Mortierella alpina diacylglycerol acyltransferase for alteration of polyunsaturated fatty acids and oil content in oleaginous organisms

• US 7,198,937 B2
• Filed: 12/29/2004
• Issued: 04/03/2007
• Est. Priority Date: 11/04/2004
• Status: Active Grant

• Current Assignee: Dupont US Holding, LLC (International Flavors & Fragrances Incorporated)
• Original Assignee: E. I. du Pont de Nemours and Company (Corteva, Inc.)
• Inventors: Xue, Zhixiong, Yadav, Narendra S., Macool, Daniel Joseph
• Primary Examiner(s): Prouty; Rebecca E.
• Assistant Examiner(s): CHOWDHURY, IQBAL HOSSAIN
• Application Number:
  US11/024,545
• Publication Number:
  US 20060094087A1
• Time in Patent Office: 825 Days
• Field of Search: 435/22.3, 435/252.3, 435/193
• US Class Current: 435/254.2
• CPC Class Codes: C12N 9/1029 transferring groups other than amino-acyl groups (2.3.1)
• CPC Class Codes: C12P 7/6463 obtained from glyceride producing microorganisms, e.g. single cell oil
• CPC Class Codes: C12P 7/6472 containing polyunsaturated fatty acid [PUFA] residues, i.e. having two or more double bonds in their backbone

Abstract

An acyltransferase is provided, suitable for use in the manufacture of microbial oils enriched in omega fatty acids in oleaginous organisms. Specifically, the gene encoding diacylglycerol acyltransferase (DGAT2) has been isolated from Mortierella alpina. This gene encodes an enzyme that participates in the terminal step in oil biosynthesis in fungi and yeast and is expected to play a key role in altering the quantity of long-chain polyunsaturated fatty acids produced in oils of oleaginous organisms. Most desirably, the substrate specificity of the instant DGAT2 will be particularly useful to enable accumulation of long-chain PUFAs having chain lengths equal to or greater than C₂₀ in oleaginous yeast, such as Yarrowia lipolytica.

Claims

1. An isolated nucleic acid molecule selected from the group consisting of:

(a) an isolated nucleic acid molecule encoding a diacylglycerol acyltransferase 2 enzyme, having the amino acid sequence as set forth in SEQ ID NO;

2;

(b) an isolated nucleic acid molecule encoding an diacylglycerol acyltransferase 2 enzyme, that hybridizes with (a) under the following hybridization conditions;

0.1×

SSC, 0.1% SDS, 65°

C. and washed with 2×

SSC, 0.1% SDS followed by 0.1×

SSC, 0.1% SDS;

or

(c) an isolated nucleic acid molecule that is completely complementary to (a) or (b).

2. The isolated nucleic acid molecule of claim 1 as set forth in SEQ ID NO:

1.

3. A chimeric gene comprising the isolated nucleic acid molecule of claim 1 operably linked to suitable regulatory sequences.

4. An isolated host cell comprising the chimeric gene of claim 3.

5. An isolated host cell of claim 4, selected from the group consisting of algae, bacteria, molds, fungi and yeast.

6. An isolated host cell of claim 5, wherein the yeast is an oleaginous yeast.

7. An isolated host cell of claim 6, wherein the oleaginous yeast cell is selected from the group consisting of Yarrowia, Candida, Rhodotorula, Rhodosporidlum, Cryptococcus, Trichosporon and Lipomyces.

8. An isolated host cell of claim 7, wherein the host cell is Yarrowia lipolyzica.