprising SEQ ID NO:91; E3 corresponds to a polypeptide comprising SEQ ID NO:95; F3 corresponds to a polypeptide comprising SEQ ID NO:92; F3 corresponds to a polypeptide comprising SEQ ID NO:92 and G3 corresponds to a polypeptide comprising SEQ ID NO:93.

[0188] A number of the candidate variants (including those with high whole-cell activity) were selected to assay for their specific enzyme activity on nylon 6,6 dimers, trimers and tetramers, using LC-MS. The activity of a selection of Phase II variants are shown in Figure 7B.

[0189] Assays of enzymatic activity against nylon 6,6 oligomers (dimer, trimer and tetramer) were conducted with 20 pM purified enzyme in a 200 pL reaction at 40 °C. Reactions were incubated for 1 hour at 40 °C and were stopped by removing the enzyme using 10 kDa MWCO centrifugal filters (15 min, 13,000 x g). The reactions were analysed using liquid-chromatography mass spectrometry (LC-MS) and compared to control reactions containing no enzyme. A number of Phase I enzymes were re-assayed in parallel with selected Phase II variants using the LC-MS method. The activity of variant polypeptides comprising SEQ ID NOs: 4, 48, 88, 63, 3, 89-94 in comparison to extant enzymes of SEQ ID NOs: 75-84 and 86 is shown in Figure 8. In particular, polypeptidescomprising SEQ ID NOs: 93, 90, 3, 63, 88 and 48 demonstrated improved activity on the nylon 6,6 oligomers.

[0190] Polypeptides comprising SEQ ID NOs: 3 and 63 showed elevated activity for nylon 6,6 dimers. Polypeptides comprising SEQ ID NOs: 3, 63, 92 and 93 showed elevated activity for nylon 6,6 trimers. Polypeptides comprising SEQ ID NOs: 90, 3, 63 and 93 showed elevated activity for nylon 6,6 tetramers.

[0191] Analysis of Phase I and Phase II sequences indicated that many of the polypeptides had increased ability to hydrolyse amide bonds in a polyamide when compared to extant sequences showed high sequence identity that can be represented by consensus sequence II of SEQ ID NO: 96 (below, and Figure 10). Table 2 provides the amino acid substitutions at the specified positions of the variant polypeptides, wherein the numbering is relative to the amino acid positions of SEQ ID NO: 1 or SEQ ID NO:96.SEQ ID NO: 96MTX1TX2LMQGX3PPAPEQX4VTLANWRX5X6PFX7RWX8FHHVREX9X10PTAX11IPR GPGX12X13X14PLPX15X16PRDLX17X18IAX19EGX20DGX21X22X23TVX24EMLX25EX26YT DX27FLVX28HRGRIVX29EX30YANGMTPHX31PHIX32FSVSKSITGX33LAGILVX34RG QLDPDAPVTX35YIPEX36X37GSAYGDATVRHVLDMTVX38IDFX39EDYLDPDGDFA RYRX40AX41GWNPX42X43DGX44TPSDLRSFLX45TLX46X47X48DGX49HGETFHYX50SP NSDLLGWIX51ERASGQRFAX52LLSEX53IWX54PMGAEX55DAYITVDRLGAPRTAG GX56CATX57RDLARFGX58MMX59NRGVANGRQX60VPX61X62WIDDIX63X64X65GDX6 6EAWARGDFAX67X68X69PX70GRYRSKWYVTGNARGAFCX71IGIHGQWIYX72DPAA EWIX73KX74SSQPX75PVDDAMDRLX76LAAFX77AIARALX78X79 (where X1-X79 is any amino acid)Example 6: Stability of engineered enzymes

[0192] For kinetic stability analysis, purified enzymes were incubated at 40°C, 50°C, 60°C, 70°C or 80°C for 15 minutes prior to reaction with nylon 6,6 trimer. The data is shown in Figure 9. Prior heat shock exposure at 50°C-60°C did not appear to result in significant loss of enzyme activity, though some variants did demonstrate a slight drop in enzyme activity at 60°C. A greater decrease in enzyme activity was observed with enzymes pre-exposed toa 70°C heat-shock. After 80°C heat-shock, the enzymes appear to have lost the capacity to convert nylon 6,6, trimer.Example 7: Acid hydrolysis of nylon 6,6 oligomers by immobilized enzyme

