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Mutant alcohol dehydrogenase leads to improved ethanol tolerance in Clostridium thermocellum.


ABSTRACT: 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.

SUBMITTER: Brown SD 

PROVIDER: S-EPMC3158198 | biostudies-literature | 2011 Aug

REPOSITORIES: biostudies-literature

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Mutant alcohol dehydrogenase leads to improved ethanol tolerance in Clostridium thermocellum.

Brown Steven D SD   Guss Adam M AM   Karpinets Tatiana V TV   Parks Jerry M JM   Smolin Nikolai N   Yang Shihui S   Land Miriam L ML   Klingeman Dawn M DM   Bhandiwad Ashwini A   Rodriguez Miguel M   Raman Babu B   Shao Xiongjun X   Mielenz Jonathan R JR   Smith Jeremy C JC   Keller Martin M   Lynd Lee R LR  

Proceedings of the National Academy of Sciences of the United States of America 20110808 33


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 ph  ...[more]

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