Production of free fatty acids (FFAs) and derivatives from renewable non-food biomass by microbial fermentation is of great interest. Here, we report the development of engineered Rhodococcus opacus strains producing FFAs, fatty acid ethyl esters (FAEEs) and long-chain hydrocarbons (LCHCs). Culture conditions were optimized to produce 82.9 g l of triacylglycerols from glucose, and an engineered strain with acyl-coenzyme A (CoA) synthetases deleted, overexpressing three lipases with lipase-specific foldase produced 50.2 g l of FFAs. Another engineered strain with acyl-CoA dehydrogenases deleted, overexpressing lipases, foldase, acyl-CoA synthetase and heterologous aldehyde/alcohol dehydrogenase and wax ester synthase produced 21.3 g l of FAEEs. A third engineered strain with acyl-CoA dehydrogenases and alkane-1 monooxygenase deleted, overexpressing lipases, foldase, acyl-CoA synthetase and heterologous acyl-CoA reductase, acyl-ACP reductase and aldehyde deformylating oxygenase produced 5.2 g l of LCHCs. Metabolic engineering strategies and engineered strains developed here may help establish oleaginous biorefinery platforms for the sustainable production of chemicals and fuels.
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