A team at S. Korea’s KAIST has engineered an oleaginous bacterium, Rhodococcus opacus, to produce fatty acids (FFAs), fatty acid ethyl esters (FAEEs) and long-chain hydrocarbons (LCHCs). A paper on their work is published in the journal Nature Chemical Biology.

Culture conditions were optimized to produce 82.9 g l⁻¹ 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⁻¹ 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⁻¹ 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⁻¹ of LCHCs.

Metabolic engineering strategies and engineered strains developed here may help establish oleaginous biorefinery platforms for the sustainable production of chemicals and fuels.

—Kim et al.

Metabolic engineering for the production of free fatty acids (FFAs), fatty acid ethyl esters (FAEEs), and long-chain hydrocarbons (LCHCs) in Rhodococcus opacus PD630.

The newly developed strain, created by Distinguished Professor Sang Yup Lee and his team, showed the highest efficiency in producing fatty acids and biodiesels ever reported.

Professor Lee’s team has already engineered Escherichia coli to produce short-chain hydrocarbons, which can be used as gasoline (published in Nature as the cover paper in 2013). However, the production efficiency of the short-chain hydrocarbons using E. coli (0.58 g/L) fell short of the levels required for commercialization.

To overcome these issues, the team employed oil-accumulating Rhodococcus opacus as a host strain in this study. First, the team optimized the cultivation conditions of Rhodococcus opacus to maximize the accumulation of oil (triacylglycerol), which serves as a precursor for the biosynthesis of fatty acids and their derivatives. Then, they systematically analyzed the metabolism of the strain and redesigned it to enable higher levels of fatty acids and two kinds of fatty acid-derived biodiesels (fatty acid ethyl esters and long-chain hydrocarbons) to be produced.

This work was supported by the Technology Development Program to Solve Climate Changes on Systems Metabolic Engineering for Biorefineries from the Ministry of Science and ICT through the National Research Foundation (NRF) of Korea (NRF-2012M1A2A2026556 and NRF-2012M1A2A2026557).

Resources

• Hye Mi Kim, Tong Un Chae, So Young Choi, Won Jun Kim & Sang Yup Lee Nature Chemical Biology (2019) “Engineering of an oleaginous bacterium for the production of fatty acids and fuels” Nature Chemical Biology doi: 10.1038/s41589-019-0295-5