1.1.1.71 D194A the mutation results in substantial catalytic deficiency (2.8% compared to the wild type enzyme) 737536 1.1.1.71 E43K/S97C/T148S/A155H/P210C/L227V/Y244F site-directed mutagenesis, mutant ADHA-0381, the mutant shows increased thermostability and altered pH conditions compared to the wild-type enzyme 760769 1.1.1.71 E43K/S97C/T148S/T194V/M206D/P210C/L227V site-directed mutagenesis, mutant ADHA-0398, the mutant shows increased thermostability and altered pH conditions compared to the wild-type enzyme -, 760769 1.1.1.71 G198D site-directed mutagenesis, the variant of the NAD(P)H-dependent ADH, TeSADH, shows a significant switch of cofactor dependence from NADPH towards NADH compared to the wild-type enzyme 760921 1.1.1.71 H198A inactive 737536 1.1.1.71 H267A inactive 737536 1.1.1.71 H281A inactive 737536 1.1.1.71 H363A the mutant with full activity is not able to grow on butanal-containing medium 737536 1.1.1.71 additional information development of improved enzyme variants ADHA-0398 and -0381 shows +26°C improved melting temperature and 17fold higher oxidative activity at pH 6.5 towards (4R,6S)-6-(chloromethyl)oxane-2,4-diol but only 6fold at pH 7.5, method development, overview -, 760769 1.1.1.71 P704L/H734R the ethanol tolerant phenotype of Clostridium thermocellum is primarily due to a mutated bifunctional acetaldehyde-CoA/alcohol dehydrogenase gene (adhE). The mutant displays a complete loss of NADH-dependent activity with concomitant acquisition of NADPH-dependent activity. The reduction in specific activity with respect to NADH is far greater (about 25fold less activity) than the increase with respect to NADPH. Although total ADH activity dropps by 25fold, ethanol production does not drop significantly between strains under these conditions 739526