OXIDATIVE ALCOHOL-DEHYDROGENASE FROM KLUYVEROMYCES-MARXIANUS CULTURESIN DOUBLE-STEP SUBSTRATE MEDIA

Experiments with different carbon sources were performed in order to o btain oxidative alcohol dehydrogenase (ADH(2)) from Kluyveromyces marx ianus. Only a double step substrate medium (glucose + reduced carbon s ource) gave good yields in terms of both biomass and enzymatic activit y. Enzyme purification was achieved by anionic exchange chromatography and by affinity chromatography. Electrophoretic patterns of purified ADH displayed two bands for ethanol or lactate grown yeasts, and three bands for 2-propanol grown cells. Velocity constants for both the fer mentative and the oxidative reaction showed a high degree of oxidative activity (maximum 94% in double step substrate media having ethanol a s the second carbon source). Michaelis-Menten constants for the substr ate indicated an excellent affinity for primary and secondary, saturat ed and unsaturated alcohols, for the enzyme.