Optimized strains of
View Patent ↗Described are engineered strains of the oleaginous yeast Yarrowia lipolytica capable of producing an oil comprising greater than 50 weight percent of eicosapentaenoic acid [“EPA”], an ω-3 polyunsaturated fatty acid, measured as a weight percent of total fatty acids [“% TFAs”] and having a ratio of at least 3.1 of EPA % TFAs, to linoleic acid, measured as % TFAs. These strains over-express at least one Δ9 elongase/Δ8 desaturase multizyme, in addition to other heterologous Δ9 elongases, Δ8 desaturases, Δ5 desaturases, Δ17 desaturases, Δ12 desaturases, C 16/18 elongases, and optionally over-express diacylglycerol cholinephosphotransferases, malonyl CoA synthetases and/or acyl-CoA lysophospholipid acyltransferases. The expression of at least one peroxisome biogenesis factor protein is down-regulated. Methods for producing EPA within said host cells, oils obtained from the cells, and products therefrom are claimed.
1. A recombinant Yarrowia host cell that produces an oil comprising:
(a) at least 50 weight percent of eicosapentaenoic acid measured as a weight percent of total fatty acids; and
(b) a ratio of at least 3.1 of eicosapentaenoic acid, measured as a weight percent of total fatty acids, to linoleic acid, measured as a weight percent of total fatty acids,
wherein the Yarrowia host cell comprises:
(i) at least one multizyme comprising a polypeptide having at least one delta-9 elongase linked to at least one delta-8 desaturase;
(ii) at least one peroxisome biogenesis factor protein whose expression has been down-regulated; and
(iii) at least one recombinant construct comprising at least one nucleotide sequence encoding an acyl-CoA lysophospholipid acyltransferase consisting essentially of the amino acid sequence of SEQ ID NO:17.
2. The recombinant host cell of claim 1 , wherein the delta-9 elongase and delta-8 desaturase of the multizyme are linked by a linker selected from the group consisting of: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7.
3. The recombinant host cell of claim 1 , wherein the multizyme consists essentially of the sequence selected from the group consisting of: SEQ ID NO:9, SEQ ID NO:11 and SEQ ID NO:13.
4. A method for making a microbial oil comprising eicosapentaenoic acid comprising:
a) culturing the recombinant host cell of any of claims 1 or 2 , wherein a microbial oil comprising eicosapentaenoic acid is produced; and
b) optionally recovering the microbial oil of step (a).
5. A recombinant host cell for the production of an oil comprising at least 50 weight percent of eicosapentaenoic acid measured as a weight percent of total fatty acids, said host cell is selected from the group consisting of: Yarrowia lipolytica Y8406 having the designation ATCC TM PTA-10025; Yarrowia lipolytica Y8412 having the designation ATCC TM PTA-10026; and Yarrowia lipolytica Y8259 having the designation ATCC TM PTA-10027 wherein the host cell comprises at least one recombinant construct comprising at least one nucleotide sequence encoding an acyl-CoA lysophospholipid acyltransferase consisting essentially of the amino acid sequence of SEQ ID NO:17.
6. The recombinant host cell of claim 1 , wherein said ratio of eicosapentaenoic acid to linoleic acid in the oil is at least 3.5.
7. The recombinant host cell of claim 1 , wherein said ratio of eicosapentaenoic acid to linoleic acid in the oil is at least 4.5.
8. The recombinant host cell of claim 6 , wherein the oil comprises at least 55 percent of eicosapentaenoic acid measured as a weight percent of total fatty acids.
9. The recombinant host cell of claim 7 , wherein the oil comprises at least 55 percent of eicosapentaenoic acid measured as a weight percent of total fatty acids.
10. The recombinant host cell of claim 8 , wherein the oil comprises at least 60 percent of eicosapentaenoic acid measured as a weight percent of total fatty acids.
11. The recombinant host cell of claim 9 , wherein the oil comprises at least 60 percent of eicosapentaenoic acid measured as a weight percent of total fatty acids.
12. The recombinant host cell of claim 1 , wherein the oil comprises at least 60 percent of eicosapentaenoic acid measured as a weight percent of total fatty acids.