Fungal xylanases and xylosidases

Abstract

The present invention provides fungal xylanase and/or xylosidase enzymes suitable for use in saccharification reactions. The present invention provides xylanase and xylosidase enzymes suitable for use in saccharification reactions. The present application further provides genetically modified fungal organisms that produce xylanase(s) and/or xylosidase(s), as well as enzyme mixtures exhibiting enhanced hydrolysis of cellulosic material to fermentable sugars, enzyme mixtures produced by the genetically modified fungal organisms, and methods for producing fermentable sugars from cellulose using such enzyme mixtures. In some embodiments, the xylanase and xylosidase enzyme(s) are M. thermophila enzymes.

We claim: 
     
         1 . A recombinant organism comprising a xylanase and/or xylosidase and/or biologically active xylanase and/or xylosidase fragment comprising (a) an amino acid sequence comprising at least about 70%, at least about 75%, at least about 80%, at least about 81%, at least 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% sequence identity to SEQ ID NO:8. 
     
     
         2 . The recombinant organism of  claim 1 , wherein said isolated xylanase and/or xylosidase or biologically active xylanase and/or xylosidase fragment is a  Myceliophthora thermophila  xylanase, xylosidase and/or fragment. 
     
     
         3 . A recombinant organism comprising at least one polynucleotide comprising at least one nucleic acid sequence encoding the xylanase and/or xylosidase of  claim 1 , wherein said polynucleotide comprises a sequence that has least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7. 
     
     
         4 . An isolated xylanase and/or xylosidase and/or biologically active xylanase and/or xylosidase fragment comprising (a) an amino acid sequence comprising at least about 70%, at least about 75%, at least about 80%, at least about 81%, at least 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% sequence identity to SEQ ID NO:8. 
     
     
         5 . An enzyme composition comprising the xylanase and/or xylosidase of  claim 4 . 
     
     
         6 . The enzyme composition of  claim 5 , further comprising at least one additional enzyme, selected from cellulases, hemicellulases, xylanases, amylases, glucoamylases, proteases, esterases, and lipases. 
     
     
         7 . A recombinant nucleic acid construct comprising at least one polynucleotide sequence, wherein the polynucleotide is selected from:
 (a) a polynucleotide that encodes a polypeptide comprising an amino acid sequence comprising at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:8;   (b) a polynucleotide that hybridizes under stringent hybridization conditions to at least a fragment of a polynucleotide that encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:8; and/or   (c) a polynucleotide that hybridizes under stringent hybridization conditions to the complement of at least a fragment of a polynucleotide that encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:8.   
     
     
         8 . The nucleic acid construct of  claim 7 , wherein the polynucleotide sequence is operably linked to a promoter. 
     
     
         9 . The nucleic acid construct of  claim 7 , wherein said nucleic acid sequence is operably linked to at least one additional regulatory sequence. 
     
     
         10 . A recombinant host cell that expresses at least one polynucleotide sequence encoding at least one xylanase and/or xylosidase of  claim 4 . 
     
     
         11 . A recombinant host cell comprising at least one nucleic acid construct as provided in  claim 7 . 
     
     
         12 . The recombinant host cell of  claim 11 , wherein said host cell comprises the polynucleotide sequence set forth in SEQ ID NO:7. 
     
     
         13 . The recombinant host cell of  claim 11 , wherein at least one xylanase and/or xylosidase is produced by said cell. 
     
     
         14 . The recombinant host cell of  claim 11 , wherein said host cell further produces at least one enzyme selected from endoglucanases (EG), β-glucosidases (BGL), Type 1 cellobiohydrolases (CBH1), Type 2 cellobiohydrolases (CBH2), and GH61 enzymes. 
     
     
         15 . The recombinant cell of  claim 11 , wherein said cell is a prokaryotic or eukaryotic cell. 
     
     
         16 . A method for producing at least one fermentable sugar from a feedstock, comprising contacting the feedstock with the enzyme composition of  claim 5 , under culture conditions whereby fermentable sugars are produced. 
     
     
         17 . The method of  claim 16 , wherein the enzyme composition further comprises at least one enzyme selected from endoglucanases (EG), β-glucosidases (BGL), Type 1 cellobiohydrolases (CBH1), Type 2 cellobiohydrolases (CBH2), and GH61 enzymes. 
     
     
         18 . A method of producing an end product from a feedstock, comprising:
 a) acting the feedstock with at least one enzyme composition of  claim 5 , under conditions whereby at least one fermentable sugar is produced from the substrate; and   b) contacting the fermentable sugar with a microorganism under conditions such that the microorganism uses the fermentable sugar to produce an end-product.   
     
     
         19 . A method for producing at least one fermentable sugar from a feedstock, comprising contacting the feedstock with the recombinant host cell of  claim 11 , under culture conditions whereby fermentable sugars are produced. 
     
     
         20 . The method of  claim 19 , wherein the recombinant host cell further comprises at least one enzyme selected from endoglucanases (EG), β-glucosidases (BGL), Type 1 cellobiohydrolases (CBH1), Type 2 cellobiohydrolases (CBH2), and/or GH61 enzymes.