Increased incorporation of gaseous CO2 into succinate by Escherichia coli overexpressing carbonic anhydrase and phosphoenolpyruvate carboxylase genes

- E. coli was engineered to increase incorporation of gaseous CO2 to succinate.
- Expression of carbonic anhydrase and phosphoenolpyruvate carboxylase genes enhanced the incorporation of gaseous CO2 to succinate in E. coli.
- Pyruvate kinase gene was deleted in E. coli for enhancing the incorporation of gaseous CO2 to succinate.

Carbon dioxide (CO2) is an abundant and cheap carbon source that is partly responsible for global warming in the atmosphere. The objective of this study was to construct a recombinant E. coli strain that can show enhanced production of succinate derived from CO2. In this study, we confirmed the enhancement of utilization by analyzing succinate containing one carbon-13 (13C) derived from 13CO2. Firstly, the carbonic anhydrase gene (SP(−)HCCA) derived from Hahella chejuensis KCTC 2396 was over-expressed to enhance carbon flux toward bicarbonate ion (HCO3−) synthesis in E. coli. The phosphoenolpyruvate carboxylase gene (ppc) was over-expressed to enhance the production of oxaloacetate by enhancing the carbon flux. Compared with the control strain, the percentage of the succinate containing one 13C (succinate119) to total succinate was enhanced by approximately 2.80-fold and the amount of succinate119 also increased by approximately 4.09-fold in SGJS120. Secondly, the lactate dehydrogenase gene (ldhA) was deleted to re-direct the utilization of the carbon source from glucose to enhance succinate production in SGJS120. However, ldhA deletion did not increase CO2 utilization in SJGS120. Finally, the phosphotransferase system gene (ptsG) and pyruvate kinase F gene (pykF) were deleted to increase the amount of phosphoenolpyruvate (PEP). SGJS126 (pykF deletion strain) showed the highest increase, which was 6.05-fold higher than the control strain. From the results, SP(−)HCCA overexpression and pykF deletion may be useful for enhancing CO2 utilization in E. coli. Additionally, engineered strains showed the potential to reduce the cost of succinate production by using an industrially cheaper carbon source such as CO2 and converting CO2 to a valuable chemical.