Mutant alcohol dehydrogenase leads to improved ethanol tolerance in Clostridium thermocellum

Brown, Steven D.; Guss, Adam M.; Karpinets, Tatiana; Parks, Jerry M.; Smolin, Nikolai; Yang, Shihui; Land, Miriam L.; Klingeman, Dawn Marie; Bhandiwad, Ashwini; Rodriguez Jr, Miguel; Raman, Babu; Shao, Xiongjun; Mielenz, Jonathan R.; Smith, Jeremy C.; Keller, Martin; Lynd, Lee R.

doi:10.1073/pnas.1102444108

Title: Mutant alcohol dehydrogenase leads to improved ethanol tolerance in Clostridium thermocellum

Journal Article · Mon Aug 01 00:00:00 EDT 2011 · Proceedings of the National Academy of Sciences of the United States of America

DOI:https://doi.org/10.1073/pnas.1102444108· OSTI ID:1037143

^[1];

^[1]; Karpinets, Tatiana ^[1];

^[1]; Smolin, Nikolai ^[1]; Yang, Shihui ^[1];

^[1];

^[1]; Bhandiwad, Ashwini ^[2]; Rodriguez Jr, Miguel ^[1]; Raman, Babu ^[3]; Shao, Xiongjun ^[2]; Mielenz, Jonathan R. ^[1];

^[1]; Keller, Martin ^[1]; Lynd, Lee R. ^[2]

ORNL
Dartmouth College, Thayer School of Engineering
Dow Chemical Company, Midland, MI

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.

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Research Organization:: Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)

Sponsoring Organization:: USDOE

DOE Contract Number:: AC05-00OR22725

OSTI ID:: 1037143

Journal Information:: Proceedings of the National Academy of Sciences of the United States of America, Vol. 108, Issue 33; ISSN 0027-8424

Country of Publication:: United States

Language:: English

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