Introducing Combinatorial Metabolic Engineering to Improve Free Fatty Acids Production

Abstract

Microbial production of fatty-acid-derived chemicals has been studied due to its promising substitute for some petroleum-based chemicals. Many studies have attempted to engineer microbes for increased production of FFA through metabolic engineering and synthetic biology. Here, we constructed a high FFA-producing Escherichia coli (E. coli) strain by combinatorial combinations of previous positive results; 1) heterologous expression of the methylmalonyl-CoA carboxyltransferase (MMC) and overexpression of phosphoenolpyruvate carboxylase (PPC) to increase malonyl-CoA pool from oxaloacetate (OAA), 2) expressing a mutant acyl-CoA thioesterase I (‘TesA) with high specific enzyme activity, and 3) deletion of FFA transport-related genes. The manipulation of E. coli increased the FFA production by 5.4-fold compared with control strain. Additionally, employing transcriptional activator of fatty acid synthesis (FadR) further increased the TRY in our strain, resulting in 9.5- and 1.4-fold increases in FFA production, compared with the control strain expressing only the wild-type thioesterase and the positive control strain overexpressing FadR, respectively.