Investigating E. coli Coculture for Resveratrol Production with ¹³C Metabolic Flux Analysis

Resveratrol, a phytoalexin produced by plants, has several beneficial effects in humans. It can be produced using Escherichia coli by introducing only three heterologous genes: TAL, 4CL, and STS. However, the resveratrol synthesis pathway requires two precursors, tyrosine and acetyl-CoA, which are produced by two branched central metabolic pathways. Therefore, overexpression of these genes in E. coli results in the production of only trace amounts of resveratrol. In this study, we attempted to produce resveratrol via coculture of two engineered strains in which the two metabolic pathways are activated. The first strain was engineered to produce p-coumaric acid using tyrosine as a precursor, which can be synthesized by the pentose phosphate pathway. The second strain produced resveratrol by combining p-coumaric acid from the first strain and malonyl-CoA synthesized from acetyl-CoA, which is produced by the glycolytic pathway. In total, 55.7 mg/L of resveratrol was produced from 20 g/L of glucose via coculture of these two strains in glucose minimal medium without any supplements. The metabolic fluxes in each of the strains producing resveratrol were successfully investigated by ¹³C metabolic flux analysis. The results showed that the balance between the citric acid cycle and the malonyl-CoA supply node was important for resveratrol production.