Glucoamylase Variants
AssigneeNovozymes A/S
InventorAllan Svendsen
Life of this patent
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Abstract
The invention relates to a variant of a parent fungal glucoamylase, which exhibits improved thermal stability and/or increased specific activity using saccharide substrates.
Claims25
A variant of a parent glucoamylase comprising one or more mutation(s) in position(s) or region(s) corresponding to the following position(s) or region(s) in the amino acid sequence published as SEQ ID NO:
- 2 in WO 00/04136:
- a) said variant comprising one or more insertion(s) in:
- Region:
- 1-35, Region:
- 40-58, Region:
- 60-62, Region:
- 73-80, Position:
- 93, Region:
- 95-101, Region:
- 103-121, Region:
- 123-124, Region:
- 126-127, Region:
- 170-175, Region:
- 177-184, Region:
- 200-212, Region:
- 234-246, Region:
- 287-312, Region:
- 314-319, Region:
- 334-339, Position:
- 341, Region:
- 354-356, Position:
- 358, Region:
- 360-374, Region:
- 388-392, Position:
- 394, Region:
- 396-401, Region:
- 403-407, Region:
- 409-414, Region:
- 445-449, Region:
- 452-467, Region:
- 469-470, and/or in a corresponding position or region in a homologous glucoamylase which displays at least 60% homology with the amino acid sequences shown in SEQ ID NO:
- 2 of WO 00/04136, or
- b) said variant comprising one or more substitution(s), insertion(s) and/or deletion(s) in:
- Region:
- 36-39, Region:
- 63-65, Position 67, Region:
- 69-71, Region:
- 81-92, Region:
- 128-169, Region:
- 185-188, Region:
- 190-199, Region:
- 213-222, Region:
- 224-226, Region:
- 228-233, Region:
- 247-271, Region:
- 273-286, Region:
- 320-333, Region:
- 343-344, Region:
- 346-347, Region:
- 349-351, Region:
- 375-378, Region:
- 380-385, Position:
- 387, Position:
- 415, Region:
- 417-424, Position:
- 426, Region:
- 428-443, Region:
- 471-485, Region:
- 487-489, Region:
- 491-493, Region:
- 495-616, and/or in a corresponding position or region in a homologous glucoamylase which displays at least 60% homology with the amino acid sequence shown in SEQ ID NO:
- 2 of WO 00/04136, except the following amino acid substitutions:
- A39V, P128S, G137A, G139A, D153N, S185H, G251A, D257E, 259D, E259Q, C320A, D375C, G383A, W417F, S431C, A435P, S436P, W437F, A442T, A471C, A479C, T480C, P481C, A495T and A495P.
The process of any of the preceding claims, wherein the saccharification is conducted in the presence of a debranching enzyme selected from the group of pullulanase and isoamylase, preferably a pullulanase derived from Bacillus acidopullulyticus or Bacillus deramificans or an isoamylase derived from Pseudomonas amyloderamosa.
The process of any of the preceding claims, wherein the saccharification is conducted at a pH of 3 to 5.5, preferably at a pH from 4 to 4.5, and at a temperature of 60-80° C., preferably 63-75° C., for 24 to 72 hours, preferably for 36-48 hours.
A method for altering the thermostability and/or for altering the specific activity of a parent glucoamylase by making one or more mutation(s) in one or more of the following position(s) or region(s) corresponding to the position(s) or region(s) of the amino acid sequence shown in SEQ ID NO:
- 2 of WO 00/04136:
- a) said mutation(s) comprising one or more insertion(s) in:
- Region:
- 1-35, Region:
- 40-58, Region:
- 60-62, Region:
- 73-80, Position:
- 93, Region:
- 95-101, Region:
- 103-121, Region:
- 123-124, Region:
- 126-127, Region:
- 170-175, Region:
- 177-184, Region:
- 200-212, Region:
- 234-246, Region:
- 287-312, Region:
- 314-319, Region:
- 334-339, Position:
- 341, Region:
- 354-356, Position:
- 358, Region:
- 360-374, Region:
- 388-392, Position:
- 94, Region:
- 396-401, Region:
- 403-407, Region:
- 409-414, Region:
- 445-449, Region:
- 452-467, Region:
- 469-470, and/or in a corresponding position or region in a homologous glucoamylase which displays at least 60% homology with the amino acid sequences shown in SEQ ID NO:
- 2 of WO 00/04136, or
- b) said mutation(s) comprising one or more substitution(s), insertion(s) and/or deletion(s) in:
- Region:
- 36-39, Region:
- 63.65, Position 67, Region:
- 69-71, Region:
- 81-92, Region:
- 128-169, Region:
- 85-188, Region:
- 190-199, Region:
- 213-222, Region:
- 224-226, Region:
- 228-233, Region:
- 247-271, Region:
- 273-286, Region:
- 320-333, Region:
- 343-344, Region:
- 346-347, Region:
- 349-351, Region:
- 375-378, Region:
- 380-385, Position:
- 387, Position:
- 415, Region:
- 417-424, Position:
- 426, Region:
- 428-443, Region:
- 471-485, Region:
- 487-489, Region:
- 491-493, Region:
- 495-616, and/or in a corresponding position or region in a homologous glucoamylase which displays at least 60% homology with the amino acid sequence shown in SEQ ID NO:
- 2 of WO 00/04136, except the following amino acid substitutions:
- A39V, P128S, G137A, G139A, D153N, S185H, G251A, D257E, F259D, E259Q, C320A, D375C, G383A, W417F, S431C, A435P, S436P, W437F, A442T, A471C, A479C, T480C, P481C, A495T and A495P.
Dependencies are inferred from the claim text.