Xyloglucanase from Malbranchea

• US 6,500,658 B1
• Filed: 09/01/2000
• Issued: 12/31/2002
• Est. Priority Date: 08/17/1999
• Status: Expired due to Fees

• Current Assignee: Novo Nordisk A/S
• Original Assignee: Novozymes A/S (Novozymes)
• Inventors: Stringer, Mary Ann, Wu, Wenping, Schulein, Martin, Kauppinen, Markus Sakari
• Primary Examiner(s): Prouty, Rebecca E.
• Assistant Examiner(s): Rao, Manjunath N.
• Application Number:
  US09/653,778
• Time in Patent Office: 851 Days
• Field of Search: 435/183, 435/200, 435/206, 435/209, 435/226, 435/252.3, 435/69.1, 435/262, 435/263, 435/264, 510/114
• US Class Current: 435/206
• CPC Class Codes: C11D 3/38636 containing enzymes other than protease, amylase, lipase, cellulase, oxidase or reductase
• CPC Class Codes: D06L 4/40 using enzymes
• CPC Class Codes: D06M 16/003 with enzymes or microorganisms

Abstract

An isolated or purified polypeptide having xyloglucanase activity which is obtained from a strain of the genus Malbranchea and has xyloglucanase activity in the pH range 4-11, measured at 50° C.; and/or a molecular mass of 25±10 kDa, as determined by SDS-PAGE; and/or an isoelectric point (pI) in the range 3-5; and/or an N-terminal sequence Ala-Asp-Phe-Cys-Gly-Gln-Xaa-Asp-Ser-Glu-Gln-Ser-Gly-Pro-Tyr-Ile-Val-Tyr-Asn-Asn-Leu is useful in industrial applications such as in laundry detergent compositions and for treatment of textiles.

Claims

1. An isolated or purified polypeptide having xyloglucanase activity which is obtained from a strain of Malbranchea and has

(a) xyloglucanase activity throughout the pH range of 4-11, measured at 50°

C.;

(b) a molecular mass of 25+/−

10 kDa, as determined by SDS-PAGE;

(c) an isoelectric point in the range of 3-5; and

(d) an N-terminal sequence of Ala-Asp-Phe-Cys-Gly-Gln-Trp-Asp-Ser-Glu-Gln-Ser-Gly-Pro-Tyr-Ile-Val-Tyr-Asn-Asn-Leu (amino acid residues 29-49 of SEQ ID NO;

     2).

2. The isolated or purified polypeptide of claim 1, which is obtained from a strain of a species selected from the group consisting of Malbranchea albolutea, Malbranchea arcuata, Malbranchea aurantiaca, Malbranchea bolognesii-chiurcoi, Malbranchea chrysosporioidea, Malbranchea cinnamomea (and the synonyms Malbranchea sulfurea, Malbranchea pulchella var. sulfurea and Thermoidium sulfureum), Malbranchea circinata, Malbranchea dendritica, Malbranchea filamentosa, Malbranchea flava, Malbranchea flavorosea, Malbranchea flocciformis, Malbranchea fulva, Malbranchea graminicola, Malbranchea gypsea, Malbranchea kambayashii, Malbranchea multicolor, Malbranchea pulchella, Malbranchea sclerotica, and Malbranchea setosa.

3. The isolated or purified polypeptide of claim 2, wherein the strain is a Malbranchea cinnamomea strain.

4. The isolated or purified polypeptide of claim 3, wherein the strain is selected from the group consisting of Malbranchea cinnamomea, CBS 115.68;

Malbranchea cinnamomea, CBS 343.55;

Malbranchea cinnamomea, CBS 423.54;

Malbranchea cinnamomea, CBS 960.72;

Malbranchea cinnamomea, MUCL 9500;

Malbranchea cinnamomea, MUCL 11291; and

Malbranchea cinnamomea, UAMH 3827.

5. The isolated or purified polypeptide of claim 1, wherein the polypeptide exhibits a relative xyloglucanase activity of at least 50% at pH 9.5 or of at least 40% at pH 10, compared to the activity at the pH optimum.

6. The isolated or purified polypeptide of claim 1, wherein the polypeptide belongs to family 12 of glycosyl hydrolases.

7. The isolated or purified polypeptide of claim 1, which exhibits no activity on carboxymethylcellulose (CMC) or microcrystalline cellulose (Avicel).

8. An enzyme preparation comprising the isolated or purified polypeptide of claim 1.

9. The preparation of claim 8, which further comprises one or more enzymes selected from the group consisting of alpha-amylases, cellobiohydrolases, cellulases (endoglucanases), cutinases, beta-glucanases, glucoamylases, hemicellulases, laccases, ligninases, lipases, other mannanases, oxidases, pectate lyases, pectinases, peroxidases, pectin acetyl esterases, pectin lyases, pectin methylesterases, phenoloxidases, polygalacturonases, proteases, pullulanases, reductases, rhamnogalacturonases, transglutaminases, xylanases, xyloglucanases, or mixtures thereof.

10. A detergent composition comprising a polypeptide of claim 1 and a surfactant.

11. A process for machine treatment of a fabric, comprising treating the fabric with a washing solution containing the polypeptide of claim 1.

12. A method of producing the polypeptide of claim 1, comprising

(a) culturing a strain of Malbranchea under conditions permitting the production of the xyloglucanase enzyme, and

(b) recovering the polypeptide from the culture.

13. An isolated or purified family 12 xyloglucanase selected from the group consisting of

(a) a polypeptide comprising a fragment encoded by the DNA sequence of positions 141-806 of SEQ ID NO:

1;

(b) a polypeptide produced by culturing a cell comprising the sequence of SEQ ID NO;

1 under conditions wherein the DNA sequence is expressed;

(c) a polypeptide having a sequence of at least 80% identity to positions 29-250 of SEQ ID NO;

2 wherein the identity is determined by GAP provided in the GCG program package version 10.0 using a GAP creation penalty of 8 and GAP extension penalty of 2.

14. The xyloglucanase of claim 13, which comprises an amino acid sequence of SEQ ID NO:

2.

15. The xyloglucanase of claim 13, which comprises an amino acid sequence of amino acid residues 29-250 of SEQ ID NO:

2.

16. An isolated xyloglucanase which is (i) free from homologous impurities, and (ii) produced by culturing a cell comprising a DNA sequence of positions 141-806 of SEQ ID NO:

1.