-4Desaturases from euglena gracilis, expressing plants, and oils containing pufa

• US 20060218668A1
• Filed: 04/06/2004
• Published: 09/28/2006
• Est. Priority Date: 04/08/2003
• Status: Active Grant

• Current Assignee: BASF Plant Science GmbH (BASF SE)
• Original Assignee: BASF Plant Science GmbH (BASF SE)
• Inventors: Meyer, Astrid, Zähringer, Ulrich, Cirpus, Petra, Bauer, Jörg, Heinz, Ernst
• Granted Patent: US 7,629,503 B2
• Time in Patent Office: Days
• US Class Current: 800/281
• CPC Class Codes: C12N 15/8247 involving modified lipid metabolism, e.g. seed oil composition
• CPC Class Codes: C12N 9/0071 acting on paired donors with incorporation of molecular oxygen (1.14)

Abstract

The present invention relates to an improved process for the specific preparation of unsaturated ω-3 fatty acids, and to a process for preparing triglycerides having an increased content of unsaturated fatty acids, particularly of ω-3 fatty acids having more than three double bonds. The invention relates to the preparation of a transgenic organism, preferably a transgenic plant or a transgenic microorganism, having increased content of fatty acids, oils or lipids having Δ-4 double bonds owing to the expression of a Δ-4-desaturase from Euglena gracilis. The invention additionally relates to expression cassettes comprising a nucleic acid sequence, a vector and organisms comprising at least one nucleic acid sequence or one expression cassette. The invention additionally relates to unsaturated fatty acids and to triglycerides having an increased content of unsaturated fatty acids and to the use thereof. Fatty acids and triglycerides have a large number of uses in the food industry, in animal nutrition, cosmetics and in the drugs sector. They are suitable for a wide variety of uses depending on whether they are free saturated or unsaturated fatty acids or triglycerides having an increased content of saturated or unsaturated fatty acids.

Claims

1. An isolated nucleic acid sequence which codes for polypeptides having Δ

-4-desaturase activity, selected from the group;

a) of a nucleic acid sequence having the sequence depicted in SEQ ID NO;

1,

b) nucleic acid sequences which, as a result of the degeneracy of the genetic code, can be derived from the coding sequence comprised in SEQ ID NO;

1,

c) derivates of the nucleic acid sequence depicted in SEQ ID NO;

1, which code for polypeptides having the amino acid sequences depicted in SEQ ID NO;

2 or a nucleic acid sequence which code for a polypeptide having at least 40% homology at the amino acid level with SEQ ID NO;

2 and having a Δ

-4-desaturase activity.

2. The isolated nucleic acid sequence according to claim 1, where the sequence is derived from a plant.

3. The isolated nucleic acid sequence according to claim 1, where the sequence is derived from the class of Euglenophyceae.

4. An amino acid sequence which is encoded by an isolated nucleic acid sequence according to claim 1.

5. A gene construct comprising an isolated nucleic acid according to claim 1, where the nucleic acid is functionally connected to one or more regulatory signals.

6. The gene construct according to claim 5, wherein the nucleic acid construct comprises additional biosynthesis genes of fatty acid or lipid metabolism selected from the group of acyl-CoA dehydrogenase(s), acyl-ACP [=acyl carrier protein] desaturase(s), acyl-ACP thioesterase(s), fatty acid acyltransferase(s), acyl-CoA:

lysophospholipid acyltransferase(s), fatty acid synthase(s), fatty acid hydroxylase(s), acetyl-coenzyme A carboxylase(s), acyl-coenzyme A oxidase(s), fatty acid desaturase(s), fatty acid acetylenases, lipoxygenases, triacylglycerol lipases, allene oxide synthases, hydroperoxide lyases or fatty acid elongase(s).

7. The gene construct according to claim 5, wherein the nucleic acid construct comprises additional biosynthesis genes of fatty acid or lipid metabolism selected from the group of Δ

-4-desaturase, Δ

-5-desaturase, Δ

-6-desaturase, Δ

-8-desaturase, Δ

-9-desaturase, Δ

-12-desaturase, Δ

-5-elongase, Δ

-6-elongase or Δ

-9-elongase.

8. A vector comprising a nucleic acid according to claim 1.

9. A transgenic nonhuman organism comprising at least one nucleic acid according to claims 1 to 3, one gene construct according to claim 1.

10. The transgenic nonhuman organism according to claim 8, where the organism is a microorganism, a nonhuman animal or a plant.

11. The transgenic nonhuman organism according to claim 9, where the organism is a plant.

12. A process for producing polyunsaturated fatty acids, wherein the process comprises the culturing transgenic organism which comprises a nucleic acid according to claim 1 encoding a Δ

-4-desaturase which specifically desaturates ω

-3-fatty acids, and recovering the polyunsaturated fatty acids.

13. The process according to claim 12, where docosahexaenoic acid is produced in the process.

14. The process according to claim 12, where the polyunsaturated fatty acid molecules are isolated from the organism in the form of an oil, lipid or a free fatty acid.

15. The process according to claim 12, where the organism is a microorganism, a nonhuman animal or a plant.

16. The process according to claim 12, where the organism is a transgenic plant.

17. An oil, lipids or fatty acids or a fraction thereof produced by the process according to claim 14.

18. An oil, lipid or fatty acid composition which comprises polyunsaturated fatty acids produced by a process according to claim 12 and is derived from transgenic plants.

19. An animal feed, human foods, cosmetics or pharmaceuticals, which contains the oil, lipids or fatty acids according to claim 17.

20. An animal feed, human foods, cosmetics or pharmaceuticals, which contains the oil, lipids or fatty acids composition according to claim 18.