Heterologous Expression of the carbon monoxide dehydrogenase Gene from Clostridium sp. to Enhance Acetic Acid and Alcohol Production from CO₂

This study evaluates the performance of carbon monoxide dehydrogenase ( codh ) embedded strains in bench-scale microbial electrochemical systems (MES) for CO₂ reduction to biofuels and biochemicals. CO ₂ fermentation efficiency was evaluated by comparing the wild-type Clostridium acetobutylicum (Wild), a negative control E. coli strain lacking the codh gene (NC-BL21), and engineered E. coli strain (Eng) alone and with IPTG induction (Eng+IPTG). Four electrochemical systems were used viz. Wild+E, NC-BL21+E, Eng+E, and Eng+IPTG+E, with a poised potential of 0.6 V applied to the working electrode. CO₂ and bicarbonate were supplemented to a total inorganic carbon (IC) concentration of 40 g/L, with a retention time of 60 h. The engineered strains demonstrated enhanced metabolic performance compared to the wild-type and negative control strains, yielding a maximum formic acid concentration of 2.1 g/L and acetic acid concentration of 7.8 g/L under the Eng+IPTG condition. Ethanol yield was highest at 3.9 g/L under the Eng+IPTG+E condition, substantially exceeding the 2.4 g/L acetic acid yield observed in the wild-type strain. The engineered strains showed superior cumulative yields (0.4075 g/g), improved codh charge flux stability (60 vs. 5 for Wild), and upregulated expression of genes in the Wood-Ljungdahl pathway. Bioelectrochemical performance analysis demonstrated elevated reductive catalytic currents, enhanced CO₂ reduction, and optimal charge transfer kinetics. This study highlights the effectiveness of genetic and process engineering, particularly codh overexpression and IPTG induction, in optimizing microbial electrosynthesis for biofuel and biochemical production from C1 gases.