Claims
- 1. An expression vector comprising a DNA encoding an L-sorbose dehydrogenase and a DNA encoding an L-sorbosone dehydrogenase,
- wherein the L-sorbose dehydrogenase has the amino acid sequence of SEQ ID NO: 1 and the L-sorbosone dehydrogenase has the amino acid sequence of SEQ ID NO: 2.
- 2. The expression vector of claim 1, wherein the DNA encoding the L-sorbose dehydrogenase and the DNA encoding the L-sorbosone dehydrogenase are constructed such that transcriptions occur successively from one promoter, or a transcription of each DNA occurs under the control of a different promoter.
- 3. The expression vector of claim 1, wherein the promoter is a promoter region of the L-sorbosone dehydrogenase gene or a part thereof having a promoter activity.
- 4. The expression vector of claim 1, wherein the promoter is derived from Escherichia coli.
- 5. The expression vector of claim 1, wherein the promoter is Escherichia coli promoter tufB, .lambda. PL, trp or tac.
- 6. The expression vector of claim 1, which is suitable for transforming a host microorganism capable of producing L-sorbose at high yields from D-sorbitol, which has no or low 2-keto-L-gulonic acid-decomposing activity.
- 7. The expression vector of claim 1, which is suitable for transforming a host microorganism capable of producing L-sorbose at high yields from D-sorbitol, which has no or low 2-keto-L-gulonic acid-decomposing activity, and no or low L-idonic acid-producing activity.
- 8. The expression vector of claim 1, wherein the DNA encoding an L-sorbose dehydrogenase and a DNA encoding an L-sorbosone dehydrogenase are obtained from a bacterium belonging to the genus Gluconobacter.
- 9. The expression vector of claim 1, wherein the DNA encoding an L-sorbose dehydrogenase and a DNA encoding an L-sorbosone dehydrogenase are obtained from Gluconobacter oxydans.
- 10. The expression vector of claim 1, wherein the DNA encoding the L-sorbose dehydrogenase and the DNA encoding the L-sorbosone dehydrogenase are DNAs encoding said enzymes derived from Gluconobacter oxydans T-100.
- 11. A transformant obtained by transforming a host cell with the expression vector of claim 1.
- 12. A transformant which produces a 2-keto-L-gulonic acid from D-sorbitol, which is obtained by transforming a host microorganism, which produces L-sorbose from D-sorbitol, with the expression vector of claim 1.
- 13. A transformant,
- (1) which produces 37 to 91 mg/ml of 2-keto-L-gulonic acid from D-sorbitol; and
- (2) which is obtained by transforming a host microorganism, which produces L-sorbose from D-sorbitol, with the expression vector of claim 1.
- 14. The transformant of claim 13, which produces 42 to 91 mg/ml of 2-keto-L-gulonic acid from D-sorbitol.
- 15. The transformant of claim 13, which produces 56 to 91 mg/ml of 2-keto-L-gulonic acid from D-sorbitol.
- 16. A transformant obtained by transforming a host cell with the expression vector of claim 1, wherein the transformant produces an undetectable quantity of L-idonic acid as measured by absorbance at 210 nm.
- 17. A process for producing a 2-keto-L-gulonic acid, comprising culturing the transformant of claim 11 in a medium containing D-sorbitol, thereby producing 2-keto-L-gulonic acid, and harvesting the 2-keto-L-gulonic acid.
- 18. A process for producing a 2-keto-L-gulonic acid, comprising culturing the transformant of claim 16 in a medium containing D-sorbitol, thereby producing 2-keto-L-gulonic acid, and harvesting the 2-keto-L-gulonic acid.
- 19. The host microorganism of claim 16, which belongs to the genus Gluconobacter or Acetobacter.
- 20. A process for producing a 2-keto-L-gulonic acid, comprising culturing the transformant of claim 11, in a medium containing D-sorbitol, and harvesting the 2-keto-L-gulonic acid from the obtained culture.
- 21. A process for producing a 2-keto-L-gulonic acid, comprising culturing the transformant of claim 13 in a medium containing D-sorbitol, and harvesting the 2-keto-L-gulonic acid from the culture.
- 22. A method of transforming a host microorganism, comprising:
- culturing a host microorganism in a culture medium containing D-mannitol; placing the cultured host cell in a solution containing an expression vector comprising a DNA encoding an L-sorbose dehydrogenase and a DNA encoding an L-sorbosone dehydrogenase, wherein the L-sorbose dehydrogenase has the amino acid sequence of SEQ ID NO:1 and the L-sorbosone dehydrogenase has the amino acid sequence of SEQ ID NO:2; and
- subjecting the cultured host cell to electroporation.
Priority Claims (1)
Number |
Date |
Country |
Kind |
6-28612 |
Feb 1994 |
JPX |
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Parent Case Info
This is a continuation of application Ser. No. 08/696,834 filed Sep. 24, 1996, now U.S. Pat. No. 5,834,263, which was filed as International Application No. PCT/JP95/00285 filed on Feb. 24, 1995.
US Referenced Citations (4)
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4902617 |
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|
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Non-Patent Literature Citations (1)
Entry |
Balbas et al. (1990) Methods in Enzymology, vol. 185, pp. 14-37. |
Continuations (1)
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Number |
Date |
Country |
Parent |
696834 |
Sep 1996 |
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