Pathway engineering of Escherichia coli for α‐ketoglutaric acid production

Abstract α‐Ketoglutaric acid (α‐KG) is a multifunctional dicarboxylic acid in the tricarboxylic acid (TCA) cycle, but microbial engineering for α‐KG production is not economically efficient, due to the intrinsic inefficiency of its biosynthetic pathway. In this study, pathway engineering was used to improve pathway efficiency for α‐KG production in Escherichia coli . First, the TCA cycle was rewired for α‐KG production starting from pyruvate, and the engineered strain E. coli W3110Δ4‐PCAI produced 15.66 g/L α‐KG. Then, the rewired TCA cycle was optimized by designing various strengths of pyruvate carboxylase and isocitrate dehydrogenase expression cassettes, resulting in a large increase in α‐KG production (24.66 g/L). Furthermore, acetyl coenzyme A (acetyl‐CoA) availability was improved by overexpressing acetyl‐CoA synthetase, leading to α‐KG production up to 28.54 g/L. Finally, the engineered strain E. coli W3110Δ4‐P (H) CAI (H) A was able to produce 32.20 g/L α‐KG in a 5‐L fed‐batch bioreactor. This strategy described here paves the way to the development of an efficient pathway for microbial production of α‐KG.