Desaturase

• US 20030152983A1
• Filed: 01/13/2003
• Published: 08/14/2003
• Est. Priority Date: 12/23/1997
• Status: Abandoned Application

• Current Assignee: University of Bristol
• Original Assignee: University of Bristol
• Inventors: Michaelson, Louise, Stobart, Keith, Napier, Johnathan A.
• Application Number:
  US10/340,779
• Publication Number:
  US 20030152983A1
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: A01K 2217/05 Animals comprising random inserted nucleic acids (transgenic)
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: C12N 15/8247 involving modified lipid metabolism, e.g. seed oil composition
• CPC Class Codes: C12N 9/0083 Miscellaneous (1.14.99)
• CPC Class Codes: C12P 7/6427 Polyunsaturated fatty acids [PUFA], i.e. having two or more double bonds in their backbone
• CPC Class Codes: C12P 7/6472 containing polyunsaturated fatty acid [PUFA] residues, i.e. having two or more double bonds in their backbone

Abstract

This invention relates to cDNA sequences encoding Δ5-fatty acid desaturases comprising the sequences shown in SEQ.1 and SEQ.2.

Claims

1. An isolated animal Δ

5-fatty acid desaturase and functional portions thereof.

3. A DNA sequence encoding a Δ

5-fatty acid desaturase according to claim 1 or claim 2.

4. A DNA sequence according to claim 3 and comprising at least a portion of the sequence shown in SEQ.2 and equivalents to that sequence, or to portions of that sequence, which encode a functional Δ

5-fatty acid desaturase by virtue of the degeneracy of the genetic code.

5. A DNA sequence according to claim 4 derived from a Caenorhabditis elegans DNA sequence.

6. A DNA sequence according to claim 3 encoding a functional Δ

5-fatty acid desaturase and comprising at least a portion of the sequence shown in SEQ.1 and equivalents to that sequence, or to portions of that sequence, which encode a functional Δ

5-fatty acid desaturase by virtue of the degeneracy of the genetic code.

7. A DNA sequence according to claim 6 derived from a Mortierella alpina DNA sequence.

8. A DNA sequence according to any one of claims 3 to 7 wherein the DNA sequence is functional in a mammal.

9. A DNA sequence according to claim 8 in which the DNA sequence is expressed in a mammal

10. A DNA sequence according to claim 9 wherein the DNA sequence is expressed in a human.

11. A DNA sequence obtained by modification of a functional natural gene encoding a Δ

-5 fatty acid desaturase according to claim 1 or claim 2.

12. A DNA sequence according to claim 11 wherein the modification includes modification by chemical, physical, or biological means without removing a catalytic activity of the enzyme which it encodes.

13. A DNA sequence according to claim 12 wherein the modification improves a catalytic activity of the enzyme which it encodes.

14. A DNA sequence according to claim 12 or 13 wherein the biological modification includes recombinant DNA methods and forced evolution techniques.

15. A DNA sequence according to claim 14 wherein the forced evolution technique is DNA shuffling.

16. A polypeptide encoded by a DNA sequence according to any of claims 3 to 15.

17. A polypeptide according to claim 16 wherein at least a portion of the polypeptide has the sequence shown in SEQ.3 or functional equivalents to that sequence or to portions of that sequence.

18. A polypeptide according to claim 16 wherein at least a portion of the polypeptide has the sequence shown in SEQ.4 or functional equivalents to that sequence or to portions of that sequence.

19. A polypeptide according to any of claims 16 to 18 wherein the polypeptide catalyses the conversion of dihomogamma linolenic acid to arachidonic acid.

20. A polypeptide according to any of claims 16 to 19 wherein the polypeptide has been modified without removing the catalytic activity of the encoded polypeptide.

21. A polypeptide according to claim 20 wherein the polypeptide has been modified in such a way as to introduce a specific level of saturation of a substrate at a specific location within the molecular structure of the substrate.

22. A vector containing a DNA sequence or any portion of a DNA sequence according to any of claims 3 to 15.

23. A method of producing polyunsaturated fatty acids comprising contacting a suitable substrate with a Δ

5-fatty acid desaturase according to claim 1 or 2 or a polypeptide according to claim 16 to 21.

24. A method of converting dihomogamma linolenic acid to arachidonic acid wherein said conversion is catalysed by a Δ

5-fatty acid desaturase according to claim 1 or 2 or a polypeptide or modified polypeptide according to any of claims 16 to 21.

25. An organism engineered to produce high levels of a polypeptide according to any of claims 16 to 21.

26. An organism engineered to produce high levels of a product of a reaction catalysed by a Δ

5-fatty acid desaturase according to claim 1 or 2 or by a polypeptide according to any one of claims 16 to 21.

27. An organism which has been engineered to carry out the method of claim 23 or claim 24.

28. An organism according to either of claims 26 and 27 wherein the organism is a microorganism.

29. An organism according to claim 28 wherein a microorganism is selected from algae, bacteria and fungi.

30. An organism according to claim 29 wherein a fungi includes phycomycetes.

31. An organism according to claim 28 wherein said microorganism is a yeast.

32. An organism according to any of claims 25 to 27 wherein the organism is a plant.

33. An organism according to claim 32 wherein the plant is selected from oil seed plants and tobacco.

34. An organism according to claim 33 wherein the oil seed plants are selected from oil seed rape, sunflower, cereals including maize, tobacco, legumes including peanut and soybean, safflower, oil palm, coconut and other palms, cotton, sesame, mustard, linseed, castor, borage and evening primrose.

35. A seed or other reproductive material derived from an organism according to claim 33 or claim 34.

36. An organism according to any of claims 25 to 27 wherein the organism is a mammal.

37. An isolated multienzyme pathway wherein the pathway includes a Δ

5 desaturase according to claim 1 or 2 or a polypeptide according to any of claims 16 to 21

38. A compound produced by a conversion of a substrate, wherein said conversion is catalysed by a Δ

5 desaturase according to claim 1 or 2 or by a polypeptide according to any of claims 16 to 21.

39. An intermediate compound produced by the reaction catalysed by a Δ

5 desaturase according to claim 1 or 2 or by a polypeptide according to any of claims 16 to 21.

40. A foodstuff or dietary supplement containing a polyunsaturated fatty acid produced by a method according to claim 23 or 24.

41. A pharmaceutical preparation containing a polyunsaturated fatty acid produced by a method according to claim 23 or 24.

42. Prostaglandins synthesised by a biosynthetic pathway including a catalytic activity of a Δ

5 desaturase according to claim 1 or 2 or by a polypeptide according to any of claims 16 to 21.

43. A method for modulation of prostaglandin synthesis by the control of the levels of expression of a DNA sequence according to any of claims 3 to 15.

44. A probe comprising all or part of a DNA sequence according to any of claims 3 to 15 or an equivalent RNA sequence.

45. A diagnostic or search probe comprising all or part of a Δ

5 desaturase according to claim 1 or 2 or of a polypeptide according to any of claims 16 to 21.

46. A method of isolating Δ

5 desaturases using a probe according to claim 44 or 45.

2. Isolated C. elegans Δ

5-fatty acid desaturase.