Proteolytic activation of grass carp (Ctenopharygodon idellus) liver alcohol dehydrogenase.

Two different forms of alcohol dehydrogenase (ADH) were prepared from the liver of grass carp, using ion-exchange chromatography on DEAE-cellulose and affinity chromatography on Affi-gel Blue agarose. In the presence of dithiothreitol and benzamidine. ADH-l, a dimeric protein with native molecular weight of 80 kDa, was obtained. However, in their absence, another form of the enzyme, ADH-C, consisting of two 27 kDa and two 13 kDa subunits tightly but non-covalently associated with each other, can be isolated. ADH-C represented a cleaved form of the enzyme and was shown to be derived from the original, intact form, ADH-l through limited and specific proteolytic cleavage. Surprisingly, such a cleavage caused an activation of the enzyme toward the oxidation of ethanol at pH 10.5. The Vmax value of ADH-C was 5.16-fold greater than that of ADH-l. However, ADH-C had much weaker binding affinity for the alcohol. Its Km value toward ethanol was 163-fold higher than that of ADH-l.