[0193] An engineered polypeptide was immobilised on an adsorption-based polymeric resin at an enzyme loading of 10% w / w. The immobilised enzyme was incubated with nylon 6,6 dimer, trimer and tetramer. The reaction solution was sampled after 1 hour for analysis by LC-MS and was compared to a control reaction containing no enzyme (time 0 hr). The data are shown in Figure 6.Example 8: Further enzyme engineering

[0194] Further rounds of enzyme engineering identified variant polypeptides that demonstrated activity in hydrolysing amide bonds in a polyamide, including nylon 6,6, oligomers. Assays of enzymatic activity against nylon 6,6 oligomers (dimer, trimer and tetramer) were conducted at 40 °C for 2 hours. Reactions were stopped by removing the enzyme using 10 kDa MWCO centrifugal filters (15 min, 13,000 x g). The reactions were analysed using ultra high performance liquid chromatography (UHPLC).

[0195] Polypeptide variants comprising amino acid sequences of any one of SEQ ID NOs: 97-123 were capable of hydrolysing nylon 6,6 oligomers. The activity of some of these polypeptides in hydrolysing nylon 6,6 trimers is demonstrated in Figure 11

[0196] A significant proportion of polypeptides capable of hydrolysing nylon 6,6 oligomers shared a consensus sequence of SEQ ID NO: 1241 consensus III.SEQ ID NO: 124MTQTNLX1QX2X3PPAX4EX5EVTLANWRQAPFSRWSFHHVRELVPTAQIPR X6PGPASPLPAAPX7X8LGEIALEGPDX9KEGTVAEMLEESYTDX10FLVLHRG RIVAEHYANGMTPHX11PHIVFSVSKSITX12TLAGILVEX13GQLDPX14APVT DYIPEVAGSAYGX15ATVRHVLDMTVX16IDFEEX17YLDPX18X19X20FARYRR AMGWNPPSX21GETPSDLX22SFX23ATLKKGX24GPHGETFHYX25SPNSDLLG WILERASGQRFADLLSERIWRPMGAEADAYX26TVDRLGAPRTAGGICX27T X28RDLARFGEMMRNRGVANGRQIVPEAWIX29DILTNGDX30EAWAX31GDFAHX32LPNGRYRX33KWYVTGNARGAFCAIGIHGQWIYX34DPAAEWIVKLS SQPLPVDX35AMDRX36MLAAFRAIAX37ALG(where X1-X37 is any amino acid)Example 9: Polypeptides capable of hydrolysing nylon 6,6 oligomers

[0197] Analysis of the polypeptides disclosed herein as capable of hydrolysing amide bonds in a polyamide showed significant sequence conservation that can be represented by consensus sequence IV of SEQ ID NO: 125 (Figure 13). This includes SEQ ID NOs:2, 4- 15, 89-95, and 97-122.SEQ ID NO: 125MTX1TX2LX3QX4X5PPAX6EX7X8VTLANWRX9X10PFX11RWX12FHHVREX13X 14PTAX15IPRX16PGX17X18X19PLPX20X21PX22X23LX24X25IAX26EGX27DX28X29X3 0X31TVX32EMLX33EX34YTDX35FLVX36HRGRIVX37EX38YANGMTPHX39PHIX 40FSVSKSITX41X42LAGILVX43X44GQLDPX45APVTX46YIPEX46X48GSAYGX49 ATVRHVLDMTVX50IDFX51EX52YLDPX53X54X55FARYRX56AX57GWNPX58X59 X60GX61TPSDLX62SFX63X64TLX65X66X67X68GX69HGETFHYX70SPNSDLLGWI X71ERASGQRFAX72LLSEX73IWX74PMGAEX75DAYX76TVDRLGAPRTAGGX7 7CX78TX79RDLARFGX80MMX81NRGVANGRQX82VPX83X84WIX85DIX86X87X88 GDX89EAWAX90GDFAX91X92X93PX94GRYRX95KWYVTGNARGAFCX96IGIH GQWIYX97DPAAEWDC98KX99SSQPX100PVDX101AMDRX102X103LAAFX104 AIAX105ALX106X107(where X1-X107 is any amino acid)

[0198] Table 4 shows the amino acid substitutions at the specified positions of the variant polypeptides, where the amino acid numbering is relative to the amino acid positions of SEQ ID NO: 1, SEQ ID NO:96 or SEQ ID NO: 125.

