Laccases for bio-bleaching

Patent metadata

Field	Value
Publication number	US-9353477-B2
Application number	US-201514591449-A
Country	US
Kind code	B2
Filing date	Jan 7, 2015
Priority date	Sep 1, 2006
Publication date	May 31, 2016
Grant date	May 31, 2016

Abstract

Provided herein are isolated laccase enzymes and nucleic acids encoding them. Also provided are mediators for laccase reactions. Also provided herein are methods for using laccases to oxidize lignins and other phenolic and aromatic compounds, such as for bio-bleaching and decolorization of wood pulp under high temperature and pH conditions to facilitate a substantial reduction in use of bleaching chemicals, as well as for treatment of fibers.

First claim

What is claimed is:

1. An isolated polypeptide, comprising:
an amino acid sequence having 90-99% sequence identity to the amino acid sequence of the polypeptide of SEQ ID NO: 2, wherein said isolated polypeptide comprises laccase activity, oxidizes lignin under conditions of pH greater than or equal to pH 8, and retains laccase activity for at least about 5 minutes at greater than or equal to 60° C.

2. The isolated polypeptide of claim 1 , lacking its associated signal peptide.

3. A method of delignifying a composition comprising lignin, comprising:
contacting a phenolic substrate with the polypeptide of claim 1 .

4. The method of claim 3 , further comprising contacting the composition with a mediator.

5. The method of claim 4 , wherein said mediator is a compound represented by Formula I or Formula II, wherein Formula I is represented by

wherein R 1 , R 2 R 3 , R 4 , R 5 and R 6 are independently selected from the group consisting of H, CH 3 , CH 2 CH 3 , CH 2 CH 2 CH 3 , CH(CH 3 ) 2 , CH 2 CH 2 (CH 3 ) 2 , C(CH 3 ) 3 , CH 2 CH 2 CH 2 , CH 3 , alkyl, aryl, phenyl, CF 3 , OC(CH 3 ) 3 , OCH(CH 3 ) 2 , CH 2 CH 3 , C(O)OCH 3 , COOH, OCH 3 , OH, OCF 3 , NH 2 , NH(CH 3 ), N(CH 3 ) 2 , CH 2 NH 2 , CH 2 CH 2 NH 2 , Br, and Cl, and wherein Formula II is represented by

wherein R 1 , R 2 R 3 , R 4 , R 5 R 6 , R 7 , R 8 , R 9 and R 10 are independently selected from the group consisting of H, CH 3 , CH 2 CH 3 , CH 2 CH 2 CH 3 , CH(CH 3 ) 2 , CH 2 CH 2 (CH 3 ) 2 , C(CH 3 ) 3 , CH 2 CH 2 CH 2 , CH 3 , alkyl, aryl, phenyl, CF 3 , OC(CH 3 ) 3 , OCH(CH 3 ) 2 , CH 2 CH 3 , C(O)OCH 3 , COOH, OCH 3 , OH, OCF 3 , NH 2 , NH(CH 3 ), N(CH 3 ) 2 , CH 2 NH 2 , CH 2 CH 2 NH 2 , Br, and Cl.

6. The method of claim 4 , wherein the mediator is

7. The method of claim 4 , wherein the mediator is selected from the group consisting of violuric acid; 2,6,6-tet-rarmethylpiperidein-1-yloxy (TEMPO); 1-hydroxybenzotriazole (HBT); 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS); syringaldazine; N-benzoyl-N-phenyl hydroxylamine (BPHA); N-hydroxyphthalimide, 3-Hydroxy-1,2,3-benzotriazin-4-one; promazine; 1, 8-Dihydroxy-4,5-dinitroanthraquinone; phenoxazine; anthraquinone; 2-hydroxy-1,4-naphthoquinone; phenothiazine; anthrone; anthracene, anthrarufin, anthrarobin; dimethoxyphenol (DMP); ferulic acid; catechin; epicatechin; homovanillic acid (HMV); and 2,3-dihydroxybenzoic acid (2,3-DHB); or any combination thereof.

8. The method of claim 3 , wherein said delignification reaction proceeds under conditions of between pH 8 and pH 11.

9. The method of claim 3 , wherein said delignification reaction proceeds at a temperature of at least 50° C.

10. A method of oxidizing a composition comprising a fiber, comprising:
contacting said composition with the polypeptide of claim 1 .

11. The method of claim 10 , further comprising contacting the composition with a mediator.

12. The method of claim 11 , wherein said mediator is a compound represented by Formula I or Formula II.

13. The method of claim 11 , wherein the mediator is

14. The method of claim 11 , wherein the mediator is selected from the group consisting of violuric acid; 2,6,6-tet-rarmethylpiperidein-1-yloxy (TEMPO); 1-hydroxybenzotriazole (HBT); 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS); syringaldazine; N-benzoyl-N-phenyl hydroxylamine (BPHA); N-hydroxyphthalimide, 3-Hydroxy-1,2,3-benzotriazin-4-one; promazine; 1, 8-Dihydroxy-4, 5-dinitroanthraquinone; phenoxazine; anthraquinone; 2-hydroxy-1,4-naphthoquinone; phenothiazine; anthrone; anthracene, anthrarufin, anthrarobin; dimethoxyphenol (DMP); ferulic acid; catechin; epicatechin; homovanillic acid (HMV); and 2,3-dihydroxybenzoic acid (2,3-DHB); or any combination thereof.

15. The method of claim 10 , wherein said oxidation reaction proceeds under conditions of between pH 8 and pH 11.

16. The method of claim 10 wherein said oxidation reaction proceeds at a temperature of at least 50° C.

17. A method of bleaching a composition comprising pulp or paper, comprising:
contacting said composition with the polypeptide of claim 1 .

18. The method of claim 17 , further comprising contacting the composition with a mediator.

19. The method of claim 18 , wherein said mediator is a compound represented by Formula I or Formula II.

20. The method of claim 19 , wherein the mediator is

21. The method of claim 19 , wherein the mediator is selected from the group consisting of violuric acid; 2,6,6-tet-rarmethylpiperidein-1-yloxy (TEMPO); 1-hydroxybenzotriazole (HBT); 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS); syringaldazine; N-benzoyl-N-phenyl hydroxylamine (BPHA); N-hydroxyphthalimide, 3-Hydroxy-1,2,3-benzotriazin-4-one; promazine; 1,8-Dihydroxy-4,5-dinitroanthraquinone; phenoxazine; anthraquinone; 2-hydroxy-1,4-naphthoquinone; phenothiazine; anthrone; anthracene; anthrarufin; anthrarobin; dimethoxyphenol (DMP); ferulic acid; catechin; epicatechin; homovanillic acid (HMV); and 2,3-dihydroxybenzoic acid (2,3-DHB); or any combination thereof.

22. The method of claim 19 , wherein said bleaching reaction proceeds under conditions of between pH 8 and pH 11.

23. The method of claim 19 , wherein said bleaching reaction proceeds at a temperature of at least 50° C.

Assignees

• Basf Enzymes Llc
• Basf Se

Inventors

• Kerovuo Janne Samuli
• Haremza Sylke
• Koch Oliver
• Habicher Tilo
• Robertson Dan E
• Desantis Grace
• Mccann Ryan
• Luginbuhl Peter

Classifications

• C12Y110/03002

  Laccase (1.10.3.2) · CPC title

• C12N9/0061 Primary

  Laccase (1.10.3.2) · CPC title

• C12P19/00

  Preparation of compounds containing saccharide radicals (ketoaldonic acids C12P7/58) · CPC title

• D21C5/005

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

• C12N15/09

  Recombinant DNA-technology · CPC title