Elongase genes and uses thereof

• US 6,677,145 B2
• Filed: 07/11/2001
• Issued: 01/13/2004
• Est. Priority Date: 09/02/1998
• Status: Expired due to Term

• Current Assignee: Abbott Laboratories
• Original Assignee: Abbott Laboratories
• Inventors: Mukerji, Pradip, Huang, Yung-Sheng, Pereira, Suzette L., Leonard, Amanda Eun-Yeong
• Primary Examiner(s): Prouty, Rebecca E.
• Assistant Examiner(s): Swope, Sheridan
• Application Number:
  US09/903,456
• Publication Number:
  US 20020138874A1
• Time in Patent Office: 916 Days
• Field of Search: 435/193, 435/252.33, 435/320.1, 435/252.3, 435/348, 435/419, 435/254.11, 435/254.21, 435/254.22, 435/254.3, 435/254.4, 435/254.5, 435/254.6, 435/252.31, 435/328, 435/254.23, 536/23.2
• US Class Current: 435/193
• CPC Class Codes: A01K 2217/05 Animals comprising random inserted nucleic acids (transgenic)
• CPC Class Codes: A23D 9/00 Other edible oils or fats, e.g. shortenings, cooking oils
• CPC Class Codes: A23K 20/158 Fatty acids; Fats; Products containing oils or fats
• CPC Class Codes: A61K 2800/86 Products or compounds obtained by genetic engineering
• CPC Class Codes: A61K 31/202 having three or more double bonds, e.g. linolenic eicosanoids, e.g. leukotrienes A61K31/557
• 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: A61K 8/361 Carboxylic acids having more than seven carbon atoms in an unbroken chain; Salts or anhydrides thereof
• CPC Class Codes: A61K 8/606 Nucleosides; Nucleotides; Nucleic acids
• CPC Class Codes: A61K 8/64 Proteins; Peptides; Derivatives or degradation products thereof
• CPC Class Codes: A61P 3/02 Nutrients, e.g. vitamins, minerals
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: A61Q 19/00 Preparations for care of the skin
• 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 subject invention relates to the identification of several genes involved in the elongation of polyunsaturated acids (i.e., “elongases”) and to uses thereof. At least two of these genes are also involved in the elongation of monounsaturated fatty acids. In particular, elongase is utilized in the conversion of gamma linolenic acid (GLA) to dihomogamma linolenic acid (DGLA) and in the conversion of AA to adrenic acid (ADA), or eicosapentaenoic acid (EPA) to ω3-docosapentaenoic acid (DPA). DGLA may be utilized in the production of polyunsaturated fatty acids, such as arachidonic acid (AA), docosahexaenoic acid (DHA), EPA, adrenic acid, ω6-docosapentaenoic acid or ω3-docosapentaenoic acid which may be added to pharmaceutical compositions, nutritional compositions, animal feeds, as well as other products such as cosmetics.

Claims

1. An isolated nucleic acid sequence comprising or complementary to a nucleic acid sequence encoding a polypeptide having elongase activity, wherein the amino acid sequence of said polypeptide has at least 80% amino acid sequence identity to SEQ ID NO:

7.

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

7.

3. The isolated nucleic acid sequence of claims 1 or 2 wherein said sequence encodes a functionally active elongase which utilizes a polyunsaturated fatty acid as a substrate.

4. The isolated nucleic acid sequence of claim 1 wherein said sequence is derived from the genus Thraustochytrium.

5. The isolated nucleic acid sequence of claim 4 wherein said sequence is derived from Thraustochytrium aureum.

6. A method of producing an elongase enzyme comprising the steps of:

a) isolating a nucleotide sequence comprising SEQ ID NO;

7 (FIG. 72);

b) constructing a vector comprising;

i) said isolated nucleotide sequence operably linked to ii) a promoter;

c) introducing said vector into a host cell under time and conditions sufficient for expression of said elongase enzyme.

7. The method of claim 6 wherein said host cell is selected from the group consisting of a eukaryotic cell or a prokaryotic cell.

8. The method of claim 7 wherein said prokaryotic cell is selected from the group consisting of E. coli, Cyanobacteria, and B. subtilis.

9. The method of claim 7 wherein said eukaryotic cell is selected from the group consisting of a mammalian cell, an insect cell, a plant cell and a fungal cell.

10. The method of claim 9 wherein said fungal cell is selected from the group consisting of Saccharomyces spp., Candida spp., Lipomyces starkey, Yarrowia spp., Kluyveromyces spp., Hansenula spp., Aspergillus spp., Penicillium spp., Neurospora spp., Trichoderma spp. and Pichia spp.

11. The method of claim 10 wherein said fungal cell is a yeast cell selected from the group consisting of Saccharomyces spp., Candida spp., Hansenula spp. and Pichia spp.

12. The method of claim 11 wherein said yeast cell is Saccharomyces cerevisiae.

13. A vector comprising:

a) a nucleotide sequence comprising SEQ ID NO;

7 (FIG.

     72) operably linked to b) a promoter.

14. A host cell comprising said vector of claim 13.

15. The host cell of claim 14 wherein said host cell is selected from the group consisting of a eukaryotic cell or a prokaryotic cell.

16. The host cell of claim 15 wherein said prokaryotic cell is selected from the group consisting of E. coli, Cyanobacteria, and B. subtilis.

17. The host cell of claim 15 wherein said eukaryotic cell is selected from the group consisting of a mammalian cell, an insect cell, a plant cell and a fungal cell.

18. The host cell of claim 17 wherein said fungal cell is selected from the group consisting of Saccharomyces spp., Candida spp., Lipomyces starkey, Yarrowia spp., Kluyveromyces spp., Hansenula spp., Aspergillus spp., Penicillium spp., Neurospora spp., Trichoderma spp. and Pichia spp.

19. The host cell of claim 18 wherein said fungal cell is a yeast cell selected from the group consisting of Saccharomyces spp., Candida spp., Hansenula spp. and Pichia spp.

20. The host cell of claim 19 wherein said yeast cell is Saccharomyces cerevisiae.

21. A plant cell comprising said vector of claim 13, wherein expression of said nucleotide sequence of said vector results in production of a polyunsaturated fatty acid by said plant cell.

22. The plant cell of claim 21 wherein said polyunsaturated fatty acid is selected from the group consisting of dihom-γ

-linolenic acid (DGLA), 20;

4n-3, adrenic acid (ADA) and ω

3-docosapentaenoic acid.