(485x) Engineering Native E. Coli Propionyl-CoA and Methylmalonyl-CoA Metabolism to Support Heterologous 6-Deoxyerythronolide B Production

Propionyl-CoA and (2S)-methylmalonyl-CoA are required for the biosynthesis of the erythromycin precursor 6-deoxyerythronolide B (6dEB). A heterologous pathway has been introduced to E. coli to produce these two acyl-CoA's and support 6dEB production. In this study, hypothesis-driven modifications were made to native E. coli metabolism to investigate the potential impact on heterologous 6dEB biosynthesis. Specifically, three native genes, predicted to link native and heterologous metabolic pathways, were separately deleted and over-expressed. These genes were sbm encoding methylmalonyl-CoA mutase, ygfG encoding methylmalonyl-CoA decarboxylase, and ygfH encoding propionyl-CoA:succinyl-CoA transferase. The study showed that the Sbm pathway did not influence heterologous 6dEB production and over-expression of ygfG decreased final 6dEB levels by 4-fold. Interestingly, the deletion of ygfH led to a 2-fold increase in 6dEB production. These results demonstrate that E. coli native metabolism does compete with the heterologous pathway for propionyl-CoA and (2S)-methylmalonyl-CoA. The study also revealed that E. coli metabolism can provide propionyl-CoA and (2S)-methylmalonyl-CoA for heterologous 6dEB biosynthesis in the absence of exogenous propionate or the heterologous pcc genes that catalyze the formation of (2S)-methlmalonyl-CoA. These findings indicate the potential to leverage native E. coli metabolism for future heterologous natural product applications.