Identification and Characterization of Diacylglycerol Acyltransferase in Oleaginous Yeast Rhodosporidium toruloides

Abstract

Diacylglycerol acyltransferase (DGAT), which catalyzes TAG formation from DAG and acyl-CoA, has been considered to play a vital role in TAG accumulation in oleaginous microorganisms. The genome of oleaginous yeast Rhodosporidium toruloides contains two putative DGAT genes, RtDGATa and RtDGATb, which shared little conserved amino acid coding sequence with each other. Phylogeny tree analysis showed that RtDGATa belonged to DGAT1 family and RtDGATb belonged to DGAT2 family. For functional identification of the DGATs, RtDGATa and RtDGATb were individually expressed in Saccharomyces cerevisiae TAG-deficient quadruple mutant (H1246). RtDGATa had obvious preference for monounsaturated fatty acids, however the expression of RtDGATa did not alter the TAG content in S. cerevisiae H1246 and it had non-involvement in TAG accumulation according to its mRNA expression level in R. toruloides. The expression of RtDGATb could completely resume TAG biosynthesis in S. cerevisiae H1246. Substrate preference experiments revealed that RtDGATb preferred unsaturated fatty acids over saturated fatty acids, but not C18:3. Only the expression pattern of RtDGATb was related to the process of fatty acid biosynthesis, suggesting that RtDGATb plays an important role in lipid accumulation in R. toruloides.