Claims
- 1. An isolated nucleic acid that encodes the glutaryl 7-ACA amidase having the amino acid sequence of SEQ ID NO: 2.
- 2. The nucleic acid according to claim 1, wherein said nucleic acid comprises the sequence of SEQ ID NO: 1.
- 3. The nucleic acid according to claim 2, wherein said nucleic acid consists of the sequence of SEQ ID NO: 1.
- 4. An isolated nucleic acid of between 10 and 1722 nucleotides, which hybridizes under stringent conditions either to a DNA molecule consisting of SEQ ID NO: 1 or to a DNA molecule consisting of the complement of SEQ ID NO: 1.
- 5. The isolated nucleic acid of claim 4, wherein the nucleic acid is between 17 and 1722 nucleotides in length.
- 6. The isolated nucleic acid of claim 4, wherein the nucleic acid is between 20 and 1722 nucleotides in length.
- 7. A vector comprising a nucleic acid sequence according to claim 1.
- 8. A host cell transformed with the vector of claim 7.
- 9. The host cell of claim 8 wherein the host cell is a bacterium.
- 10. A vector comprising a nucleic acid sequence according to claim 4.
- 11. A host cell transformed with the vector of claim 10.
- 12. The host cell of claim 11 wherein the host cell is a bacterium.
- 13. A glutaryl 7-ACA amidase having the following characteristics: isolatable from Pseudomonas diminuta BS-203, catalyzes the hydrolysis of 7-β-(4-carboxybutanamido)-cephalosporanic acid to 7-aminocephalosporanic acid and glutaric acid, and is composed of two subunits having apparent molecular weights of 42 kd and 26 kd by SDS PAGE electrophoresis.
- 14. The glutaryl 7-ACA amidase according to claim 13, wherein said amidase comprises the amino acid sequence shown in SEQ ID NO: 2.
- 15. The glutaryl 7-ACA amidase according to claim 13, wherein said amidase consists of the amino acid sequence shown in SEQ ID NO: 2.
- 16. A glutaryl 7-ACA amidase having at least 80% homology to the amidase of claim 15.
- 17. The glutaryl 7-ACA amidase of claim 16, wherein the amino acid sequence comprises SEQ ID NO: 2 with up to 113 conservative amino acid substitutions.
- 18. The glutaryl 7-ACA amidase of claim 16, wherein the amino acid sequence comprises SEQ ID NO: 2 with the addition or deletion of up to 20 amino acid residues.
- 19. The glutaryl 7-ACA amidase of claim 13, wherein the glutaryl 7-ACA amidase is immobilized on an insoluble support.
- 20. An isolated nucleic acid that encodes a glutaryl 7-ACA amidase of claim 16.
- 21. A vector comprising a nucleic acid sequence according to claim 20.
- 22. A host cell transformed with the vector of claim 21.
- 23. A method for preparing a glutaryl 7-ACA amidase from Pseudomonas diminuta BS-203, said method comprising
(a) culturing the Pseudomonas diminuta BS-203 bacterium under aerobic conditions in a suitable medium; and (b) recovering from the obtained culture a protein fraction having glutaryl 7-ACA amidase activity.
- 24. A method for preparing a glutaryl 7-ACA amidase, said method comprising:
(a) culturing host cells according to claim 8 under conditions suitable for expression of the glutaryl 7-ACA amidase; and (b) recovering from the obtained culture a protein fraction having glutaryl 7-ACA amidase activity.
- 25. A method for preparing a glutaryl 7-ACA amidase, said method comprising:
(a) culturing host cells according to claim 11 under conditions suitable for expression of the glutaryl 7-ACA amidase; and (b) recovering from the obtained culture a protein fraction having glutaryl 7-ACA amidase activity.
- 26. A method for preparing a glutaryl 7-ACA amidase, said method comprising:
(a) culturing host cells according to claim 22 under conditions suitable for expression of the glutaryl 7-ACA amidase; and (b) recovering from the obtained culture a protein fraction having glutaryl 7-ACA amidase activity.
