Method for the production by fermentation of sulphur-containing fine chemicals (metF)
View Patent ↗The invention relates to methods for the fermentative production of sulfur-containing fine chemicals, in particular L-methionine, by using bacteria which express a nucleotide sequence coding for a methionine synthase (metF) gene.
1. A method for the fermentative production of L-methionine, which comprises the following steps:
a) fermenting in a medium cells of a coryneform bacterium for producing L-methionine, the coryneform bacteria expressing at least one heterologous nucleotide sequence which codes for a protein with methylenetetrahydrofolate reductase (metF) activity, wherein said heterologous nucleotide sequence comprises a nucleotide sequence encoding a metF protein having an amino acid sequence as set forth in SEQ ID NO: 2 or comprises a nucleotide sequence encoding a metF protein having an amino acid sequence with 95% homology or more to the sequence as set forth in SEQ ID NO: 2;
b) concentrating L-methionine in the medium or in the bacterial cells, and
c) isolating L-methionine.
2. The method as claimed in claim 1 , wherein the metF-encoding sequence comprises a coding sequence as set forth in SEQ ID NO:1.
3. The method as claimed in claim 1 , wherein the metF-encoding sequence codes for a protein with metF activity, said protein comprising an amino acid sequence as set forth in SEQ ID NO:2.
4. The method as claimed in claim 1 , wherein the coding metF sequence is a DNA or RNA which can be replicated in coryneform bacteria or is stably integrated into the chromosome.
5. The method as claimed in claim 4 , wherein
a) a bacteria strain transformed with a plasmid vector carrying at least one copy of the coding metF sequence under the control of regulatory sequences is used, or
b) a strain in which the coding metF sequence has been integrated into the bacteria chromosome is used.
6. The method as claimed in claim 1 , wherein the coding metF sequence is overexpressed.
7. The method as claimed in claim 1 , wherein bacteria are fermented in which additionally at least one further gene of the biosynthetic pathway of L-methionine has been overexpressed.
8. The method as claimed in claim 1 , wherein coryneform bacteria are fermented in which, at the same time, at least one of the genes from among
a) a lysC gene derived from a coryneform bacterium, which encodes an aspartate kinase,
b) the glyceraldehyde-3-phosphate dehydrogenase-encoding gene gap,
c) the 3-phosphoglycerate kinase-encoding gene pgk,
d) the pyruvate carboxylase-encoding gene pyc,
e) the triose phosphate isomerase-encoding gene tpi,
f) the homoserine O-acetyltransferase-encoding gene metA,
g) the cystathionine gamma-synthase-encoding gene metB,
h) the cystathionine gamma-lyase-encoding gene metC,
i) the serine hydroxymethyltransferase-encoding gene glyA,
j) the O-acetylhomoserine sulfhydrylase-encoding gene metY,
k) the vitamin B12-dependent methionine synthase-encoding gene metH,
l) the phosphoserine aminotransferase-encoding gene serC,
m) the phosphoserine phosphatase-encoding gene serB,
n) the serine acetyltransferase-encoding gene cysE, or
o) the horn gene, which encodes a homoserine dehydrogenase, is overexpressed.
9. The method as claimed in claim 1 , wherein the coryneform bacterium is of the species Corynebacterium glutamicum.
10. A method for the production of L-methionine, which comprises the following steps:
a) fermenting in a medium cells of a coryneform bacterium fur producing of L-methionine, said coryneform bacteria expressing at least one heterologous nucleotide sequence which codes for a protein with methylenetetrahydrofolate reductase (metF) activity, wherein the heterologous nucleotide sequence comprises a nucleotide sequence having 95% identity or more to the sequence as set forth in SEQ ID NO; 1;
b) concentrating L-methionine in the medium or in the bacterial cells; and
c) isolating L-methionine.
11. The method of claim 10 , wherein the coding metF sequence is a DNA or RNA which can be replicated in coryneform bacteria or is stably integrated into the chromosome.
12. The method of claim 10 , wherein
a) a bacteria strain transformed with a plasmid vector carrying at least one copy of the coding metF sequence under the control of regulatory sequences is used, or
b) a strain in which the coding metF sequence has been integrated into the bacteria chromosome is used.
13. The method of claim 10 , wherein the coding metF sequence is overexpressed.
14. The method of claim 10 , wherein bacteria are fermented in which additionally at least one further gene of the biosynthetic pathway of L-methionine has been overexpressed.
15. The method of claim 10 , wherein the coryneform bacterium is of the species Corynebacterium glutamicum.
16. The method as claimed in claim 1 , wherein coryneform bacteria are fermented in which, at the same time, a lysC gene derived from a coryneform bacterium, which encodes an aspartate kinase, is overexpressed.
17. The method as claimed in claim 16 , wherein the lysC gene is derived from C. glutamicum .