Heterologous production of 10-methylstearic acid
Nucleic acids and cells comprising a methyltransferase gene and/or a reductase gene are disclosed. These nucleic acids and cells may be used to produce branched (methyl)lipids, such as 10-methylstearate.
1. A yeast cell comprising a methyltransferase gene encoding a Thermomonospora curvata enzyme tmsB and either a branched (methyl)lipid or an exomethylene-substituted lipid, wherein:
the branched (methyl)lipid or exomethylene-substituted lipid is a carboxylic acid, carboxylate, ester, thioester, or amide, and
the branched (methyl)lipid comprises a saturated or unsaturated branched aliphatic chain comprising a branching methyl group or the exomethylene-substituted lipid comprises a branched aliphatic chain that is branched because the aliphatic chain is substituted with an exomethylene group.
2. The cell of claim 1 , wherein the branched (methyl)lipid or the exomethylene-substituted lipid comprises a linear lipid with a chain length of 14-20 carbons and a methyl branch at the Δ9, Δ10, or Δ11 position.
3. The cell of claim 2 , wherein the branched (methyl)lipid or the exomethylene-substituted lipid is a diacylglycerol, triacylglycerol, or phospholipid, and the diacylglycerol, triacylglycerol, or phospholipid comprises an ester of 10-methylstearate or an ester of 10-methylenestearate.
4. The cell of claim 1 , wherein at least 1% by weight of the fatty acids of the cell are one or more linear fatty acids with a chain length of 14-20 carbons and a methyl branch at the Δ9, Δ10, or Δ11 position.
5. The cell of claim 1 , wherein the cell comprises at least 1% lipid as measured by % dry cell weight.
6. The cell of claim 1 , further comprising a recombinant reductase gene.
7. The cell of claim 6 , wherein the reductase gene encodes tmsA from Thermomonospora curvata.
8. The cell of claim 1 , wherein:
the methyltransferase gene is codon-optimized for the cell; or
the cell further comprises a reductase gene and the reductase gene is codon-optimized for the cell.
9. The cell of claim 1 , wherein the cell is selected from the group consisting of Arxula, Saccharomyces , and Yarrowia.
10. The cell of claim 9 , wherein the cell is selected from the group consisting of Arxula adeninivorans, Saccharomyces cerevisiae , and Yarrowia lipolytica.
11. The cell of claim 1 , wherein the cell comprises a methyltransferase protein encoded by the methyltransferase gene, and the methyltransferase protein comprises either: an amino acid sequence with at least 95% sequence identity with the amino acid sequence of SEQ ID NO:76; or the amino acid sequence of SEQ ID NO:76.
12. The cell of claim 1 , wherein the methyltransferase gene comprises either: a nucleotide sequence with at least 95% sequence identity with the nucleotide sequence of SEQ ID NO:75; or the nucleotide sequence of SEQ ID NO:75.
13. A method of producing a branched (methyl)lipid or exomethylene-substituted lipid, comprising contacting the cell of claim 1 with oleic acid, methionine, or both oleic acid and methionine.
14. The cell of claim 6 , wherein the reductase protein is in a fusion protein with the tmsB enzyme.