Polypeptides having cellulolytic enhancing activity and polynucleotides encoding same

Patent metadata

Field	Value
Publication number	US-9409958-B2
Application number	US-201214004141-A
Country	US
Kind code	B2
Filing date	Mar 9, 2012
Priority date	Mar 10, 2011
Publication date	Aug 9, 2016
Grant date	Aug 9, 2016

Abstract

The present invention relates to isolated polypeptides having cellulolytic enhancing activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

First claim

What is claimed is:

1. A nucleic acid construct comprising a polynucleotide encoding a GH61 polypeptide having cellulolytic enhancing activity, wherein the polynucleotide is operably linked to one or more heterologous control sequences that direct the production of the polypeptide, and wherein the GH61 polypeptide having cellulolytic enhancing activity is selected from the group consisting of:
(a) a GH61 polypeptide having at least 90% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2;
(b) a GH61 polypeptide encoded by a polynucleotide that hybridizes under high stringency conditions with (i) the full-length complement of nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (ii) the cDNA of (i), wherein high stringency conditions are defined as prehybridization and hybridization at 42° C. in 5X SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 50% formamide, and washing three times each for 15 minutes using 2× SSC, 0.2% SDS at 65° C.;
(c) a GH61 polypeptide encoded by (i) a polynucleotide having at least 90% sequence identity to nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (ii) the cDNA of (i); and
(d) a fragment of the GH61 polypeptide of (a), (b), or (c), wherein the fragment has cellulolytic enhancing activity.

2. An isolated recombinant host cell transformed with the nucleic acid construct of claim 1 .

3. A method of producing a GH61 polypeptide having cellulolytic enhancing activity, comprising cultivating the recombinant host cell of claim 2 under conditions conducive for production of the polypeptide.

4. An isolated plant cell transformed with the nucleic acid construct of claim 1 .

5. A method of producing a GH61 polypeptide having cellulolytic enhancing activity, comprising cultivating the plant cell of claim 4 under conditions conducive for production of the polypeptide.

6. A nucleic acid construct encoding a protein and a signal peptide operably linked to the protein, wherein the signal peptide consists of amino acids 1 to 20 of SEQ ID NO: 2, wherein the signal peptide is foreign to the protein, and wherein the C-terminus of the signal peptide is linked to the N-terminus of the protein.

7. A method of producing a protein, comprising cultivating a recombinant host cell comprising the nucleic acid construct of claim 6 , under conditions conducive for production of the protein.

8. The nucleic acid construct of claim 4 , wherein the GH61 polypeptide having cellulolytic enhancing activity has at least 95% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2.

9. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity has at least 96% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2.

10. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity has at least 97% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2.

11. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity has at least 98% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2.

12. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity comprises (i) the polypeptide of SEQ ID NO: 2; (ii) amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2; or (iii) a fragment of (i) or (ii) having cellulolytic enhancing activity.

13. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity is encoded by a polynucleotide that hybridizes under high stringency conditions with (i) the full-length complement of nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (ii) the cDNA of (i), wherein high stringency conditions are defined as prehybridization and hybridization at 42° C. in 5×SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 50% formamide, and washing three times each for 15minutes using 2× SSC, 0.2% SDS at 65° C.

14. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity is encoded by a polynucleotide that hybridizes under very high stringency conditions with (i) the full-length complement of nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (ii) the cDNA of (i), wherein very high stringency conditions are defined as prehybridization and hybridization at 42° C. in 5×SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 50% formamide, and washing three times each for 15minutes using 2× SSC, 0.2% SDS at 70° C.

15. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity is encoded by (i) a polynucleotide having at least 95% sequence identity to nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (ii) the cDNA of (i).

16. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity is encoded by (i) a polynucleotide having at least 96% sequence identity to nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (ii) the cDNA of (i).

17. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity is encoded by (i) a polynucleotide having at least 97% sequence identity to nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (ii) the cDNA of (i).

18. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity is encoded by (i) a polynucleotide having at least 98% sequence identity to nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (ii) the cDNA of (i).

19. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity is encoded by (i) a polynucleotide comprising SEQ ID NO: 1, (ii) a polynucleotide comprising nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (iii) the cDNA of (i) or (ii).

20. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity has at least 99% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2.

21. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity is encoded by (i) a polynucleotide having at least 99% sequence identity to nucleotides 61 to 969 of the polynucleotide of SEQ ID NO: 1, or (ii) the cDNA of (i).

22. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity has at least 90% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2.

23. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity has at least 91% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2.

24. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity has at least 92% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2.

25. The nucleic acid construct of claim 1 , wherein the GH61 polypeptide having cellulolytic enhancing activity has at least 93% sequence identity to amino acids 21 to 322 of the polypeptide of SEQ ID NO: 2.

26. The nucleic acid

Assignees

• Schnorr Kirk
• Kramer Randall
• Novozymes Inc

Inventors

• Schnorr Kirk
• Kramer Randall

Classifications

• C12N9/0071 Primary

  acting on paired donors with incorporation of molecular oxygen (1.14) · CPC title

• C07K14/37 Primary

  from fungi · CPC title

• C07K2319/02

  containing a signal sequence · CPC title

• C12N9/2434

  acting on beta-1,4-glucosidic bonds · CPC title

• Y02P20/52

  using catalysts, e.g. selective catalysts · CPC title