Metabolic Engineering for Chemicals Production By Solving Reducing Equivalent Problems

Redox balance is very important to realize efficient chemicals production in metabolic engineering. However, there are many redox imbalance problems during metabolic engineering for chemicals production. For example, the reducing equivalent type generated during anaerobic glucose metabolism is not consistent with that required for isobutanol production. The reducing equivalent quantities generated in the native host are not enough for those required for succinate and terpenoids production. We developed several strategies to solve these reducing equivalent problems. Three strategies were used for isobutanol production, including (1) activation of transhydrogenase PntAB and NAD kinase in combination for converting NADH to NADPH; (2) recruiting NADH-dependent keto acid reductoisomerase together with activating PntAB and NAD kinase; (3) recruiting Entner-Doudoroff pathway to change the reducing equivalent type generated during anaerobic glucose metabolism. Two strategies were used for succinate production, including (1) activation of pentose phosphate pathway to increase the reducing equivalent quantities generated during anaerobic glucose metabolism together with activating transhydrogenase SthA to convert NADPH to NADH for succinate production; (2) activation of pyruvate dehydrogenase under anaerobic condition in the transcriptional and protein levels. Pentose phosphate pathway and TCA cycle were activated to increase the reducing equivalent quantities generated during aerobic glucose metabolism for improving terpenoids production.