Increasing cocoa butter-like lipid production of Saccharomyces cerevisiae by expression of selected cocoa genes

Cocoa butter (CB) extracted from cocoa beans mainly consists of three different kinds of triacylglycerols (TAGs), 1,3-dipalmitoyl-2-oleoyl-glycerol (POP, C16:0-C18:1-C16:0), 1-palmitoyl-3-stearoyl-2-oleoyl-glycerol (POS, C16:0-C18:1-C18:0) and 1,3-distearoyl-2-oleoyl-glycerol (SOS, C18:0-C18:1-C18:0), but CB supply is limited. Therefore, CB-like lipids (CBL, which are composed of POP, POS and SOS) are in great demand. Saccharomyces cerevisiae produces TAGs as storage lipids, which are also mainly composed of C16 and C18 fatty acids. However, POP, POS and SOS are not among the major TAG forms in yeast. TAG synthesis is mainly catalyzed by three enzymes: glycerol-3-phosphate acyltransferase (GPAT), lysophospholipid acyltransferase (LPAT) and diacylglycerol acyltransferase (DGAT). In order to produce CBL in S. cerevisiae, we selected six cocoa genes encoding GPAT, LPAT and DGAT potentially responsible for CB biosynthesis from the cocoa genome using a phylogenetic analysis approach. By expressing the selected cocoa genes in S. cerevisiae, we successfully increased total fatty acid production, TAG production and CBL production in some S. cerevisiae strains. The relative CBL content in three yeast strains harboring cocoa genes increased 190, 230 and 196% over the control strain, respectively; especially, the potential SOS content of the three yeast strains increased 254, 476 and 354% over the control strain. Moreover, one of the three yeast strains had a 2.25-fold increased TAG content and 6.7-fold higher level of CBL compared with the control strain. In summary, CBL production by S. cerevisiae were increased through expressing selected cocoa genes potentially involved in CB biosynthesis.