GH61 Polypeptide Variants and Polynucleotides Encoding Same

Abstract

The present invention relates to GH61 polypeptide variants. The present invention also relates to polynucleotides encoding the variants; nucleic acid constructs, vectors, and host cells comprising the polynucleotides; and methods of using the variants.

1 - 21 . (canceled) 
     
     
         22 . A variant GH61 polypeptide having cellulolytic enhancing activity, comprising a substitution at one or more positions corresponding to positions 105, 154, 188, 189, 216, and 229 of the mature polypeptide of SEQ ID NO: 30, wherein the substitution at corresponding position 105 is with Pro or Lys, the substitution at corresponding position 154 is with Ile or Leu, the substitution at corresponding position 188 is with Ala, Phe, Met, or Trp, the substitution at corresponding position 189 is with His or Lys, the substitution at corresponding position 216 is with Leu or Tyr, and the substitution at corresponding position 229 is with Trp, His, Ile, or Tyr, wherein the variant has cellulolytic enhancing activity, wherein the variant has increased thermostability relative to a GH61 polypeptide without the substitution at the one or more positions corresponding to positions 105, 154, 188, 189, 216, and 229 of the mature polypeptide of SEQ ID NO: 30, and wherein the variant has at least 95% sequence identity, but less than 100% sequence identity, to the mature polypeptide of SEQ ID NO: 2, 4, 6, 8, 10, 12, 16, 18, 20, 22, 24, 26, 28, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, or 216. 
     
     
         23 . The variant of  claim 22 , wherein the variant has at least 96% sequence identity, but less than 100% sequence identity, to the mature polypeptide of the GH61 polypeptide of SEQ ID NO: 2, 4, 6, 8, 10, 12, 16, 18, 20, 22, 24, 26, 28, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, or 216. 
     
     
         24 . The variant of  claim 22 , wherein the variant has at least 97% sequence identity, but less than 100% sequence identity, to the mature polypeptide of the GH61 polypeptide of SEQ ID NO: 2, 4, 6, 8, 10, 12, 16, 18, 20, 22, 24, 26, 28, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, or 216. 
     
     
         25 . The variant of  claim 22 , wherein the variant has at least 98% sequence identity, but less than 100% sequence identity, to the mature polypeptide of the GH61 polypeptide of SEQ ID NO: 2, 4, 6, 8, 10, 12, 16, 18, 20, 22, 24, 26, 28, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, or 216. 
     
     
         26 . The variant of  claim 22 , wherein the variant has at least 99% sequence identity, but less than 100% sequence identity, to the mature polypeptide of the GH61 polypeptide of SEQ ID NO: 2, 4, 6, 8, 10, 12, 16, 18, 20, 22, 24, 26, 28, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, or 216. 
     
     
         27 . The variant of  claim 22 , which further comprises a substitution at one or more positions corresponding to positions 111, 152, 155, and 162 of the mature polypeptide of SEQ ID NO: 30, wherein the substitution at corresponding position 111 is with Val, the substitution at corresponding position 152 is with Ser, the substitution at corresponding position 155 is with Leu, and the substitution at corresponding position 162 is with Trp, and wherein the variant has cellulolytic enhancing activity. 
     
     
         28 . The variant of  claim 27 , wherein said variant comprises one or more substitutions or corresponding substitutions selected from the group consisting of a substitution of Leu at position 111 with Val, a substitution of Asp at position 152 with Ser, a substitution of Met at position 155 with Leu, and a substitution of Ala at position 162 with Trp. 
     
     
         29 . The variant of  claim 22 , wherein the thermostability of the variant is increased at least 1.01-fold compared to the parent. 
     
     
         30 . An isolated polynucleotide encoding the GH61 polypeptide variant of  claim 22 . 
     
     
         31 . A recombinant host cell transformed with the polynucleotide of  claim 30 . 
     
     
         32 . A method of producing a GH61 polypeptide variant, comprising: (a) cultivating the recombinant host cell of  claim 31  under conditions suitable for expression of the variant; and optionally (b) recovering the variant. 
     
     
         33 . A transgenic plant, plant part or plant cell transformed with the polynucleotide of  claim 30 . 
     
     
         34 . A method of producing a GH61 polypeptide variant, comprising: (a) cultivating the transgenic plant or a plant cell of  claim 33  under conditions conducive for production of the variant; and optionally (b) recovering the variant. 
     
     
         35 . A process for degrading or converting a cellulosic material, comprising: treating the cellulosic material with an enzyme composition comprising the GH61 polypeptide variant having cellulolytic enhancing activity of  claim 22 . 
     
     
         36 . A process for producing a fermentation product, comprising: (a) saccharifying a cellulosic material with an enzyme composition comprising the GH61 polypeptide variant having cellulolytic enhancing activity of  claim 22 ; (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. 
     
     
         37 . A process of fermenting a cellulosic material, comprising: fermenting the cellulosic material with one or more fermenting microorganisms, wherein the cellulosic material is saccharified with an enzyme composition comprising the GH61 polypeptide variant having cellulolytic enhancing activity of  claim 22 . 
     
     
         38 . A whole broth formulation or cell culture composition, comprising the GH61 polypeptide variant of  claim 22 . 
     
     
         39 . A detergent composition, comprising a surfactant and the GH61 polypeptide variant of  claim 22 .