Synergistic Metabolism of Glucose and Formate Increases the Yield of Short-Chain Organic Acids in Escherichia coli

Microbial cell factories using a single carbon source (e.g., sugars) have been used to produce a wide variety of chemicals. However, this process is often accompanied by stoichiometric constraints on carbons and redox cofactors. Here, a synthetic pathway was designed and constructed in Escherichia coli to synergistically use glucose and formate as mixed carbon sources. By optimizing this synthetic pathway via enzyme mining, protein engineering, and bioprocess approaches, the yield of pyruvate from glucose was enhanced to 94% of the theoretical glycolysis yield, reaching 1.88 mol/mol. Finally, the optimized synthetic pathway was integrated with a phosphite reductase-based NADH regeneration system in malate-producing E. coli, resulting in the conversion of glucose into l -malate with a high yield of up to 1.65 mol/mol. This synergistic carbon metabolism strategy can be used to establish carbon- and energy-efficient productive processes.