Synthesis of high-titer alka(e)nes in Yarrowia lipolytica is enabled by a discovered mechanism

Jingbo Li,Bekir E Eser,Peter Ruhdal Jensen,Nian Liu,Gregory Stephanopoulos,Zheng Guo,Yongshuo Ma

doi:10.1038/s41467-020-19995-0

Abstract

Alka(e)nes are ideal fuel components for aviation, long-distance transport, and shipping. They are typically derived from fossil fuels and accounting for 24% of difficult-to-eliminate greenhouse gas emissions. The synthesis of alka(e)nes in Yarrowia lipolytica from CO2-neutral feedstocks represents an attractive alternative. Here we report that the high-titer synthesis of alka(e)nes in Yarrowia lipolytica harboring a fatty acid photodecarboxylase (CvFAP) is enabled by a discovered pathway. We find that acyl-CoAs, rather than free fatty acids (FFAs), are the preferred substrate for CvFAP. This finding allows us to debottleneck the pathway and optimize fermentation conditions so that we are able to redirect 89% of acyl-CoAs from the synthesis of neutral lipids to alka(e)nes and reach titers of 1.47 g/L from glucose. Two other CO2-derived substrates, wheat straw and acetate, are also demonstrated to be effective in producing alka(e)nes. Overall, our technology could advance net-zero emissions by providing CO2-neutral and energy-dense liquid biofuels.

Highlights

Alka(e)nes are ideal fuel components for aviation, long-distance transport, and shipping
Expression of the codon-optimized CvFAP in Y. lipolytica po1f resulted in YLjbl-2 which produced 15.3 mg/L alka (e)nes
Since free fatty acids (FFAs) were believed to be the substrate of CvFAP in in vitro studies[11,15], harnessing FFA-producing pathways and blocking FFA-consuming pathways in YLjbl-2 were expected to improve alka(e)ne synthesis

Summary

Introduction

Alka(e)nes are ideal fuel components for aviation, long-distance transport, and shipping. 1234567890():,; Increasing greenhouse gas (GHG) emissions is the leading cause of climate change[1] and efforts to minimize the use of fossil fuels have mainly focused on deploying alternative energy sources such as bioethanol and electrical power Due to their low energy densities, both of these approaches struggle to transform the aviation, trucking, and shipping industries, making long-distance transport difficult to decarbonize[2]. For fatty acid-derived hydrocarbons, two major metabolic pathways have been identified, the two-step cyanobacterial pathway involving acyl-ACP reduction and aldehyde decarbonylation[7], and the one-step decarboxylation reaction from free fatty acids (FFAs)[8,9,10,11] In prior studies, both pathways were harnessed in E. coli to synthesize alka(e)nes with varying chain lengths, with reported titers ranging from 1.3 to 1310 mg/L and corresponding yields ranging from 0.43 to 11 mg/g glucose (Supplementary Table 1). The production of 10.87 mg/L and 58.7 mg/L alka(e)nes were achieved in the presence of light with batch and fed-batch fermentation modes, respectively, no information regarding the responsible metabolic pathways was detailed[14]

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Results

Conclusion