Novel Gene and Method for Producing L-Amino Acids

Patent metadata

Field	Value
Publication number	US-2016289716-A1
Application number	US-201615185371-A
Country	US
Kind code	A1
Filing date	Jun 17, 2016
Priority date	Dec 23, 1998
Publication date	Oct 6, 2016
Grant date	—

Abstract

The present invention describes a bacterium which has an ability to produce an amino acid and in which the rhtC gene encoding a protein having an enhanced activity of imparting L-threonine resistance to a bacterium expressing the protein. Preferably, the bacterium further includes an rhtB gene encoding for a protein having an enhanced activity of imparting to a bacterium L-homoserine resistance expressing the protein. The present invention also describes a method of cultivating the bacterium in a culture medium to produce and accumulate amino acids in the medium, and the amino acid is recovered from the medium.

First claim

What is claimed is:

1 . A method for producing the amino acid L-hormoserine or L-threonine, comprising the steps of:
i) cultivating a bacterium belonging to the genus Escherichia , which has the ability to produce the amino acid, in a culture medium, to produce and accumulate the amino acid in the medium, and ii) recovering the amino acid from the medium, wherein L-threonine resistance of said bacterium is enhanced by increasing the activity of a protein in said bacterium, wherein the protein is selected from the group consisting of: (A) a protein comprising the amino acid sequence shown in SEQ ID NO: 4; and (B) a protein comprising the amino acid sequence shown in SEQ ID NO: 4, but which includes deletion, substitution, insertion or addition of one or several amino acids in the amino acid sequence, and which has the activity of conferring resistance to L-threonine to the bacterium, wherein the protein is encoded by a DNA selected from the group consisting of: (a) a DNA comprising the nucleotide sequence of nucleotide numbers 187 to 804 in SEQ ID NO: 3; (b) a DNA which hybridizes with a nucleotide sequence of nucleotide numbers 187 to 804 in SEQ ID NO: 3 under stringent conditions, and which encodes a protein having the activity of conferring resistance to L-threonine to the bacterium, wherein the stringent conditions comprise washing at 60° C. and at a salt concentration corresponding to 1×SSC and 0.1% SDS.

2 . The method according to claim 1 , wherein the L-homoserine resistance of said bacterium is enhanced by increasing the activity of a protein in the bacterium, wherein the protein is selected from the group consisting of:
(C) a protein comprising the amino acid sequence shown in SEQ ID NO: 2; and (D) a protein comprising the amino acid sequence of SEQ ID NO: 2, but which includes deletion, substitution, insertion or addition of one or several amino acids in the amino acid sequence shown in SEQ ID NO: 2, and which has the activity of making a bacterium having the protein L-homoserine-resistant.

3 . The method according to claim 2 , wherein the protein as defined in (C) or (D) is encoded by a DNA selected from the group consisting of:
(c) a DNA comprising the nucleotide sequence of nucleotide numbers 557 to 1171 in SEQ ID NO: 1; (d) a DNA which hybridizes with a nucleotide sequence of nucleotide numbers 557 to 1171 in SEQ ID NO: 1 under stringent conditions and which encodes a protein having the activity of conferring resistance to L-homoserine to the bacterium, wherein the stringent conditions comprise washing at 60° C., and at a salt concentration corresponding to 1×SSC and 0.1% SDS.

4 . A method for producing an amino acid, comprising the steps of:
i) cultivating a bacterium belonging to the genus Escherichia , which has the ability to produce the amino acid, in a culture medium, to produce and accumulate the amino add in the medium, and ii) recovering the amino acid from the medium, wherein L-threonine resistance of said bacterium is enhanced by increasing the activity of a protein in the bacterium, wherein the protein is selected from the group consisting of: (A) a protein comprising the amino acid sequence shown in SEQ ID NO: 4; and (B) a protein which comprises the amino acid sequence including deletion, substitution, insertion or addition of one or several amino acids in the amino acid sequence shown in SEQ ID NO: 4, and which has the activity of conferring resistance to L-threonine to bacterium, wherein the protein is encoded by a DNA selected from the group consisting of: (a) a DNA comprising the nucleotide sequence of nucleotide numbers 187 to 804 in SEQ ID NO: 3; (b) a DNA which hybridizes with a nucleotide sequence of nucleotide numbers 187 to 804 in SEQ ID NO: 3 under stringent conditions and which encodes a protein having the activity of conferring resistance to L-threonine to the bacterium, wherein the stringent conditions comprise washing at 60° C. and at a salt concentration corresponding to 1×SSC and 0.1% SDS, and wherein the L-homoserine resistance of said bacterium is enhanced by increasing the activity of a protein in the bacterium, wherein said protein is selected from the group consisting of: (C) a protein comprising the amino acid sequence shown in SEQ ID NO: 2; and (D) a protein comprising the amino acid sequence of SEQ ID NO: 2, but which includes deletion, substitution, insertion or addition of one or several amino acids in the amino acid sequence shown in SEQ ID NO: 2, and which has the activity of conferring resistance to L-homoserine to the bacterium.

5 . The method according to claim 4 , wherein said amino acid is branched chain amino acid.

6 . The method according to claim 5 , wherein said branched chain amino acid is L-valine or L-leucine.

7 . The method according to claim 4 , wherein the protein as defined in (C) or (D) is encoded by a DNA selected from the group consisting of:
(c) a DNA comprising the nucleotide sequence of nucleotide numbers 557 to 1171 in SEQ ID NO: 1; (d) a DNA which hybridizes with a nucleotide sequence of nucleotide numbers 557 to 1171 in SEQ ID NO: 1 under stringent conditions and which encodes a protein having the activity of conferring resistance to L-homoserine to the bacterium, wherein the stringent conditions comprise washing at 60° C., and at a salt concentration corresponding to 1×SSC and 0.1% SDS.

Assignees

• Ajinomoto Kk

Inventors

• Livshits Vitaliy Arkadyevich
• Zakataeva Natalia Pavlovna
• Aleshin Vladimir Veniaminovich
• Belareva Alla Valentinovna
• Tokhmakova Irina Lyvovna

Classifications

• C12P13/06

  Alanine; Leucine; Isoleucine; Serine; Homoserine · CPC title

• C07K14/245

  Escherichia (G) · CPC title

• C12P13/08 Primary

  Lysine; Diaminopimelic acid; Threonine; Valine · CPC title

• C12N1/20 Primary

  Bacteria; Culture media therefor · CPC title