Method for producing L-amino acid of glutamate family

Patent metadata

Field	Value
Publication number	US-9932614-B2
Application number	US-201615334444-A
Country	US
Kind code	B2
Filing date	Oct 26, 2016
Priority date	Oct 30, 2015
Publication date	Apr 3, 2018
Grant date	Apr 3, 2018

Abstract

A method for producing an L-amino acid of glutamate family such as L-glutamic acid is provided. An L-amino acid of glutamate family is produced by culturing a coryneform bacterium having an ability for producing an L-amino acid of glutamate family, which has been modified so that the activity of an α-ketoglutaric acid (α-KG) uptake carrier is increased, in a medium, and collecting the L-amino acid of glutamate family from the medium.

First claim

The invention claimed is:

1. A method for producing an L-amino acid, the method comprising:
A) culturing a coryneform bacterium having an L-amino acid-producing ability in a medium; and
B) collecting the L-amino acid from the medium, wherein the bacterium has been modified so that the activity of an α-ketoglutaric acid (α-KG) uptake carrier is increased as compared with a non-modified bacterium, wherein the α-KG uptake carrier is a protein encoded by kgtP gene, and
wherein the L-amino acid is an L-amino acid of glutamate family.

2. The method according to claim 1 , wherein the α-KG uptake carrier is a protein selected from the group consisting of:
(a) a protein comprising the amino acid sequence of SEQ ID NO: 8;
(b) a protein comprising the amino acid sequence of SEQ ID NO: 8, but which includes substitution, deletion, insertion, and/or addition of 1 to 10 amino acid residues, and wherein said protein has α-KG uptake activity; and
(c) a protein comprising an amino acid sequence having an identity of 90% or higher to the amino acid sequence of SEQ ID NO: 8, and wherein said protein has α-KG uptake activity.

3. The method according to claim 1 , wherein the activity of the α-KG uptake carrier is increased by increasing the expression of a gene encoding the α-KG uptake carrier.

4. The method according to claim 3 , wherein the expression of the gene is increased by increasing the copy number of the gene and/or modifying an expression control sequence of the gene.

5. The method according to claim 1 , wherein the bacterium has further been modified so that the activity of phosphoketolase is increased as compared with a non-modified bacterium.

6. The method according to claim 5 , wherein the phosphoketolase comprises D-xylulose-5-phosphate phosphoketolase and/or fructose-6-phosphate phosphoketolase.

7. The method according to claim 5 , wherein the activity of the phosphoketolase is increased by increasing the expression of a gene encoding the phosphoketolase.

8. The method according to claim 1 , wherein the bacterium has further been modified so that the activity of α-ketoglutarate dehydrogenase and/or succinate dehydrogenase is reduced as compared with a non-modified bacterium.

9. The method according to claim 1 , wherein the bacterium is a Corynebacterium bacterium.

10. The method according to claim 9 , wherein the bacterium is Corynebacterium glutamicum.

11. The method according to claim 1 , wherein the L-amino acid of glutamate family is selected from the group consisting of L-glutamic acid, L-glutamine, L-proline, L-arginine, L-citrulline, L-ornithine, and combinations thereof.

12. The method according to claim 1 , wherein the L-amino acid of glutamate family is L-glutamic acid.

13. The method according to claim 11 , wherein the L-glutamic acid is monoammonium L-glutamate or monosodium L-glutamate.

14. The method according to claim 12 , wherein the bacterium has been further modified to harbor a mutant yggB gene.

15. The method according to claim 14 , wherein the mutant yggB gene is a yggB gene having a mutation that improves the L-glutamic acid-producing ability of the coryneform bacterium.

16. The method according to claim 14 , wherein the mutant yggB gene encodes a mutant YggB protein, and wherein said mutant yggB gene has a mutation selected from the group consisting of:
(1) a mutation in the region coding for the amino acid residues at positions 419 to 533 of a wild-type YggB protein of SEQ ID NO: 12,
(2) a mutation in the region coding for a transmembrane region of a wild-type YggB protein, and
(3) a combination thereof.

17. The method according to claim 16 , wherein the mutant YggB protein is a protein selected from the group consisting of:
(a) a protein comprising the amino acid sequence of SEQ ID NO: 12, but which includes substitution, deletion, insertion, and/or addition of 1 to 10 amino acid residues, and wherein said protein is overexpressed in the coryneform bacterium, which provides an improved L-glutamic acid-producing ability of the coryneform bacterium; and
(b) a protein comprising an amino acid sequence having an identity of 90% or higher to the amino acid sequence of SEQ ID NO: 12, and wherein said protein is overexpressed in the coryneform bacterium, which provides an improved L-glutamic acid-producing ability of the coryneform bacterium.

Assignees

• Ajinomoto Kk

Inventors

• Hirano Seiko
• Hayashi Kazuyuki
• Takayashiki Hitoshi
• Fukui Keita

Classifications

• C12N9/001

  acting on the CH-CH group of donors (1.3) · CPC title

• C07K14/34

  from Corynebacterium (G) · CPC title

• C12Y401/02009

  Phosphoketolase (4.1.2.9) · CPC title

• C12N9/1022

  transferring aldehyde or ketonic groups (2.2) · CPC title

• C12Y202/01001

  Transketolase (2.2.1.1) · CPC title