[0199] The disclosure of every patent, patent application, and publication cited herein is hereby incorporated herein by reference in its entirety.

[0200] The citation of any reference herein should not be construed as an admission that such reference is available as “Prior Art” to the instant application.

[0201] Throughout the specification the aim has been to describe the preferred embodiments of the invention without limiting the invention to any one embodiment or specific collection of features. Those of skill in the art will therefore appreciate that, in light of the instant disclosure, various modifications and changes can be made in the particular embodiments exemplified without departing from the scope of the present invention. All such modifications and changes are intended to be included within the scope of the appended claims.Table 1: Amino acid sequences of the disclosed polypeptides.IllTable 2: Amino acid substitutions across variant polypeptides0Table 3: Expression yieldTable 4: Amino acid substitutions across variant polypeptides

Claims

THE CLAIMS DEFINING THE INVENTION ARE AS FOLLOWS:

1. A polypeptide capable of hydrolysing an amide bond in a polyamide, wherein the polypeptide comprises an amino acid sequence of amino acid residues 2-398 of SEQ ID NO: 2 or an amino acid sequence that has at least 70% sequence identity thereto.

2. The polypeptide of claim 1, wherein the polypeptide comprises an amino acid sequence of amino acid residues 2-398 of SEQ ID NO: 125 or an amino acid sequence that has at least 80% sequence identity thereto.

3. The polypeptide of claim 1 or claim 2, wherein the polypeptide comprises an amino acid sequence of amino acid residues 2-398 of SEQ ID NO: 96 or an amino acid sequence that has at least 80% sequence identity thereto.

4. The polypeptide of any one of claims 1-3, wherein the polypeptide comprises an amino acid sequence of amino acid residues 2-398 of SEQ ID NO: 1 or an amino acid sequence that has at least 80% sequence identity thereto.

