Compositions for saccharification of cellulosic material
The present invention relates to enzyme compositions for high temperature saccharification of cellulosic material and to uses thereof.
1. An enzyme composition, comprising:
(I) a polypeptide having cellobiohydrolase I activity selected from the group consisting of: (a) a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 158; (b) a polypeptide encoded by a polynucleotide that hybridizes under at least high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 157, (ii) the genomic DNA sequence of the mature polypeptide coding sequence of SEQ ID NO: 157, or (iii) the full-length complement of (i) or (ii), 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 15 minutes using 2×SSC, 0.2% SDS at 65° C.; and (c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence having at least 95% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 157;
(II) a polypeptide having cellobiohydrolase II activity selected from the group consisting of: (a) a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 18; (b) a polypeptide encoded by a polynucleotide that hybridizes under at least high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 17, (ii) the cDNA sequence of the mature polypeptide coding sequence of SEQ ID NO: 17, or (iii) the full-length complement of (i) or (ii), 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 15 minutes using 2×SSC, 0.2% SDS at 65° C.; and (c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence having at least 95% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 17;
(III) a polypeptide having endoglucanase I activity selected from the group consisting of: (a) a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 20; (b) a polypeptide encoded by a polynucleotide that hybridizes under at least high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 19, (ii) the cDNA sequence of the mature polypeptide coding sequence of SEQ ID NO: 19, or (iii) the full-length complement of (i) or (ii), 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 15 minutes using 2×SSC, 0.2% SDS at 65° C.; and (c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence having to the mature polypeptide coding sequence of SEQ ID NO: 19;
(IV) a polypeptide having endoglucanase II activity selected from the group consisting of: (a) a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 26; (b) a polypeptide encoded by a polynucleotide that hybridizes under at least high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 25, (ii) the genomic DNA sequence of the mature polypeptide coding sequence of SEQ ID NO: 25, or (iii) the full-length complement of (i) or (ii), 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 15 minutes using 2×SSC, 0.2% SDS at 65° C.; and (c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence having at least 95% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 25; and
(V) a polypeptide having beta-glucosidase activity selected from the group consisting of: (a) a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 178; (b) a polypeptide encoded by a polynucleotide that hybridizes under at least high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 177, (ii) the cDNA sequence of the mature polypeptide coding sequence of SEQ ID NO: 177, or (iii) the full-length complement of (i) or (ii), 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 15 minutes using 2×SSC, 0.2% SDS at 65° C.; and (c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence having at least 95% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 177;
wherein saccharification of a cellulosic material is performed with the enzyme composition at a temperature in the range of about 40° C. to about 70° C.
2. The enzyme composition of claim 1 , further comprising a polypeptide having cellulolytic enhancing activity selected from the group consisting of: (a) a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 42; (b) a polypeptide encoded by a polynucleotide that hybridizes under at least high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 41, (ii) the cDNA sequence of the mature polypeptide coding sequence of SEQ ID NO: 41, or (iii) the full-length complement of (i) or (ii), 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 15 minutes using 2×SSC, 0.2% SDS at 65° C.; and (c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence having at least 95% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 41.
3. The enzyme composition of claim 1 , further comprising a polypeptide having xylanase activity.
4. The enzyme composition of claim 3 , wherein the polypeptide having xylanase activity is a Family 10 polypeptide having xylanase activity.
5. The enzyme composition of claim 4 , wherein the Family 10 polypeptide having xylanase activity is selected from the group consisting of: (a) a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 50; (b) a polypeptide encoded by a polynucleotide that hybridizes under at least high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 49, (ii) the genomic DNA sequence of the mature polypeptide coding sequence of SEQ ID NO: 49, or (iii) the full-length complement of (i) or (ii), 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 15 minutes using 2×SSC, 0.2% SDS at 65° C.; and (c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence having at least 95% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 49.
