Variants of polypeptides having cellulolytic enhancing activity and polynucleotides encoding same

Patent metadata

Field	Value
Publication number	US-10246691-B2
Application number	US-201113876759-A
Country	US
Kind code	B2
Filing date	Sep 29, 2011
Priority date	Sep 30, 2010
Publication date	Apr 2, 2019
Grant date	Apr 2, 2019

Abstract

The present invention relates to variants of GH61 polypeptides having cellulolytic enhancing activity, wherein the variants comprise a substitution at one or more positions corresponding to positions 90, 131, 134, and 141 of the full-length polypeptide of SEQ ID NO: 2, and have cellulolytic enhancing activity. The present invention also relates to polynucleotides encoding the variants; nucleic acid constructs, vectors, and host cells comprising the polynucleotides; and methods of using the variants.

First claim

What is claimed is:

1. A variant of a parent GH61 polypeptide having cellulolytic enhancing activity, comprising a substitution at one or more positions corresponding to positions 90, 131, 134, and 141 of the full-length polypeptide of SEQ ID NO: 2, wherein the substitution at a position corresponding to position 90 is with Val, the substitution at a position corresponding to position 131 is with Ser, the substitution at a position corresponding to position 134 is with Leu, and the substitution at a position corresponding to position 141 is with Trp, wherein the variant has cellulolytic enhancing activity and increased thermal activity and/or increased thermostability, and wherein the variant has
at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 2.

2. The variant of claim 1 , which comprises one or more substitutions selected from the group consisting of L90V, D131S, M134L, and A141W.

3. The variant of claim 1 , which comprises the substitutions L90V+D131S+M134L+A141W of the mature polypeptide of SEQ ID NO: 2.

4. An isolated polynucleotide encoding the variant of claim 1 .

5. An isolated recombinant host cell transformed with the polynucleotide of claim 4 .

6. A method of producing a variant, comprising:
(a) cultivating the recombinant host cell of claim 5 under conditions suitable for the expression of the variant; and
(b) recovering the variant.

7. A transgenic plant, plant part or plant cell transformed with the polynucleotide of claim 4 .

8. A method of producing the variant of claim 1 , comprising:
(a) cultivating a transgenic plant or a plant cell comprising a polynucleotide encoding the variant under conditions conducive for production of the variant; and
(b) recovering the variant.

9. A method for obtaining a variant of a parent GH61 polypeptide having cellulolytic enhancing activity, comprising introducing into the parent GH61 polypeptide a substitution at one or more positions corresponding to positions 90, 131, 134, and 141 of the full-length polypeptide of SEQ ID NO: 2, wherein the substitution at a position corresponding to position 90 is with Val, the substitution at a position corresponding to position 131 is with Ser, the substitution at a position corresponding to position 134 is with Leu, and the substitution at a position corresponding to position 141 is with Trp, wherein the variant has cellulolytic enhancing activity and increased thermal activity and/or increased thermostability, and wherein the variant has
at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 2.

10. A method for degrading a cellulosic material, comprising: treating the cellulosic material with an enzyme composition comprising the variant of claim 1 .

11. The method of claim 10 , further comprising recovering the degraded cellulosic material.

12. A method for producing a fermentation product, comprising:
(a) saccharifying a cellulosic material with an enzyme composition comprising the variant of claim 1 ;
(b) fermenting the saccharified cellulosic material with one or more fermenting microorganisms to produce the fermentation product; and
(c) recovering the fermentation product from the fermentation.

13. A method of fermenting a cellulosic material, comprising: fermenting the cellulosic material with one or more fermenting microorganisms, wherein the cellulosic material is saccharified with an enzyme composition comprising the variant of claim 1 .

14. The method of claim 13 , wherein the fermenting of the cellulosic material produces a fermentation product.

15. The method of claim 14 , further comprising recovering the fermentation product from the fermentation.

16. A detergent composition comprising the variant of claim 1 and a surfactant.

17. A whole broth formulation or cell culture composition comprising the variant of claim 1 .

18. The variant of claim 1 , wherein the variant has at least 96% sequence identity to the mature polypeptide of SEQ ID NO: 2.

19. The variant of claim 1 , wherein the variant has at least 97% sequence identity to the mature polypeptide of SEQ ID NO: 2.

20. The variant of claim 1 , which comprises a substitution at two positions corresponding to any of positions 90, 131, 134, and 141.

21. The variant of claim 1 , which comprises a substitution at three positions corresponding to any of positions 90, 131, 134, and 141.

22. The variant of claim 1 , which comprises a substitution at each position corresponding to positions 90, 131, 134, and 141.

23. The method of claim 10 , wherein the enzyme composition further comprises one or more enzymes selected from the group consisting of a cellulase, a polypeptide having cellulolytic enhancing activity, a hemicellulase, an expansin, an esterase, a laccase, a ligninolytic enzyme, a pectinase, a peroxidase, a protease, and a swollenin.

24. The method of claim 12 , wherein the enzyme composition further comprises one or more enzymes selected from the group consisting of a cellulase, a polypeptide having cellulolytic enhancing activity, a hemicellulase, an expansin, an esterase, a laccase, a ligninolytic enzyme, a pectinase, a peroxidase, a protease, and a swollenin.

25. The method of claim 13 , wherein the enzyme composition further comprises one or more enzymes selected from the group consisting of a cellulase, a polypeptide having cellulolytic enhancing activity, a hemicellulase, an expansin, an esterase, a laccase, a ligninolytic enzyme, a pectinase, a peroxidase, a protease, and a swollenin.

26. The variant of claim 1 , wherein the variant has at least 98% sequence identity to the mature polypeptide of SEQ ID NO: 2.

27. The variant of claim 1 , wherein the variant has at least 99% sequence identity to the mature polypeptide of SEQ ID NO: 2.

Assignees

• Sweeney Matthew
• Wogulis Mark
• Novozymes Inc

Inventors

• Sweeney Matthew
• Wogulis Mark

Classifications

• C12Y302/01004

  Cellulase (3.2.1.4), i.e. endo-1,4-beta-glucanase · CPC title

• C12N9/18 Primary

  Carboxylic ester hydrolases {(3.1.1)} · CPC title

• D21C5/005

  Treatment of cellulose-containing material with microorganisms or enzymes · CPC title

• C12Y302/01091

  Cellulose 1,4-beta-cellobiosidase (3.2.1.91) · CPC title

• C12Y302/01021

  Beta-glucosidase (3.2.1.21) · CPC title