5. The polypeptide of any one of claims 1-4, wherein i. the amino acid at position 3 is Q or T; ii. the amino acid at position 5 is N or D; iii. the amino acid at position 7 is M or F; iv. the amino acid at position 9 is G or T; v. the amino acid at position 10 is S or F; vi. the amino acid at position 14 is P or A; vii. the amino acid at position 16 is Q or G viii. the amino acid at position 17 is Q or E; ix. the amino acid at position 25 is Q or T; x. the amino acid at position 26 is A or P; xi. the amino acid at position 29 is N or S; xii. the amino acid at position 32 is A or S; xiii. the amino acid at position 39 is I or L; xiv. the amino acid at position 40 is I or V;xv. the amino acid at position 44 is N or Q; xvi. the amino acid at position 48 is G or A xvii. the amino acid at position 51 is A or P; xviii. the amino acid at position 52 is A or V; xix. the amino acid at position 53 is S or W; xx. the amino acid at position 57 is A or R; xxi. the amino acid at position 58 is A or S; xxii. the amino acid at position 60 is R or A; xxiii. the amino acid at position 61 is D, A or G; xxiv. the amino acid at position 63 is D or G; xxv. the amino acid at position 64 is G, E or R; xxvi. the amino acid at position 67 is F or L xxvii. the amino acid at position 70 is P or deleted; xxviii. the amino acid at position 72 is G or A; xxix. the amino acid at position 73 is R or K; xxx. the amino acid at position 74 is S or E; xxxi. the amino acid at position 75 is T, M, G or W; xxxii. the amino acid at position 78 is A or G; xxxiii. the amino acid at position 82 is A or E; xxxiv. the amino acid at position 84 is S or T; xxxv. the amino acid at position 88 is A or G; xxxvi. the amino acid at position 92 is L or M; xxxvii. the amino acid at position 99 is A or S; xxxviii. the amino acid at position 101 is W or H; xxxix. the amino acid at position 110 is S or T; xl. the amino acid at position 114 is L or V; xli. the amino acid at position 123 is G or A; xlii. the amino acid at position 124 is I or T; xliii. the amino acid at position 131 is D or E; xliv. the amino acid at position 132 is R or D; xlv. the amino acid at position 138 is D or A; xlvi. the amino acid at position 143 is H, R or D; xlvii. the amino acid at position 148 is A or V; xlviii. the amino acid at position 149 is A or K;xlix. the amino acid at position 155 is D or G;1. the amino acid at position 167 is S or G; li. the amino acid at position 171 is E or D; lii. the amino acid at position 173 is D or A; liii. the amino acid at position 178 is D or A; liv. the amino acid at position 179 is G or S;Iv. the amino acid at position 180 is D or A;Ivi. the amino acid at position 186 is R, E or Q;Ivii. the amino acid at position 188 is T or M;Iviii. the amino acid at position 193 is A or P; lix. the amino acid at position 194 is S or P; lx. the amino acid at position 195 is D or P;Ixi. the amino acid at position 197 is A or E;Ixii. the amino acid at position 203 is R or Y;Ixiii. the amino acid at position 206 is L or I;Ixiv. the amino acid at position 207 is A or V;Ixv. the amino acid at position 210 is R or K;Ixvi. the amino acid at position 211 is R, K or deleted;Ixvii. the amino acid at position 212 is S, D or G;Ixviii. the amino acid at position 213 is D or A;Ixix. the amino acid at position 215 is P or E;Ixx. the amino acid at position 223 is A or V;Ixxi. the amino acid at position 234 is L or I;Ixxii. the amino acid at position 244 is D or E;Ixxiii. the amino acid at position 249 is H or R;Ixxiv. the amino acid at position 252 is Q or R;Ixxv. the amino acid at position 258 is H or A;Ixxvi. the amino acid at position 262 is I or V;Ixxvii. the amino acid at position 276 is L or I;Ixxviii. the amino acid at position 278 is A or C;Ixxix. the amino acid at position 280 is L, A or P;Ixxx. the amino acid at position 288 is Q or E;Ixxxi. the amino acid at position 291 is L or R;Ixxxii. the amino acid at position 301 is I or V;Ixxxiii. the amino acid at position 304 is E, A or G;Ixxxiv. the amino acid at position 305 is A or W;Ixxxv. the amino acid at position 308 is A or D;Ixxxvi. the amino acid at position 311 is L or R;Ixxxvii. the amino acid at position 312 is Q or T;Ixxxviii. the amino acid at position 313 is N or G;Ixxxix. the amino acid at position 316 is R, K or P; xc. the amino acid at position 321 is R or Q; xci. the amino acid at position 326 is K or H; xcii. the amino acid at position 327 is F or L; xciii. the amino acid at position 328 is F or L; xciv. the amino acid at position 330 is N or G; xcv. the amino acid at position 335 is S or N; xcvi. the amino acid at position 349 is A or G; xcvii. the amino acid at position 359 is I or V; xcviii. the amino acid at position 368 is A or V; xcix. the amino acid at position 370 is L or F; c. the amino acid at position 375 is L or E; ci. the amino acid at position 379 is D or A; cii. the amino acid at position 384 is L or E; ciii. the amino acid at position 385 is C, M or N; civ. the amino acid at position 390 is D, E or R; cv. the amino acid at position 394 is R or A; cvi. the amino acid at position 397 is G or A; and / or cvii. the amino acid at position 398 is G or deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1, SEQ ID NO:96 or SEQ ID NO: 125.

