Processes for producing fermentation products

Patent metadata

Field	Value
Publication number	US-9816112-B2
Application number	US-201113994310-A
Country	US
Kind code	B2
Filing date	Dec 21, 2011
Priority date	Dec 22, 2010
Publication date	Nov 14, 2017
Grant date	Nov 14, 2017

Abstract

The present invention relates to processes for producing fermentation products from starch-containing material, wherein a thermostable alpha-amylase and optionally a thermostable protease are present and/or added during liquefaction. The invention also relates to a composition suitable for use in a process of the invention.

First claim

The invention claimed is:

1. A process for producing fermentation products from starch-containing material comprising the steps of:
i) liquefying the starch-containing material at a pH in the range from 4.5-5.0 at a temperature in the range from 80-90° C. using:
a Bacillus stearothermophilus alpha-amylase having a T½ (min) at pH 4.5, 85° C., 0.12 mM CaCl 2 ) of at least 10, wherein the alpha-amylase is truncated and has altered amino acids selected from the group consisting of:

I181*+G182*+N193F+V59A+Q89R+E129V+K177L+R179E+H208Y+K220P+N224L+Q254S;
I181*+G182*+N193F+E129V+K177L+R179E; and
I181*+G182*+N193F+E129V+K177L+R179E+K220P+N224L+S242Q+Q254S, using SEQ ID NO: 1 for numbering;
a Pyrococcus protease having at least 95% identity to SEQ ID NO: 13 and a thermostability value of more than 20% determined as Relative Activity at 80° C./70° C.;

ii) saccharifying using a carbohydrate-source generating enzyme;
iii) fermenting using a fermenting organism.

2. A composition comprising an alpha-amylase and a protease, wherein the
i) alpha-amylase is a Bacillus stearothermophilus alpha-amylase having a T½ (min) at pH 4.5, 85° C., 0.12 mM CaCl 2 ) of at least 10; and
ii) the protease is Pyrococcus protease having at least 95% identity to SEQ ID NO: 13 and a thermostability value of more than 20% determined as Relative Activity at 80° C./70° C., wherein the alpha-amylase is truncated and has altered amino acids selected from the group consisting of:
I181*+G182*+N193F+V59A+Q89R+E129V+K177L+R179E+H208Y+K220P+N224L+Q254S;
I181*+G182*+N193F+E129V+K177L+R179E; and
I181*+G182*+N193F+E129V+K177L+R179E+K220P+N224L+S242Q+Q254S, using SEQ ID NO: 1 for numbering.

3. The composition of claim 2 , wherein the alpha-amylase is a variant of a Bacillus stearothermophilus alpha-amylase having at least 95% sequence identity, but less than 100% identity to the polypeptide of SEQ ID NO: 1.

4. The composition of claim 2 , further wherein the composition comprises a second alpha-amylase.

5. The composition of claim 2 , comprising:
an alpha-amylase having a T½ (min) at pH 4.5, 85° C., 0.12 mM CaCl 2 ) of at least 10;
a second alpha-amylase having a T½ (min) at pH 4.5, 85° C., 0.12 mM CaCl 2 ) of less than 10;
a protease having a thermostability value of more than 20% determined as Relative Activity at 80° C./70° C.;
a thermostable glucoamylase.

6. The composition of claim 2 , wherein the protease has a thermostability of more than 30% determined as Relative Activity at 80° C./70° C.

7. A composition comprising an alpha-amylase and a protease, wherein the
alpha amylase is the Bacillus stearothermophilus alpha-amylase shown in SEQ ID NO: 1 having the following mutations: I181*+G182*+N193F+E129V+K177L+R179E or an alpha-amylase which has at least 95% sequence identity to the polypeptide of SEQ ID NO: 1 and having a T½ (min) at pH 4.5, 85° C., 0.12 mM CaCl 2 of at least 10; and wherein the protease is Pyrococcus furiosus protease having at least 95% sequence identity to SEQ ID NO: 13 and has a thermostability value of more than 20% determined as Relative Activity at 80° C./70° C.

8. The composition of claim 7 , wherein the alpha-amylase has at least 99% sequence identity to the polypeptide of SEQ ID NO: 1.

9. The composition of claim 7 , wherein the alpha-amylase has at least 97% sequence identity to the polypeptide of SEQ ID NO: 1.

10. The composition of claim 7 , wherein the Pyrococcus furiosus protease has at least 97% sequence identity to SEQ ID NO: 13.

11. The composition of claim 7 , wherein the Pyrococcus furiosus protease has at least 99% sequence identity to SEQ ID NO: 13.

12. A composition comprising an alpha-amylase and a protease, wherein the
i) alpha-amylase is a variant of the Bacillus stearothermophilus alpha-amylase of SEQ ID NO: 1 having a double deletion in I181+G182 and substitution N193F, wherein the variant has at least 95% identity, but less than 100% identity to the polypeptide of SEQ ID NO: 1 and has a T½ (min) at pH 4.5, 85° C., 0.12 mM CaCl 2 of at least 10; and
ii) Pyrococcus furiosus protease has at least 95% sequence identity to SEQ ID NO: 13 and has a thermostability value of more than 20% determined as Relative Activity at 80° C./70° C.

13. A composition comprising an alpha-amylase and a protease, wherein the
i) alpha-amylase is a variant of the Bacillus stearothermophilus alpha-amylase of SEQ ID NO: 1 having a double deletion in I181+G182, wherein the variant has at least 95% identity, but less than 100% identity to the polypeptide of SEQ ID NO: 1 and has a T½ (min) at pH 4.5, 85° C., 0.12 mM CaCl 2 of at least 10; and
ii) Pyrococcus furiosus protease has at least 90% sequence identity to SEQ ID NO: 13 and has a thermostability value of more than 20% determined as Relative Activity at 80° C./70° C.

Assignees

• Deinhammer Randy
• Clark Suzanne
• Quiros Mauricio
• Matthews John
• Hjulmand Anne Glud
• Soong Chee Leong
• Matsui Tomoko
• Takagi Shinobu
• Novozymes As

Inventors

• Deinhammer Randy
• Clark Suzanne
• Quiros Mauricio
• Matthews John
• Hjulmand Anne Glud
• Soong Chee Leong
• Matsui Tomoko
• Takagi Shinobu

Classifications

• C12Y302/01001

  Alpha-amylase (3.2.1.1) · CPC title

• C12P7/06 Primary

  Ethanol, i.e. non-beverage · CPC title

• C12P19/20

  produced by the action of an exo-1,4 alpha-glucosidase, e.g. dextrose · CPC title

• C12Y302/01003

  Glucan 1,4-alpha-glucosidase (3.2.1.3), i.e. glucoamylase · CPC title

• C12Y304/24

  Metalloendopeptidases (3.4.24) · CPC title