Oleic acid production in yeast
Disclosed are transformed cells comprising one or more genetic modifications that affect the lipid content of the cell, e.g., by increasing the concentration of oleic acid in the cell relative to an unmodified cell of the same type. Also disclosed are methods for modifying the lipid content of a cell by increasing the activity of one or more proteins in the cell and/or by decreasing the activity of one or more proteins in the same cell.
1. A yeast cell comprising:
a first genetic modification, wherein the first genetic modification is a knockout mutation of a native Δ12 desaturase protein;
a second genetic modification, wherein the second genetic modification increases the expression of an elongase protein, a diacylglycerol acyltransferase protein, or a glycerol-3-phosphate acyltransferase protein; and
a third genetic modification, wherein the third genetic modification results from a transformation with a nucleic acid comprising a gene encoding an exogenous fungal Δ9 desaturase protein.
2. The yeast cell of claim 1 , wherein the second genetic modification results from a transformation with a nucleic acid comprising a gene encoding an elongase protein, a diacylglycerol acyltransferase protein, or a glycerol-3-phosphate acyltransferase protein.
3. The yeast cell of claim 1 , wherein:
the exogenous Δ9 desaturase protein is a Puccinia graminis, Arxula adeninivorans or Microbotryum violaceum Δ9 desaturase protein.
4. The yeast cell of claim 1 , wherein the yeast cell is selected from the group consisting of Arxula, Aspergillus, Aurantiochytrium, Candida, Claviceps, Cryptococcus, Cunninghamella, Geotrichum, Hansenula, Kluyveromyces, Kodamaea, Leucosporidiella, Lipomyces, Mortierella, Ogataea, Pichia, Prototheca, Rhizopus, Rhodosporidium, Rhodotorula, Saccharomyces, Schizosaccharomyces, Tremella, Trichosporon, Wickerhamomyces , and Yarrowia.
5. The yeast cell of claim 1 , wherein the yeast cell is selected from the group consisting of Arxula adeninivorans, Saccharomyces cerevisiae , and Yarrowia lipolytica.
6. The yeast cell of claim 1 , wherein:
the yeast cell is Yarrowia lipolytica ; and the native Δ12 desaturase protein has the amino acid sequence set forth in SEQ ID NO: 1.
7. The yeast cell of claim 1 , wherein the yeast cell comprises at least 50% lipid as measured by % dry cell weight.
8. The yeast cell of claim 1 , wherein the yeast cell comprises oleic acid at a concentration of at least 70% as a percentage of total C16 and C18 fatty acids in the yeast cell.
9. The yeast cell of claim 1 , wherein the second genetic modification increases the expression of a glycerol-3-phosphate acyltransferase protein.
10. The yeast cell of claim 2 , wherein the glycerol-3-phosphate acyltransferase protein is an exogenous glycerol-3-phosphate acyltransferase protein.
11. The yeast cell of claim 1 , wherein the second genetic modification increases the expression of an elongase protein.
12. The yeast cell of claim 1 , wherein the second genetic modification increases the expression of a diacylglycerol acyltransferase protein.
13. The yeast cell of claim 1 , wherein the yeast cell comprises a knockout mutation of a native Δ9 desaturase protein.
14. The yeast cell of claim 1 , wherein the exogenous Δ9 desaturase protein has an increased specificity for C18 saturated fatty acids relative to a native Δ9 desaturase protein of the yeast cell.
15. The yeast cell of claim 10 , wherein the exogenous glycerol-3-phosphate acyltransferase protein is an Arxula adeninivorans glycerol-3-phosphate acyltransferase.
16. The yeast cell of claim 2 , wherein the elongase protein is an exogenous elongase protein.
17. The yeast cell of claim 16 , wherein the exogenous elongase protein is a Rattus norvegicus or Yarrowia lipolytica elongase.
18. The yeast cell of claim 2 , wherein the diacylglycerol acyltransferase protein is an exogenous diacylglycerol acyltransferase protein.
19. The yeast cell of claim 18 , wherein the exogenous diacylglycerol acyltransferase protein is a Rhodosporidium toruloides, Yarrowia lipolytica, Arxula adeninivorans , or Claviceps purpurea diacylglycerol acyltransferase.
20. The yeast cell of claim 1 , wherein the second genetic modification results from transformation with a nucleic acid comprising a gene encoding a diacylglycerol acyltransferase type 1 protein, a gene encoding a diacylglycerol acyltransferase type 2 protein, or a gene encoding a diacylglycerol acyltransferase type 1 protein and a gene encoding a diacylglycerol acyltransferase type 2 protein.
21. The yeast cell of claim 1 , wherein the second genetic modification results from a transformation with one or more nucleic acids comprising a gene encoding an elongase protein, a diacylglycerol acyltransferase protein, and a glycerol-3-phosphate acyltransferase protein.
22. The yeast cell of claim 21 , wherein the elongase protein is a Rattus norvegicus or Yarrowia lipolytica elongase, the diacylglycerol acyltransferase protein is a Rhodosporidium toruloides, Yarrowia lipolytica, Arxula adeninivorans and/or Claviceps purpurea diacylglycerol acyltransferase, and the glycerol-3-phosphate acyltransferase protein is an Arxula adeninivorans glycerol-3-phosphate acyltransferase.