6. The polypeptide of any one of claims 1-5, wherein i. the amino acid at position 3 is Q or T; ii. the amino acid at position 5 is N or D; iii. the amino acid at position 10 is S or F; iv. the amino acid at position 17 is Q or E; v. the amino acid at position 25 is Q or T;vi. the amino acid at position 26 is A or P; vii. the amino acid at position 29 is N or S; viii. the amino acid at position 32 is A or S; ix. the amino acid at position 39 is I or L; x. the amino acid at position 40 is I or V; xi. the amino acid at position 44 is N or Q; xii. the amino acid at position 51 is A or P; xiii. the amino acid at position 52 is A or V; xiv. the amino acid at position 53 is S or W; xv. the amino acid at position 57 is A or R; xvi. the amino acid at position 58 is A or S; xvii. the amino acid at position 63 is D or G; xviii. the amino acid at position 64 is G, E or R; xix. the amino acid at position 67 is F or L xx. the amino acid at position 70 is P or deleted; xxi. the amino acid at position 73 is R or K; xxii. the amino acid at position 74 is S or E; xxiii. the amino acid at position 75 is T, M, G or W; xxiv. the amino acid at position 78 is A or G; xxv. the amino acid at position 82 is A or E; xxvi. the amino acid at position 84 is S or T; xxvii. the amino acid at position 88 is A or G; xxviii. the amino acid at position 92 is L or M; xxix. the amino acid at position 99 is A or S; xxx. the amino acid at position 101 is W or H; xxxi. the amino acid at position 110 is S or T; xxxii. the amino acid at position 114 is L or V; xxxiii. the amino acid at position 124 is I or T; xxxiv. the amino acid at position 131 is D or E; xxxv. the amino acid at position 143 is H, R or D; xxxvi. the amino acid at position 148 is A or V; xxxvii. the amino acid at position 149 is A or K; xxxviii. the amino acid at position 167 is S or G; xxxix. the amino acid at position 171 is E or D;xl. the amino acid at position 186 is R, E or Q; xli. the amino acid at position 188 is T or M; xlii. the amino acid at position 193 is A or P; xliii. the amino acid at position 194 is S or P; xliv. the amino acid at position 197 is A or E; xlv. the amino acid at position 207 is A or V; xlvi. the amino acid at position 210 is R or K; xlvii. the amino acid at position 211 is R, K or deleted; xlviii. the amino acid at position 212 is S, D or G; xlix. the amino acid at position 215 is P or E;1. the amino acid at position 223 is A or V; li. the amino acid at position 234 is L or I; lii. the amino acid at position 244 is D or E; liii. the amino acid at position 249 is H or R; liv. the amino acid at position 252 is Q or R;Iv. the amino acid at position 258 is H or A;Ivi. the amino acid at position 276 is L or I;Ivii. the amino acid at position 280 is L or A;Iviii. the amino acid at position 288 is Q or E; lix. the amino acid at position 291 is L or R; lx. the amino acid at position 301 is I or V;Ixi. the amino acid at position 304 is E, A or G;Ixii. the amino acid at position 305 is A or W;Ixiii. the amino acid at position 311 is L or R;Ixiv. the amino acid at position 312 is Q or T;Ixv. the amino acid at position 313 is N or G;Ixvi. the amino acid at position 316 is R, K or P;Ixvii. the amino acid at position 326 is K or H;Ixviii. the amino acid at position 327 is F or L;Ixix. the amino acid at position 328 is F or L;Ixx. the amino acid at position 330 is N or G;Ixxi. the amino acid at position 349 is A or G;Ixxii. the amino acid at position 359 is I or V;Ixxiii. the amino acid at position 368 is A or V;Ixxiv. the amino acid at position 370 is L or F;Ixxv. the amino acid at position 375 is L or E;Ixxvi. the amino acid at position 385 is C, M or N;Ixxvii. the amino acid at position 390 is D, E or R;Ixxviii. the amino acid at position 397 is G or A; and / orIxxix. the amino acid at position 398 is G or deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96.

