Engineered Coenzyme A Biosynthesis and Butyrate Transporter Drives High‐Efficient Butyrate Synthesis in Escherichia coli

Butyric acid is a four-carbon fatty acid with a range of applications in chemical, food and pharmaceutical industries. In this study, a heterologous butyrate biosynthetic pathway was engineered in Escherichia coli, which was afflicted by low titer and low yield. To address these issues, two key strategies for metabolic engineering of E. coli were implemented by enhancing coenzyme A (CoA) biosynthesis and optimizing butyrate transport. First, the CoA biosynthesis pathway was engineered through alleviating CoA-mediated inhibition, and enhancing the supply of pantothenate and cysteine precursors. Second, a TolC-associated MdtEF efflux pumps was identified and optimized to mitigate butyrate reuptake. The combined implementation in strain JH016 led to 11.1-fold and 86% increase of butyrate titer and yield, resulting in production of 21.12 g/L butyrate with a yield of 0.95 mol/mol. Our results suggested that CoA engineering and butyrate transporter optimization had a synergistic effect on butyrate production. Furthermore, these strategies could be broadly utilized for the production of various other useful chemicals in the fields of metabolic engineering and synthetic biology.