Method of producing succinic acid and other chemicals using facilitated diffusion for sugar import

Patent metadata

Field	Value
Publication number	US-10934565-B2
Application number	US-201916588183-A
Country	US
Kind code	B2
Filing date	Sep 30, 2019
Priority date	Jul 23, 2013
Publication date	Mar 2, 2021
Grant date	Mar 2, 2021

Abstract

This invention relates to the production of succinic acid and other chemicals derived from phosphoenolpyruvate (PEP) by fermentation with a microorganism in which the fermentation medium contains one or more sugars, and in which one or more of the sugars is imported into the cell by facilitated diffusion. As a specific example, succinic acid is produced from a glucose-containing renewable feedstock through fermentation using a biocatalyst. Examples of such a biocatalyst include microorganisms that have been enhanced in their ability to utilize glucose as a carbon and energy source. The biocatalysts of the present invention are derived from the genetic manipulation of parental strains that were originally constructed with the goal to produce one or more chemicals (for example succinic acid and/or a salt of succinic acid) at a commercial scale using feedstocks that include, for example, glucose, fructose, or sucrose. The genetic manipulations of the present invention involve the introduction of exogenous genes involved in the transport and metabolism of glucose or fructose into the parental strains. The genes involved in the transport and metabolism of glucose or fructose can also be introduced into a microorganism prior to developing the organism to produce a particular chemical. The genes involved in the transport and metabolism of sucrose can also be used to augment or improve the efficiency of sugar transport and metabolism by strains already known to have some ability for glucose utilization in biological fermentations.

First claim

What is claimed is:

1. An Escherichia coli bacterium, which produces more than 30 g/L of a succinic acid in 48 hours when grown in a minimal medium, wherein a biosynthetic intermediate for said succinic acid is phosphoenolpyruvate, and said Escherichia coli bacterium comprises:
at least one exogenous gene that encodes a protein that functions in the facilitated diffusion of a sugar,
a mutation or deletion in one or more genes that encode one or more proteins that function in a phosphotransferase system for sugar import, and
at least one additional copy of a gene that encodes a Crr protein that functions in catabolite repression.

2. The Escherichia coli bacterium of claim 1 , further comprising a deletion in a gene that encodes a sugar importer that functions using proton symport.

3. The Escherichia coli bacterium of claim 1 , wherein said one or more genes comprise a ptsH gene.

4. The Escherichia coli bacterium of claim 1 , wherein said one or more genes comprise a ptsI gene.

5. The Escherichia coli bacterium of claim 1 , wherein said one or more genes comprise a gene selected from the group consisting of a ptsH gene, and a ptsI gene.

6. The Escherichia coli bacterium of claim 1 , wherein said at least one exogenous gene is a glf gene.

7. The Escherichia coli bacterium of claim 1 , wherein said at least one exogenous gene is a glf gene and a glk gene.

8. The Escherichia coli bacterium of claim 1 , wherein said at least one exogenous gene is a glf gene and a frk gene.

9. The Escherichia coli bacterium of claim 2 , wherein said gene is a galP gene.

10. A method of producing succinic acid, the method comprising:
growing the Escherichia coli bacterium of claim 1 in a minimal medium, to produce succinic acid; and
optionally purifying said succinic acid from the minimal medium.

11. The method of claim 10 , wherein the Escherichia coli bacterium produces, in 48 hours, at least 60 g/L of succinic acid and 4.2 g/L or less of acetate.

12. The method of claim 10 , wherein the growing is microaerobic.

13. The Escherichia coli bacterium of claim 1 , wherein said at least one additional copy is integrated at a locus separate from a native crr locus.

Assignees

• Ptt Global Chemical Public Co Ltd

Inventors

• Yocum R Rogers
• Collard Andrew Christopher
• Hermann Theron
• Yu Xiaohui
• Gong Wei

Classifications

• Y02E50/10

  Biofuels, e.g. bio-diesel · CPC title

• C12P7/46 Primary

  Dicarboxylic acids having four or less carbon atoms, e.g. fumaric acid, maleic acid · CPC title

• C07K14/195

  from bacteria · CPC title

• C12N15/01

  Preparation of mutants without inserting foreign genetic material therein; Screening processes therefor · CPC title

• C12P7/44

  Polycarboxylic acids · CPC title