7. The polypeptide of any one of claims 1-6; wherein i. the amino acid at position 3 is Q or T; ii. the amino acid at position 5 is N or D; iii. the amino acid at position 25 is Q or T; iv. the amino acid at position 26 is A or P; v. the amino acid at position 32 is A or S; vi. the amino acid at position 40 is I or V; vii. the amino acid at position 51 is A or P; viii. the amino acid at position 52 is A or V; ix. the amino acid at position 53 is S or W; x. the amino acid at position 57 is A or R; xi. the amino acid at position 58 is A or S; xii. the amino acid at position 63 is D or G; xiii. the amino acid at position 64 is G, E or R; xiv. the amino acid at position 70 is P or deleted; xv. the amino acid at position 73 is R or K; xvi. the amino acid at position 74 is S or E; xvii. the amino acid at position 75 is T, M or W; xviii. the amino acid at position 78 is A or G; xix. the amino acid at position 82 is A or E; xx. the amino acid at position 84 is S or T; xxi. the amino acid at position 88 is A or G; xxii. the amino acid at position 92 is L or M; xxiii. the amino acid at position 99 is A or S; xxiv. the amino acid at position 101 is W or H;xxv. the amino acid at position 110 is S or T; xxvi. the amino acid at position 114 is L or V; xxvii. the amino acid at position 124 is I or T; xxviii. the amino acid at position 131 is D or E; xxix. the amino acid at position 143 is H, R or D; xxx. the amino acid at position 148 is A or V; xxxi. the amino acid at position 149 is A or K; xxxii. the amino acid at position 167 is S or G; xxxiii. the amino acid at position 171 is E or D; xxxiv. the amino acid at position 186 is R, E or Q; xxxv. the amino acid at position 193 is A or P; xxxvi. the amino acid at position 194 is S or P; xxxvii. the amino acid at position 197 is A or E; xxxviii. the amino acid at position 207 is A or V; xxxix. the amino acid at position 210 is R or K; xl. the amino acid at position 211 is R, K or deleted; xli. the amino acid at position 212 is S, D or G; xlii. the amino acid at position 215 is P or E; xliii. the amino acid at position 223 is A or V; xliv. the amino acid at position 234 is L or I; xlv. the amino acid at position 244 is D or E; xlvi. the amino acid at position 276 is L or I; xlvii. the amino acid at position 280 is L or A; xlviii. the amino acid at position 288 is Q or E; xlix. the amino acid at position 291 is L or R; 1. the amino acid at position 301 is I or V; li. the amino acid at position 304 is E, A or G; lii. the amino acid at position 305 is A or W; liii. the amino acid at position 311 is L or R; liv. the amino acid at position 312 is Q or T; Iv. the amino acid at position 313 is N or G; Ivi. the amino acid at position 316 is R, K or P; Ivii. the amino acid at position 328 is F or L; Iviii. the amino acid at position 330 is N or G;lix. the amino acid at position 349 is A or G; lx. the amino acid at position 368 is A or V;Ixi. the amino acid at position 370 is L or F;Ixii. the amino acid at position 375 is L or E;Ixiii. the amino acid at position 385 is C or N;Ixiv. the amino acid at position 390 is D, E or R; and / orIxv. the amino acid at position 397 is G or A, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96.

8. The polypeptide of any one of claims 1-7, wherein i. the amino acid at position 3 is Q; ii. the amino acid at position 5 is N; iii. the amino acid at position 10 is S; iv. the amino acid at position 17 is E; v. the amino acid at position 25 is Q; vi. the amino acid at position 26 is A; vii. the amino acid at position 29 is S; viii. the amino acid at position 32 is S; ix. the amino acid at position 39 is L; x. the amino acid at position 40 is V; xi. the amino acid at position 44 is Q; xii. the amino acid at position 51 is P; xiii. the amino acid at position 52 is A; xiv. the amino acid at position 53 is S; xv. the amino acid at position 57 is A; xvi. the amino acid at position 58 is A; xvii. the amino acid at position 63 is G; xviii. the amino acid at position 64 is E; xix. the amino acid at position 67 is L; xx. the amino acid at position 70 is P; xxi. the amino acid at position 73 is K; xxii. the amino acid at position 74 is E; xxiii. the amino acid at position 75 is G;xxiv. the amino acid at position 78 is A; xxv. the amino acid at position 82 is E; xxvi. the amino acid at position 84 is S; xxvii. the amino acid at position 88 is G; xxviii. the amino acid at position 92 is L; xxix. the amino acid at position 99 is A; xxx. the amino acid at position 101 is H; xxxi. the amino acid at position 110 is S; xxxii. the amino acid at position 114 is V; xxxiii. the amino acid at position 124 is T; xxxiv. the amino acid at position 131 is E; xxxv. the amino acid at position 143 is D; xxxvi. the amino acid at position 148 is V; xxxvii. the amino acid at position 149 is A; xxxviii. the amino acid at position 167 is S; xxxix. the amino acid at position 171 is E; xl. the amino acid at position 186 is R; xli. the amino acid at position 188 is M; xlii. the amino acid at position 193 is P; xliii. the amino acid at position 194 is S; xliv. the amino acid at position 197 is E; xlv. the amino acid at position 207 is A; xlvi. the amino acid at position 210 is K; xlvii. the amino acid at position 211 is K; xlviii. the amino acid at position 212 is G; xlix. the amino acid at position 215 is P;1. the amino acid at position 223 is V ; li. the amino acid at position 234 is L; lii. the amino acid at position 244 is D; liii. the amino acid at position 249 is R; liv. the amino acid at position 252 is R;Iv. the amino acid at position 258 is A;Ivi. the amino acid at position 276 is I;Ivii. the amino acid at position 280 is L;Iviii. the amino acid at position 288 is E; lix. the amino acid at position 291 is R; lx. the amino acid at position 301 is I;Ixi. the amino acid at position 304 is E;Ixii. the amino acid at position 305 is A;Ixiii. the amino acid at position 311 is L;Ixiv. the amino acid at position 312 is T;Ixv. the amino acid at position 313 is N;Ixvi. the amino acid at position 316 is R;Ixvii. the amino acid at position 326 is H;Ixviii. the amino acid at position 327 is L;Ixix. the amino acid at position 328 is L;Ixx. the amino acid at position 330 is N;Ixxi. the amino acid at position 349 is A;Ixxii. the amino acid at position 359 is V ;Ixxiii. the amino acid at position 368 is V ;Ixxiv. the amino acid at position 370 is L;Ixxv. the amino acid at position 375 is L;Ixxvi. the amino acid at position 385 is M;Ixxvii. the amino acid at position 390 is R;Ixxviii. the amino acid at position 397 is G; andIxxix. the amino acid at position 398 is deleted, wherein the numbering is relative to the amino acid positions of SEQ ID NO:1 or SEQ ID NO:96.

