Proceedings of the National Academy of Sciences of the United States of America
Thayer School of Engineering
Clostridium thermocellum is a thermophilic, obligately anaerobic, Gram-positive bacterium that is a candidate microorganism for converting cellulosic biomass into ethanol through consolidated bioprocessing. Ethanol intolerance is an important metric in terms of process economics, and tolerance has often been described as a complex and likely multigenic trait for which complex gene interactions come into play. Here, we resequence the genome of an ethanol-tolerant mutant, show that the tolerant phenotype is primarily due to a mutated bifunctional acetaldehyde-CoA/alcohol dehydrogenase gene (adhE), hypothesize based on structural analysis that cofactor specificity may be affected, and confirm this hypothesis using enzyme assays. Biochemical assays confirm a complete loss of NADH-dependent activity with concomitant acquisition of NADPH-dependent activity, which likely affects electron flow in the mutant. The simplicity of the genetic basis for the ethanol-tolerant phenotype observed here informs rational engineering of mutant microbial strains for cellulosic ethanol production.
Brown SD, Guss AM, Karpinets TV, Parks JM, Smolin N, Yang S, Land ML, Klingeman DM, Bhandiwad A, Rodriguez M Jr, Raman B, Shao X, Mielenz JR, Smith JC, Keller M, Lynd LR. Mutant alcohol dehydrogenase leads to improved ethanol tolerance in Clostridium thermocellum. Proc Natl Acad Sci U S A. 2011 Aug 16;108(33):13752-7. doi: 10.1073/pnas.1102444108. Epub 2011 Aug 8. PMID: 21825121; PMCID: PMC3158198.
Dartmouth Digital Commons Citation
Brown, Steven D.; Guss, Adam M.; Karpinets, Tatiana V.; and Parks, Jerry M., "Mutant Alcohol Dehydrogenase Leads to Improved Ethanol Tolerance in Clostridium Thermocellum" (2011). Dartmouth Scholarship. 1532.