- 27. A process for obtaining 7-aminocephalosporanic acid from a 7-β-(acylamido)cephalosporanic acid, comprising contacting a 7-β-(acylamido)cephalosporanic acid with the glutaryl 7-ACA amidase according to claim 13 in a suitable solvent.
- 28. A process for obtaining 7-aminocephalosporanic acid from a 7-β-(acylamido)cephalosporanic acid, comprising contacting a 7-β-(acylamido)cephalosporanic acid with the glutaryl 7-ACA amidase according to claim 14 in a suitable solvent.
- 29. A process for obtaining 7-aminocephalosporanic acid from a 7-β-(acylamido)cephalosporanic acid, comprising contacting a 7-β-(acylamido)cephalosporanic acid with the glutaryl 7-ACA amidase according to claim 15 in a suitable solvent.
- 30. A process for obtaining 7-aminocephalosporanic acid from a 7-β-(acylamido)cephalosporanic acid, comprising contacting a 7-β-(acylamido)cephalosporanic acid with the glutaryl 7-ACA amidase according to claim 19 in a suitable solvent.
- 31. The process of claim 27, wherein the 7-β-(acylamido)cephalosporanic acid is 7-β-(4-carboxybutanamido)cephalosporanic acid.
- 32. The process of claim 28, wherein the 7-β-(acylamido)cephalosporanic acid is 7-β-(4-carboxybutanamido)cephalosporanic acid.
- 33. The process of claim 29, wherein the 7-β-(acylamido)cephalosporanic acid is 7-β-(4-carboxybutanamido)cephalosporanic acid.
- 34. The process of claim 30, wherein the 7-β-(acylamido)cephalosporanic acid is 7-β-(4-carboxybutanamido)cephalosporanic acid.
- 35. A process for obtaining desacetyl 7-aminocephalosporanic acid from a desacetyl 7-β-(acylamido)cephalosporanic acid, comprising contacting a desacetyl 7-β-(acylamido)cephalosporanic acid with the glutaryl 7-ACA amidase according to claim 13 in a suitable solvent.
- 36. A process for obtaining desacetyl 7-aminocephalosporanic acid from a desacetyl 7-β-(acylamido)cephalosporanic acid, comprising contacting a desacetyl 7-β-(acylamido)cephalosporanic acid with the glutaryl 7-ACA amidase according to claim 14 in a suitable solvent.
- 37. A process for obtaining desacetyl 7-aminocephalosporanic acid from a desacetyl 7-β-(acylamido)cephalosporanic acid, comprising contacting a desacetyl 7-β-(acylamido)cephalosporanic acid with the glutaryl 7-ACA amidase according to claim 15 in a suitable solvent.
- 38. A process for obtaining desacetyl 7-aminocephalosporanic acid from a desacetyl 7-β-(acylamido)cephalosporanic acid, comprising contacting a desacetyl 7-β-(acylamido)cephalosporanic acid with the glutaryl 7-ACA amidase according to claim 19 in a suitable solvent.
- 39. The process of claim 35, wherein the desacetyl 7-β-(acylamido)cephalosporanic acid is desacetyl 7-β-(4-carboxybutanamido) cephalosporanic acid.
- 40. The process of claim 36, wherein the desacetyl 7-β-(acylamido)cephalosporanic acid is desacetyl 7-β-(4-carboxybutanamido) cephalosporanic acid.
- 41. The process of claim 37, wherein the desacetyl 7-β-(acylamido)cephalosporanic acid is desacetyl 7-β-(4-carboxybutanamido) cephalosporanic acid.
- 42. The process of claim 38, wherein the desacetyl 7-β-(acylamido)cephalosporanic acid is desacetyl 7-β-(4-carboxybutanamido) cephalosporanic acid.
Parent Case Info
[0001] This application claims the benefit of provisional U.S. Application Serial No. 60/234,532, filed Sep. 22, 2000, which is incorporated herein by reference in its entirety.
Provisional Applications (1)
|
Number |
Date |
Country |
|
60234532 |
Sep 2000 |
US |
Divisions (1)
|
Number |
Date |
Country |
Parent |
09957960 |
Sep 2001 |
US |
Child |
10346662 |
Jan 2003 |
US |