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Lipid production in Yarrowia lipolytica is maximized by engineering cytosolic redox metabolism

Abstract

Microbial factories have been engineered to produce lipids from carbohydrate feedstocks for production of biofuels and oleochemicals. However, even the best yields obtained to date are insufficient for commercial lipid production. To maximize the capture of electrons generated from substrate catabolism and thus increase substrate-to-product yields, we engineered 13 strains of Yarrowia lipolytica with synthetic pathways converting glycolytic NADH into the lipid biosynthetic precursors NADPH or acetyl-CoA. A quantitative model was established and identified the yield of the lipid pathway as a crucial determinant of overall process yield. The best engineered strain achieved a productivity of 1.2 g/L/h and a process yield of 0.27 g–fatty acid methyl esters/g-glucose, which constitutes a 25% improvement over previously engineered yeast strains. Oxygen requirements of our highest producer were reduced owing to decreased NADH oxidization by aerobic respiration. We show that redox engineering could enable commercialization of microbial carbohydrate-based lipid production.

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Figure 1: Improving the lipid yields of engineered Y. lipolytica by introducing synthetic pathways that recycles NADH.c to NADPH.c or acetyl-CoA.
Figure 2: Yield model-directed optimization of strain engineering and the resultant fermentation profiles of engineered Y. lipolytica contains combination of synthetic pathways.

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Acknowledgements

This work is supported by the Department of Energy (grant DE-SC0008744). Some material from this paper has been included in a proposal submitted to the Department of Energy for a Bioenergy Research Center. The authors would like to acknowledge J. Shaw from Novogy, Inc. and H. Zhou, Woo-suk Ahn and A.Silverman from Massachusetts Institute of Technology for useful discussions and review of the manuscript.

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Contributions

K.Q. and G.S. conceived the project and wrote the manuscript. K.Q., T.M.W., K.Z. and P. X. designed and performed all the experiments. K.Q., T.M.W., K.Z., P.X. and G.S. analyzed the results.

Corresponding author

Correspondence to Gregory Stephanopoulos.

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The authors have filed a patent (US Provisional Application No.: 62/243,824) on the process yield optimization methods.

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Supplementary Figures 1–13, Supplementary Tables 1–7, and Supplementary Notes 1–3 (PDF 3548 kb)

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Qiao, K., Wasylenko, T., Zhou, K. et al. Lipid production in Yarrowia lipolytica is maximized by engineering cytosolic redox metabolism. Nat Biotechnol 35, 173–177 (2017). https://doi.org/10.1038/nbt.3763

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