Enzyme with xylanase activity

• US 20080274231A1
• Filed: 05/03/2006
• Published: 11/06/2008
• Est. Priority Date: 02/21/2000
• Status: Active Grant

• Current Assignee: Puratos NV (Coprem NV)
• Original Assignee: Puratos NV (Coprem NV)
• Inventors: Jonniaux, Jean-Luc, Dauvrin, Thierry
• Granted Patent: US 7,998,720 B2
• Time in Patent Office: Days
• US Class Current: 426/11
• CPC Class Codes: A21D 2/267 Microbial proteins
• CPC Class Codes: A21D 8/042 with enzymes
• CPC Class Codes: C12N 9/2434 acting on beta-1,4-glucosidic bonds

Abstract

The present invention is related to an isolated and purified enzyme with xylanolytic activity having more than 70% homology, preferably more than 80% homology with the amino acid sequence SEQ ID NO: 11.

Claims

1. An isolated or purified enzyme with xylanolytic activity having more than 80% homology (sequence identity) with the amino acid sequence SEQ ID NO:

11.

2. The enzyme of claim 1, having more than 90% homology (sequence identity) with the amino acid sequence SEQ ID NO:

11.

3. The enzyme of claim 1, wherein the % sequence identity is over the length of the mature enzyme.

4. The enzyme of claim 1, which is a family 11 (formerly G) xylanase sequence.

5. The enzyme of claim 1, wherein one or more of the amino acids of SEQ ID NO:

11 have been modified.

6. The enzyme of claim 1, wherein about 5, about 10, about 13, about 15, about 16, about 17, up to about 20% of the amino acids of SEQ ID NO:

11 have been modified.

7. The enzyme of claim 5, wherein said modification may be deletion, an insertion and/or a substitution.

8. The enzyme of claim 7, wherein said modification is a substitution.

9. The enzyme of claim 8, wherein said substitution is a conservative substitution.

10. The enzyme of claim 8, wherein said substitution is a neutral substitution.

11. The enzyme of claim 7, wherein the first 8 amino acids of SEQ ID NO:

11 have been deleted.

12. The enzyme of claim 5, wherein the following preferably are not modified:

the first 30 amino acids at the N-terminal part of the mature enzyme, the Ser44, the three putative CK2-Phospho sites (at residues 3-6, 112-115, 132-135), the nine putative myristyl N-myristoylation sites, the putative N-glycosilation site (at residues 60-63) and the putative CAMP-Phospho site (at residues 144-147).

13. The enzyme of claim 1, having one or more amino acid substitutions at one or more of the following positions in the mature enzyme:

32, 40, 41, 44, 52, 53, 55, 56, 57, 58, 94, 100, 101, 102, 103, 104, 106, 107, 108, 109, 112, 113, 129, 131, 132, 140, 142, 143, 144, 145, 146, 147, 162, 164, 166, 168 or 180.

14. The enzyme of claim 1, comprising the amino acid sequence SEQ ID NO:

11, SEQ ID NO;

15, SEQ ID NO;

16, SEQ ID NO;

17 or SEQ ID NO;

18.

15. The enzyme of claim 1, having the amino acid sequence SEQ ID NO:

11, SEQ ID NO;

15, SEQ ID NO;

16, SEQ ID NO;

17 or SEQ ID NO;

18.

16. The enzyme of claim 1, wherein said enzyme has xylanolytic activity at a pH between about 4.5 and about 7.0 and optimum xylanolytic activity at a temperature between about 35 and about 55°

C.

17. The enzyme of claim 1, with an N-terminal sequence as defined in SEQ ID NO:

1.

18. The enzyme of claim 1, wherein the xylanase enzyme is a Penicillium enzyme.

19. The enzyme of claim 1, wherein the xylanase enzyme is a Penicillium griseofulvum enzyme.

20. An isolated or purified enzyme comprising SEQ ID NO:

11, SEQ ID NO;

15, SEQ ID NO;

16, SEQ ID NO;

17, SEQ ID NO;

18, or comprising the enzyme of claim 1.

21. An isolated or purified polynucleotide encoding the enzyme according to claim 1.

24. The polynucleotide of claim 21 comprising SEQ ID NO:

8.

25. The polynucleotide of claim 21 consisting of SEQ ID NO:

8.

26. A recombinant polynucleotide comprising the polynucleotide of claim 21, operably linked to one or more regulatory sequence(s).

27. The recombinant polynucleotide of claim 26, wherein said one or more regulatory sequences(s) originate from microorganisms which are homologous to that the polynucleotide originated from.

28. A vector comprising the polynucleotide of claim 21.

29. The vector of claim 28 comprising a plasmid incorporated in Escherichia coli and having the deposit number LMBP-3987.

30. A recombinant host cell transformed by the polynucleotide of claim 21.

31. The recombinant host cell of claim 30, wherein said host cell is selected from the group consisting of archaea, bacteria and fungi.

32. The recombinant host cell of claim 31, wherein said fungi is a yeast.

33. The recombinant host cell of claim 30, wherein said host cell expresses the enzyme of SEQ ID NO:

11, SEQ ID NO;

15, SEQ ID NO;

16, SEQ ID NO;

17, SEQ ID NO;

18, or a portion thereof intracellularly.

34. The recombinant host cell of claim 30 wherein said host cell expresses the enzyme of SEQ ID NO:

11, SEQ ID NO;

15, SEQ ID NO;

16, SEQ ID NO;

17, SEQ ID NO;

18, or a portion thereof extracellularly.

35. A solid support fixing an element selected from the group consisting of the host cell according to claim 30, a cell extract of the host cell according to claim 30 that comprises an enzyme with xylanolytic activity having more than 80% homology (sequence identity) with the amino acid sequence SEQ ID NO:

11, and the isolated or purified enzyme with xylanolytic activity expressed by the cell according to claim 30.

36. The solid support of claim 35 to which the isolated or purified enzyme with zyzlanolytic activity is fixed.

37. A method for the degradation of plant cell wall components comprising adding the enzyme of claim 1 to said plant cell wall components.

38. A method for the decomposition of plants and fruits comprising adding the enzyme of claim 1 to the preparation processes of fruit, legume juices, beer, paper, starch, gluten or vegetable oil.

39. A method for the decomposition of wastes comprising adding the enzyme of claim 1 to waste materials such as agricultural wastes or wastes from paper mills.

40. A method for increasing the volume of baked products comprising adding the enzyme of claim 1 to said baked products before baking.

41. A method for the separation of starch and gluten comprising adding the enzyme of claim 1 to a batter comprising starch and gluten.

22. An isolated or purified polynucleotide having more than 70% homology (sequence identity) with SEQ ID NO:

8, or a portion thereof, wherein said polynucleotide encodes a polypeptide having xylanolytic activity.

23. The polynucleotide of claim 22, wherein said polynucleotide has more than 80%, preferably more than 90% homology (sequence identity) with SEQ ID NO:

8.