9. The polypeptide of claim 8, comprising or consisting of the amino acid sequence of SEQ ID NO: 93.

10. The polypeptide of any one of claims 1-7, comprising the amino acid sequence of amino acid residues 2-394 of any one of SEQ ID NOs: 2, 4-15, 88-92 and 94-95.

11. The polypeptide of any one of claims 1-7, consisting of the amino acid sequence of any one of SEQ ID NOs: 2, 4-15, 88-92 and 94-95.

12. A polypeptide capable of hydrolysing an amide bond in a polyamide, wherein the polypeptide comprises an amino acid sequence of amino acid residues 2-394 of SEQ ID NO: 88 or an amino acid sequence that has at least 75% sequence identity thereto.

13. The polypeptide of claim 12 having at least 80% sequence identity to SEQ ID NO:88.

14. The polypeptide of claim 12 or claim 13, having at least 90% sequence identity to SEQ ID NO:88.

15. The polypeptide of any one of claims 12-14, having at least 95% sequence identity to SEQ ID NO:88.

16. The polypeptide of claim 12 or claim 13, comprising amino acid residues 2-394 of SEQ ID NO: 123.

17. The polypeptide of 16, consisting of amino acid sequence of SEQ ID NO: 123.

18. The polypeptide of claim 12 or claim 13, comprising amino acid residues 2-392 of SEQ ID NO:63.

19. The polypeptide of 18, consisting of amino acid sequence of SEQ ID NO:63.

20. The polypeptide of claim 12 or claim 13, comprising amino acid residues 2-392 of SEQ ID NO:3.

21. The polypeptide of 20, consisting of amino acid sequence of SEQ ID NO:3.

22. The polypeptide of any one of claims 1-21, wherein the polypeptide is capable of hydrolysing a nylon polymer.

23. The polypeptide of any one of claims 1-22, wherein the polypeptide has adipic acid mono- and di-N-alkyl amide hydrolase activity.

24. The polypeptide of claim 23, wherein the nylon polymer is a nylon oligomer.

25. The polypeptide of any one of claims 22-24, wherein the nylon polymer or nylon oligomer is a nylon 6 polymer, a nylon 6 oligomer, a nylon 6,6 polymer or a nylon 6,6 oligomer.

26. The polypeptide of claim 25, wherein the nylon polymer or the nylon oligomer is a nylon 6,6 polymer or a nylon 6,6 oligomer.

27. The polypeptide of claim 26, wherein the nylon 6,6 oligomer is an aqueous soluble nylon 6,6 oligomer.

28. The polypeptide of claim 26 or claim 27, wherein the nylon 6,6 oligomer is selected from the group consisting of a dimer, a trimer, a tetramer, a pentamer and hexamer of nylon 6,6.

