Medium-chain triglyceride production in Nannochloropsis via a fatty acid chain length discriminating mechanism

Depending on their fatty acid (FA) chain length, triacylglycerols (TAGs) have distinct applications; thus, a feedstock with a genetically designed chain length is desirable to maximize process efficiency and product versatility. Here, ex vivo, in vitro, and in vivo profiling of the large set of type-2 diacylglycerol acyltransferases (NoDGAT2s) in the industrial oleaginous microalga Nannochloropsis oceanica revealed two endoplasmic reticulum-localized enzymes that can assemble medium-chain FAs (MCFAs) with 8-12 carbons into TAGs. Specifically, NoDGAT2D serves as a generalist that assembles C8-C18 FAs into TAG, whereas NoDGAT2H is a specialist that incorporates only MCFAs into TAG. Based on such specialization, stacking of NoDGAT2D with MCFA- or diacylglycerol-supplying enzymes or regulators, including rationally engineering Cuphea palustris acyl carrier protein thioesterase, Cocos nucifera lysophosphatidic acid acyltransferase, and Arabidopsis thaliana WRINKLED1, elevated the medium-chain triacylglycerol (MCT) share in total TAG 66-fold and MCT productivity 64.8-fold at the peak phase of oil production. Such functional specialization of NoDGAT2s in the chain length of substrates and products reveals a dimension of control in the cellular TAG profile, which can be exploited for producing designer oils in microalgae. In Nannochloropsis oceanica, two chain-length discriminating diacylglycerol acyltransferases assemble medium-chain fatty acids into triacylglycerols.

Automated summary

Depending on their chain length, triacylglycerols (TAGs) have different applications, so it is desirable to have a genetically designed feedstock with specific chain length for maximizing process efficiency and product versatility. In the microalga Nannochloropsis oceanica, two enzymes were found to assemble medium-chain fatty acids (MCFAs) into TAGs, with one being a generalist and the other a specialist. By manipulating these enzymes, the production of medium-chain triacylglycerols (MCTs) in TAGs can be significantly increased in microalgae, allowing for the production of designer oils.