6. The enzyme composition of claim 3 , wherein the polypeptide having xylanase activity is a Family 11 polypeptide having xylanase activity.
7. The enzyme composition of claim 6 , wherein the Family 11 polypeptide having xylanase activity is selected from the group consisting of: (a) a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 56; (b) a polypeptide encoded by a polynucleotide that hybridizes under at least high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 55 or (ii) the full-length complement of (i) or (ii), 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 15 minutes using 2×SSC, 0.2% SDS at 65° C.; and (c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence having at least 95% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 55.
8. The enzyme composition of claim 1 , further comprising a beta-xylosidase.
9. The enzyme composition of claim 8 , wherein the beta-xylosidase is selected from the group consisting of: (a) a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the mature polypeptide of SEQ ID NO: 60; (b) a polypeptide encoded by a polynucleotide that hybridizes under at least high stringency conditions with the mature polypeptide coding sequence of SEQ ID NO: 59 or its full-length complement, 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 15 minutes using 2×SSC, 0.2% SDS at 65° C.; and (c) a polypeptide encoded by a polynucleotide comprising a nucleotide sequence having at least 95% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 59.
10. The enzyme composition of claim 1 , further comprising one or more enzymes selected from the group consisting of a hemicellulase, a carbohydrate-esterase, a pectinase, a protease, a laccase, a peroxidase, an H 2 O 2 -producing enzyme, an oxidoreductase, an expansin, a swollenin, or a mixture thereof.
11. The enzyme composition of claim 1 , wherein the saccharification of the cellulosic material is performed at a temperature in the range of about 50° C. to about 65° C.
12. The enzyme composition of claim 1 , which reduces the amount of protein required to achieve the same degree of cellulose hydrolysis of pretreated corn stover at 55° C., pH 3.5 to pH 6.0 at least 1.1-fold compared to a Trichoderma reesei -based enzyme composition SaMe-MF268 at 50° C., pH 5.0.
13. The enzyme composition of claim 1 , which reduces the amount of protein required to achieve the same degree of cellulose hydrolysis of pretreated corn stover at 60° C., pH 3.5 to pH 6.0 at least 1.1-fold compared to a Trichoderma reesei -based enzyme composition SaMe-MF268 at 50° C., pH 5.0.
14. The enzyme composition of claim 1 , which reduces the amount of protein required to achieve the same degree of cellulose hydrolysis of pretreated corn stover at 65° C., pH 3.5 to pH 6.0 at least 1.1-fold compared to a Trichoderma reesei -based enzyme composition SaMe-MF268 at 50° C., pH 5.0.
15. The enzyme composition of claim 1 , which reduces the amount of protein required to achieve the same degree of cellulose hydrolysis of pretreated corn stover at 70° C., pH 3.5 to pH 6.0 at least 1.1-fold compared to a Trichoderma reesei -based enzyme composition SaMe-MF268 at 50° C., pH 5.0.
16. An isolated recombinant host cell encoding the enzyme composition of claim 1 .
17. A method of producing an enzyme composition, comprising: (a) cultivating the recombinant host cell of claim 16 under conditions conducive for production of the enzyme composition; and (b) recovering the enzyme composition.
18. A method for degrading or converting a cellulosic material, comprising: treating the cellulosic material with the enzyme composition of claim 1 .
19. A method for producing a fermentation product, comprising:
(a) saccharifying a cellulosic material with the enzyme composition 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.
20. 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 the enzyme composition of claim 1 .
21. The enzyme composition of claim 11 , wherein the saccharification of the cellulosic material with the enzyme composition is performed at a temperature in the range of about 55° C. to about 63° C.
22. The enzyme composition of claim 21 , wherein the saccharification of the cellulosic material with the enzyme composition is performed at a temperature in the range of about 55° C. to about 63° C.
23. The enzyme composition of claim 22 , wherein the saccharification of the cellulosic material with the enzyme composition is performed at a temperature in the range of about 55° C. to about 60° C.