Claims
- 1. An isolated polynucleotide encoding the polypeptide of SEQ ID NO: 4.
- 2. An isolated polynucleotide encoding a polypeptide having the same biological activity as the polypeptide encoded by SEQ ID NO: 1, the polynucleotide having the nucleotide sequence of SEQ ID NO: 1 or a nucleotide sequence that hybridizes thereto under conditions of 5×SSC, 50% formamide and 42° C.
- 3. The isolated polynucleotide of claim 2, wherein the polynucleotide encodes the polypeptide encoded by SEQ ID NO: 1.
- 4. The isolated polynucleotide of claim 2, wherein the polynucleotide is isolated from a bacterium.
- 5. The isolated polynucleotide of claim 4, wherein the bacterium is a gram negative bacterium.
- 6. The isolated polynucleotide of claim 5, wherein the bacterium is Acinetobacter calcoacefcus.
- 7. The isolated polynucleotide of claim 2, comprising the nucleic acid sequence of SEQ ID NO: 1 or the complement thereof.
- 8. A recombinant vector comprising an isolated polynucleotide encoding a polypeptide having wax ester synthase activity and having the nucleotide sequence of SEQ ID NO: 1 or a nucleotide sequence that hybridizes thereto under conditions of 5×SSC, 50% formamide and 42° C.
- 9. The recombinant vector of claim 8 further comprising at least one additional sequence selected from the group consisting of
(a) a regulatory sequence operatively coupled to the polynucleotide; (b) a selection marker operatively coupled to the polynucleotide; (c) a marker sequence operatively coupled to the polynucleotide; (d) a purification moiety operatively coupled to the polynucleotide; (e) a secretion sequence operatively coupled to the polynucleotide; and (f) a targeting sequence operatively coupled to the polynucleotide.
- 10. The recombinant vector of claim 9, wherein the vector comprises a developmentally-regulated, an organelle-specific, a tissue-specific, an inducible, a constitutive or a cell-specific promoter.
- 11. The recombinant vector of claim 8, wherein the recombinant vector is selected from the group consisting of pKS:waxEB19, pSER200-4:wax, pKS:wax, pBBRIMCS-2:wax, and pKS :wax-His6C.
- 12. The recombinant vector of claim 8, wherein the polypeptide has DGAT activity.
- 13. The recombinant vector of claim 8, wherein the polynucleotide encodes at least one conserved amino acid sequence selected from the group consisting of HHAXVDGV, NDVVLA, GALRXYL, PLXAMVP, ISNVPGP, REPLYXNGA.
- 14. A host cell transformed with the recombinant vector of claim 8.
- 15. The host cell of claim 14, wherein the host cell is selected from the group consisting of a mammalian cell, plant cell, insect cell, yeast cell, fungus cell, and bacterial cell.
- 16. The host cell of claim 15, wherein the host cell is selected from E. coli and P. citronellolis.
- 17. The host cell of claim 14, wherein the host cell expresses a protein encoded by the recombinant vector.
- 18. The host cell of claim 17, wherein the expressed protein is secreted by the host cell.
- 19. A protein or polypeptide encoded by the nucleotide sequence of SEQ ID NO: 1 or a nucleotide sequence that hybridizes thereto under conditions of 5×SSC, 50% formamide and 42° C. and having wax ester synthase activity.
- 20. The protein or polypeptide of claim 19, further defined as comprising at least one conserved amino acid sequence selected from the group consisting of HHAXVDGV, NDVVLA, GALRXYL, PLXAMVP, ISNVPGP, REPLYXNGA.
- 21. The protein or polypeptide of claim 19, further defined as having DGAT activity.
- 22. A polypeptide comprising the amino acid sequence of SEQ ID NO: 4, or a fragment thereof having wax ester synthase activity.
- 23. The polypeptide of claim 22, wherein one or more of the amino acids have been substituted with a conserved amino acid and the biological function of the protein has been maintained.
- 24. The polypeptide of claim 22, wherein one or more of the amino acid residues includes a substituent group.
- 25. The polypeptide of claim 19, further defined as having DGAT activity.
- 26. A method for producing a wax ester comprising culturing a host cell of claim 14 under conditions permitting expression of the polypeptide having wax ester synthase activity.
- 27. The method of claim 26, further comprising isolating the polypeptide having wax ester synthase activity from the host cell or from the medium in which the host cell is cultured.
- 28. The method of claim 26, wherein the host cell is cultured in the presence of at least one substrate of said was ester synthase activity and/or of DGAT activity.
- 29. The method of claim 26, wherein the host cell is a bacterial cell.
- 30. A method of modifying accumulation of wax esters in a host cell comprising transforming a host cell with a recombinant construct containing a regulatory sequence operably linked to a nucleic acid sequence, said nucleic acid sequence encoding a polypeptide having wax ester synthase activity, and culturing said host cell under conditions wherein said host cell expresses the polypeptide, whereby the host cell has a modified wax ester composition compared to host cells without the recombinant construct.
- 31. The method of claim 30, wherein said polypeptide having wax ester synthase activity is a bacterial wax ester synthase polypeptide.
- 32. The method of claim 30, wherein said polynucleotide encoding a polypeptide having wax ester synthase activity is from A. calcoaceticus.
- 33. The method of claim 30, wherein said nucleic acid sequence has the nucleotide sequence of SEQ ID NO: 1, or a nucleotide sequence that hybridizes thereto under conditions of 5×SSC, 50% formamide and 42° C. and encodes a polypeptide having the same biological activity as the polypeptide encoded by SEQ ID NO: 1.
- 34. The method of claim 30, wherein said regulatory sequence comprises a constitutive promoter.
- 35. The method of claim 30, wherein said regulatory sequence comprises an inducible promoter.
- 36. The method of claim 30, wherein the polypeptide has DGAT activity.
- 37. The method of claim 30, wherein said host cell is a bacterial cell.
- 38. The method of claim 37, wherein the bacterial cell is E. coli or P. citronellolis.
- 39. The method of claim 30, wherein the modified accumulation of wax esters is an increase in the accumulation of wax esters.
- 40. A method of altering the DGAT activity of a host cell comprising transforming the cell with the recombinant vector of claim 12.
- 41. The method of claim 40, wherein the host cell is selected from the group consisting of a mammalian cell, plant cell, insect cell, yeast cell, fungus cell, and bacterial cell.
- 42. The method of claim 41, wherein the host cell is selected from E. coli and P. citronellolis.
Parent Case Info
[0001] This application claims priority to U.S. Provisional Patent Application Serial No. 60/360,774, filed on Mar. 1, 2002, the entire disclosure of which is specifically incorporated herein by reference.
Provisional Applications (1)
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Number |
Date |
Country |
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60360774 |
Mar 2002 |
US |