29. The polypeptide of claim 28, wherein the nylon 6,6 oligomer is a nylon 6,6 trimer.

30. The polypeptide of claim 28, wherein the nylon 6,6 oligomer is a nylon 6,6 dimer.

31. The polypeptide of claim 28, wherein the nylon 6,6 oligomer is a nylon 6,6 tetramer.

32. The polypeptide of any one of claims 27-31, wherein the polypeptide is capable of hydrolysing the nylon 6,6 oligomer to produce adipic acid and hexamethylenediamine.

33. The polypeptide of any one of claims 1-32, further comprising: i. increased recombinant expression in a host cell system; ii. increased enzyme activity; and / or iii. increased thermostability, when compared to a polypeptide comprising the amino acid sequence of any one of SEQ ID NOs: 73-86.

34. The polypeptide of claim 33, having increased enzyme activity when compared to a polypeptide having the amino acid sequence of any one of SEQ ID NOs: 73-86.

35. A composition comprising the polypeptide of any one of claims 1-34.

36. A polynucleotide comprising a nucleic acid sequence encoding the polypeptide of any one of claims 1-34.

37. An expression vector comprising the polynucleotide of claim 36.

38. A host cell comprising the polynucleotide of claim 36, or the expression vector of claim 37.

39. A method of producing a polypeptide capable of hydrolysing an amide bond in a polyamide, the method comprising: i. providing the polynucleotide of claim 36; ii. expressing the nucleic acid sequence in a host cell culture, thereby producing the polypeptide; and iii. collecting the polypeptide produced in (ii) from the host cell culture.

40. A method of hydrolysing nylon polymer or nylon oligomer, the method comprising exposing the nylon polymer or nylon oligomer to the polypeptide of any one of claims 1 to 34, the composition of claim 35 or the host cell of claim 38 under conditions sufficient to convert the nylon-6, 6 polymer to adipic acid and / or hexamethylenediamine.

41. A method of degrading a nylon-containing product, the method comprising exposing the nylon-containing product to the polypeptide of any one of claims 1 to 34, the composition of claim 35 or the host cell of claim 38.

42. The method of claim 40 or claim 41, comprising: i. chemical processing of nylon polymer, nylon oligomer or nylon-containing product to generate nylon oligomers; ii. exposing the nylon oligomers generated in step (i) to the polypeptide of any one of claims 1 to 34, the composition of claim 35 or the host cell of claim 38, under conditions sufficient to produce adipic acid and / or hexamethylenediamine.

43. The method of any one of claims 40-42, wherein the nylon in the nylon polymer, nylon oligomer or the nylon-containing product is nylon 6 or nylon 6,6.

44. The method of claim 43, wherein the nylon is nylon 6,6.

45. The method of claim 44, wherein the nylon 6,6 oligomer is an aqueous soluble nylon 6,6 oligomer.

46. The method of any one of claims 42-45, wherein the nylon 6,6 oligomer is selected from the group consisting of a dimer, a trimer, a tetramer, a pentamer and hexamer of nylon 6,6.

47. The method of claim 46, wherein the nylon 6,6 oligomer is a nylon 6,6 trimer.

48. The method of claim 46, wherein the nylon 6,6 oligomer is a nylon 6,6 dimer.

49. The method of claim 46, wherein the nylon 6,6 oligomer is a nylon 6,6 tetramer.

50. A method of hydrolysing a nylon 6,6 polymer, a nylon 6,6 oligomer or a nylon 6,6- comprising product, the method comprising exposing the nylon 6,6 polymer, nylon 6,6 oligomer or nylon 6,6 comprising product to a polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 73, 74 and 87, or an amino acid sequence that has at least 70% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 73,74 and 87.

51. The method of claim 50, wherein the polypeptide comprises the amino acid residues 2- 392 of any one of SEQ ID NO:73, 74 and 87.

52. The method of any one of claims 42-49, further comprising recovering the adipic acid and / or the hexamethylenediamine produced in step (ii).

53. A composition comprising the adipic acid and / or the hexamethylenediamine recovered by the method of claim 52.

54. A method of producing a nylon polymer using the composition of claim 53.

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