Mutants for the preparation of D-amino acids

Patent metadata

Field	Value
Publication number	US-9194009-B2
Application number	US-35721809-A
Country	US
Kind code	B2
Filing date	Jan 21, 2009
Priority date	Nov 4, 2002
Publication date	Nov 24, 2015
Grant date	Nov 24, 2015

Abstract

Escherichia. coli mutants which can be used for the synthesis of D-amino acids, and to such a process. The mutants are distinguished by deficiencies in particular enzymes which break down D-amino acids and include those which produce D-amino acids via the carbamoylase/hydantoinase route.

First claim

The invention claimed is:

1. An isolated Escherichia coli strain
having its dadA gene mutated or deleted so that said gene does not express a functional D-amino acid oxidase; and
having its dsdA gene mutated or deleted so that said gene does not express a functional D-serine dehydratase;
wherein said strain has been transformed with and expresses a D-carbamoylase gene and a D-hydantoinase gene,
wherein the strain is Escherichia coli strain DSM15182 (ET4), and
wherein the isolated strain exhibits reduced breakdown of produced D-amino acids when cultured in a suitable medium.

2. A process for preparing a D-amino acid comprising:
culturing in a suitable medium the isolated Escherichia coli strain of claim 1 , and
recovering a D-amino acid,
wherein the isolated strain exhibits reduced breakdown of produced D-amino acids when cultured.

3. The process of claim 2 , wherein the D-amino acid that is recovered is D-serine.

4. The process of claim 2 , wherein the D-amino acid that is recovered is D-methionine.

5. The process of claim 2 , wherein the D-amino acid that is recovered is D-tryptophan.

6. The process of claim 2 , wherein the D-amino acid that is recovered is D-phenylalanine.

7. The process of claim 2 , wherein the D-amino acid that is recovered is D-aminobutyric acid.

8. A method for producing a D-amino acid via the carbamoylase/hydantoinase route comprising culturing an isolated microorganism that has been transformed with and expresses a D-carbamoylase gene and a D-hydantoinase gene;
wherein said microorganism lacks at least one gene that expresses a functional D-amino oxidase and D-amino acid dehydratase;
wherein said microorganism is selected from the group consisting of Escherichia coli DCM15181 (ET3) and Escherichia coli DSM15182 (ET4), and
wherein the isolated strain exhibits reduced breakdown of produced D-amino acids when cultured.

9. The isolated Escherichia coli strain of claim 1 , wherein the isolated strain exhibits reduced breakdown of produced D-amino acids to less than 10% when cultured for at least 10 hours.

10. The process of claim 2 , wherein the isolated strain exhibits reduced breakdown of produced D-amino acids to less than 10% when cultured for at least 10 hours.

11. The process of claim 8 , wherein the isolated strain exhibits reduced breakdown of produced D-amino acids to less than 10% when cultured for at least 10 hours.

12. The isolated Escherichia coli strain of claim 1 , wherein the amino acid breakdown provided by the isolated strain is less than an amino acid breakdown provided by a strain which has unaltered dadA and dsdA genes.

13. The process of claim 2 , wherein the amino acid breakdown provided by the isolated strain is less than an amino acid breakdown provided by a strain which has unaltered dadA and dsdA genes.

14. The method of claim 8 , wherein the amino acid breakdown provided by the isolated strain is less than an amino acid breakdown provided by a strain which has unaltered dadA and dsdA genes.

15. An isolated Escherichia coli strain that has been transformed with and expresses a D-carbamoylase gene and a D-hydantoinase gene;
wherein said strain is Escherichia coli DCM15181 (ET3) that lacks a dadA gene that expresses a functional D-amino oxidase.

Assignees

• May Oliver
• Buchholz Stefan
• Schwarm Michael
• Drauz Karlheinz
• Turner Robert J
• Fotheringham Ian
• Evonik Degussa Gmbh

Inventors

• May Oliver
• Buchholz Stefan
• Schwarm Michael
• Drauz Karlheinz
• Turner Robert J
• Fotheringham Ian

Classifications

• C12P13/04 Primary

  Alpha- or beta- amino acids {(other amino acids C12P13/005)} · CPC title

• C12N9/86

  acting on amide bonds in cyclic amides, e.g. penicillinase {(3.5.2)} · CPC title

• C12R1/19 Primary

  Chemistry & Metallurgy · mapped topic

• C12P41/009

  by reactions involving hydantoins or carbamoylamino compounds · CPC title

• C12N9/90

  Isomerases (5.) · CPC title