Inactivation and conformational changes of yeast alcohol dehydrogenase in trifluoroethanol solutions.

Conformational changes of yeast alcohol dehydrogenase in trifluoroethanol solutions have been followed by fluorescence emission and circular dichroism spectroscopy. At low concentration (less than 5%), trifluoroethanol shows a reversible inhibition competitive to ethanol and noncompetitive to NAD+. The inhibition constants for native and structural-zinc-removed yeast alcohol dehydrogenase were 5.8 and 1.1 mM, respectively, suggesting that the active site becomes more flexible after the structural zinc is removed. At higher trifluoroethanol concentrations the enzyme was irreversibly inactivated. Comparison of inactivation and conformational changes of yeast alcohol dehydrogenase denatured in trifluoroethanol solutions shows that the extent of inactivation is larger than the extent of conformational changes at the same trifluoroethanol concentration. The results obtained from circular dichroism spectra show that the presence of trifluoroethanol can induce the formation of secondary structure of the enzyme.