Journal ArticleMetabolic engineering strategies to produce medium-chain oleochemicals via acyl-ACP:CoA transacylase activityYan, Qiang; Cordell, William T.; Jindra, Michael A.; Courtney, Dylan K.; Kuckuk, Madeline K.; Chen, Xuanqi; Pfleger, Brian F.<title>Abstract</title> <p>Microbial lipid metabolism is an attractive route for producing oleochemicals. The predominant strategy centers on heterologous thioesterases to synthesize desired chain-length fatty acids. To convert acids to oleochemicals (e.g., fatty alcohols, ketones), the narrowed fatty acid pool needs to be reactivated as coenzyme A thioesters at cost of one ATP per reactivation - an expense that could be saved if the acyl-chain was directly transferred from ACP- to CoA-thioester. Here, we demonstrate such an alternative acyl-transferase strategy by heterologous expression of PhaG, an enzyme first identified in<italic>Pseudomonads</italic>, that transfers 3-hydroxy acyl-chains between acyl-carrier protein and coenzyme A thioester forms for creating polyhydroxyalkanoate monomers. We use it to create a pool of acyl-CoA’s that can be redirected to oleochemical products. Through bioprospecting, mutagenesis, and metabolic engineering, we develop three strains of<italic>Escherichia coli</italic>capable of producing over 1 g/L of medium-chain free fatty acids, fatty alcohols, and methyl ketones.</p>Nature Communications2022-12-0110481151Nature Communications1312041-1723https://doi.org/10.1038/s41467-022-29218-31703504National Science Foundation