Method for manufacturing cis-5-hydroxy-l-pipecolic acid

Patent metadata

Field	Value
Publication number	US-2017306367-A1
Application number	US-201515526031-A
Country	US
Kind code	A1
Filing date	Nov 2, 2015
Priority date	Nov 12, 2014
Publication date	Oct 26, 2017
Grant date	—

Abstract

A method for producing cis-5-hydroxy-L-pipecolic acid, the method comprising allowing a 2-oxoglutarate-dependent L-pipecolic acid hydroxylase to act on L-pipecolic acid to generate cis-5-hydroxy-L-pipecolic acid, wherein the 2-oxoglutarate-dependent L-pipecolic acid hydroxylase comprises the polypeptide (A), (B) or (C) below: (A) a polypeptide comprising the amino acid sequence represented by SEQ ID NO: 4 or 11; (B) a polypeptide comprising the amino acid sequence represented by SEQ ID NO: 4 or 11 except that one or several amino acids are deleted, substituted, and/or added, which polypeptide has 2-oxoglutarate-dependent
L-pipecolic acid hydroxylase activity; or (C) a polypeptide comprising an amino acid sequence with an identity of not less than 60% to the amino acid sequence represented by SEQ ID NO: 4 or 11, which polypeptide has 2-oxoglutarate-dependent L-pipecolic acid hydroxylase activity.

First claim

1 . A method for producing cis-5-hydroxy-L-pipecolic acid, the method comprising allowing a 2-oxoglutarate-dependent L-pipecolic acid hydroxylase, a microorganism or cell having the ability to produce the enzyme, a processed product of the microorganism or cell, and/or a culture liquid comprising the enzyme and obtained by culturing the microorganism or cell, to act on L-pipecolic acid to generate cis-5-hydroxy-L-pipecolic acid,
wherein the 2-oxoglutarate-dependent L-pipecolic acid hydroxylase comprises the polypeptide (A), (B) or (C) below: (A) a polypeptide comprising the amino acid sequence represented by SEQ ID NO: 4 or 11; (B) a polypeptide comprising the amino acid sequence represented by SEQ ID NO: 4 or 11 except that one or several amino acids are deleted, substituted, and/or added, which polypeptide has 2-oxoglutarate-dependent L-pipecolic acid hydroxylase activity; or (C) a polypeptide comprising an amino acid sequence with an identity of not less than 60% to the amino acid sequence represented by SEQ ID NO: 4 or 11, which polypeptide has 2-oxoglutarate-dependent L-pipecolic acid hydroxylase activity.

2 . The method for producing cis-5-hydroxy-L-pipecolic acid according to claim 1 , wherein DNA encoding the 2-oxoglutarate-dependent L-pipecolic acid hydroxylase comprises the DNA (D), (E) or (F) below:
(D) DNA comprising the nucleotide sequence represented by SEQ ID NO: 1, 9, or 10; (E) DNA comprising the nucleotide sequence represented by SEQ ID NO: 1, 9, or 10 except that one or several nucleotides are substituted, deleted, and/or added, which DNA encodes a polypeptide having 2-oxoglutarate-dependent L-pipecolic acid hydroxylase activity; or (F) DNA comprising a nucleotide sequence which hybridizes with the complementary strand of the nucleotide sequence represented by SEQ ID NO: 1, 9, or 10 under stringent conditions, which DNA encodes a polypeptide having 2-oxoglutarate-dependent L-pipecolic acid hydroxylase activity.

3 . The method for producing cis-5-hydroxy-L-pipecolic acid according to claim 1 , wherein the 2-oxoglutarate-dependent L-pipecolic acid hydroxylase, the microorganism or cell having the ability to produce the enzyme, the processed product of the microorganism or cell, and/or the culture liquid comprising the enzyme and obtained by culturing the microorganism or cell is/are allowed to act on said L-pipecolic acid in the presence of 2-oxoglutaric acid and ferrous ion.

4 . A 2-oxoglutarate-dependent L-pipecolic acid hydroxylase protein having an activity to act on L-pipecolic acid to generate cis-5-hydroxy-L-pipecolic acid and comprising the polypeptide (A), (B) or (C) below:
(A) a polypeptide comprising the amino acid sequence represented by SEQ ID NO: 4 or 11; (B) a polypeptide comprising the amino acid sequence represented by SEQ ID NO: 4 or 11 except that one or several amino acids are deleted, substituted, and/or added, which polypeptide has 2-oxoglutarate-dependent L-pipecolic acid hydroxylase activity; or (C) a polypeptide comprising an amino acid sequence with an identity of not less than 60% to the amino acid sequence represented by SEQ ID NO: 4 or 11, which polypeptide has 2-oxoglutarate-dependent L-pipecolic acid hydroxylase activity.

5 . A polypeptide comprising the amino acid sequence represented by SEQ ID NO: 11.

6 . The method for producing cis-5-hydroxy-L-pipecolic acid according to claim 2 , wherein the 2-oxoglutarate-dependent L-pipecolic acid hydroxylase, the microorganism or cell having the ability to produce the enzyme, the processed product of the microorganism or cell, and/or the culture liquid comprising the enzyme and obtained by culturing the microorganism or cell is/are allowed to act on said L-pipecolic acid in the presence of 2-oxoglutaric acid and ferrous ion.

Assignees

• Api Corp

Inventors

• Miyake Ryoma
• Dekishima Yasumasa

Classifications

• C12N9/0071 Primary

  acting on paired donors with incorporation of molecular oxygen (1.14) · CPC title

• C12P17/12 Primary

  containing a six-membered hetero ring · CPC title

• C12Y114/11

  with 2-oxoglutarate as one donor, and incorporation of one atom each of oxygen into both donors (1.14.11) · CPC title

• C12N15/09

  Recombinant DNA-technology · CPC title