Method for the production of polyunsaturated fatty acids

• US 7,537,920 B2
• Filed: 01/29/2004
• Issued: 05/26/2009
• Est. Priority Date: 02/27/2003
• Status: Expired due to Fees

• Current Assignee: BASF Plant Science GmbH (BASF SE)
• Original Assignee: BASF Plant Science GmbH (BASF SE)
• Inventors: Renz, Andreas, Zank, Thorsten, Abbadi, Amine, Heinz, Ernst, Domergue, Frederic
• Primary Examiner(s): Saidha; Tekchand
• Application Number:
  US10/547,447
• Publication Number:
  US 20060168687A1
• Time in Patent Office: 1,944 Days
• Field of Search: 435/194, 435/252.3, 435/320.1, 536/23.2
• US Class Current: 435/194
• CPC Class Codes: C12N 15/8247 involving modified lipid metabolism, e.g. seed oil composition
• CPC Class Codes: C12N 9/1029 transferring groups other than amino-acyl groups (2.3.1)
• 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 7/6432 Eicosapentaenoic acids [EPA]
• CPC Class Codes: C12P 7/6434 Docosahexenoic acids [DHA]
• CPC Class Codes: C12P 7/6472 containing polyunsaturated fatty acid [PUFA] residues, i.e. having two or more double bonds in their backbone

Abstract

The present invention relates to a process for producing polyunsaturated fatty acids in an organism by introducing nucleic acids into the organism which code for polypeptides having acyl-CoA:lysophospholipid a cyltransferase activity. Advantageously, these nucleic acid sequences may, if appropriate together with further nucleic acid sequences coding for biosynthesis polypeptides of the fatty acid or lipid metabolism, be expressed in the transgenic organism. The invention furthermore relates to the nucleic acid sequences, to nucleic acid constructs comprising the nucleic acid sequences of the invention, to vectors comprising the nucleic acid sequences and/or the nucleic acid constructs and to transgenic organisms comprising the abovementioned nucleic acid sequences, nucleic acid constructs and/or vectors. A further part of the invention relates to oils, lipids and/or fatty acids produced by the process of the invention and to their use.

Claims

1. An isolated nucleic acid coding for a polypeptide having acyl-CoA:

lysophospholipid-acyltransferase activity, wherein the isolated nucleic acid comprises a nucleotide sequence having at least 95% identity to the nucleotide sequence of SEQ ID NO;

1, or encodes a polypeptide having at least 95% identity to the polypeptide sequence of SEQ ID NO;

2, wherein the acyl-CoA;

lysophospholipid acyltransferase encoded by said nucleic acid specifically uses C₁₆, C₁₈-, C₂₀- or C₂₂-fatty acids having at least one double bond as substrate.

2. The isolated nucleic acid of claim 1, wherein the nucleic acid comprises the nucleotide sequence of SEQ ID NO:

1 or encodes a polypeptide comprising the amino acid sequence of SEQ ID NO;

2.

3. The isolated nucleic acid of claim 1, which is obtained from a eukaryote.

4. An expression cassette comprising the isolated nucleic acid of claim 1, wherein said nucleic acid is functionally linked to one or more regulatory signals.

5. The expression cassette of claim 4, further comprising additional biosynthetic genes of the fatty acid or lipid metabolism, selected from the group consisting of acyl-CoA dehydrogenase(s), acyl-ACP[=acyl carrier protein] desaturase(s), acyl-ACP thioesterase(s), fatty acid 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, allenoxide synthases, hydroperoxide lyases and fatty acid elongase(s).

6. The expression cassette of claim 4, further comprising additional biosynthetic genes of the fatty acid or lipid metabolism, selected from the group consisting 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 and Δ

9-elongase.

7. A vector comprising the nucleic acid of claim 1, or an expression cassette comprising said nucleic acid functionally linked to one or more regulatory signals.

8. A transgenic plant or microorganism comprising at least one nucleic acid of claim 1, an expression cassette comprising said nucleic acid functionally linked to one or more regulatory signals, or a vector comprising said nucleic acid or said expression cassette.

9. A process for producing polyunsaturated fatty acids in an organism, wherein said process comprises:

a) introducing into an organism at least one nucleic acid coding for a polypeptide having acyl-CoA;

lysophospholipid-acyltransferase activity, and

b) culturing and harvesting said organism,

wherein the nucleic acid comprises a nucleotide sequence selected from the group consisting of;

i) the nucleotide sequence of SEQ ID NO;

1,

ii) a nucleotide sequence having at least 95% identity to the nucleotide sequence of SEQ ID NO;

1,

iii) a nucleotide sequence encoding the polypeptide sequence of SEQ ID NO;

2, and

iv) a nucleotide sequence encoding a polypeptide having at least 95% identity to the polypeptide sequence of SEQ ID NO;

2.

10. The process of claim 9, wherein the process further comprises introducing additional nucleic acid sequences into said organism, wherein the additional nucleic acid sequences code for polypeptides of the fatty acid or lipid metabolism selected from the group consisting of acyl-CoA-dehydrogenase(s), acyl-ACP[=acyl carrier protein] desaturase(s), acyl-ACP thioesterase(s), fatty acid 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, allenoxide synthases, hydroperoxide lyases and fatty acid elongase(s).

11. The process of claim 9, wherein the process further comprises introducing additional nucleic acid sequences into the organism, wherein the additional nucleic acid sequences code for polypeptides selected from the group consisting 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 and Δ

9-elongase activity.

12. The process of claim 9, wherein the polypeptide having acyl-CoA:

lysophospholipid acyltransferase activity uses C16-, C18-, C20- or C22-fatty acids as substrate.

13. The process of claim 9, wherein the polyunsaturated fatty acids are isolated from the organism in the form of an oil, lipid or a free fatty acid.

14. The process of claim 9, wherein the polyunsaturated fatty acid produced in said process is a C₁₈-, C₂₀- or C₂₂-fatty acid having at least two double bonds in the molecule.

15. The process of claim 9, wherein the polyunsaturated fatty acid produced is dihomo-γ

-linolenic acid, arachidonic acid, eisosapentaenoic acid, docosapentaenoic acid or docosahexaenoic acid.

16. The process of claim 9, wherein the organism is a microorganism or a plant.

17. The process of claim 16, wherein the plant is an oil crop plant.