To engineer acetogen biocatalyst selectively overproducing ethanol from synthesis gas or CO2/H2 as the only liquid carbonaceous product.Methods and Results
Ethanol-resistant mutant originally capable of producing only acetate from CO2/CO was engineered to eliminate acetate production and spore formation using our proprietary Cre-lox66/lox71-system. Bi-functional aldehyde/alcohol dehydrogenase was inserted into the chromosome of the engineered mutant using Tn7-based approach. Recombinants with three or six copies of the inserted gene produced 525 mmol l−1 and 1018 mmol l−1 of ethanol, respectively, in five independent single-step fermentation runs 25 days each (P < 0·005) in five independent repeats using syngas blend 60% CO and 40% H2. Ethanol production was 64% if only CO2 + H2 blend was used compared with syngas blend (P < 0·005).Conclusions
Elimination of genes unnecessary for syngas fermentation can boost artificial integrated pathway performance.Significance and Impact of the Study
Cell energy released via elimination of phosphotransacetylase, acetate kinase and early-stage sporulation genes boosted ethanol production. Deletion of sporulation genes added theft-proof feature to the engineered biocatalyst. Production of ethanol from CO2/H2 blend might be utilized as a tool to mitigate global warming proportional to CO2 fermentation scale.