The role of Lys-228 residue in horse liver alcohol dehydrogenase activity

Abstract

Lys-228 in horse liver alcohol dehydrogenase isoenzyme E (HLADH-E) was mutated to glycine by site-directed mutagenesis. The specific activity of the mutant enzyme was increased about 4-fold and Michaelis constants for NAD+(K(a)) and NADH (K(q)) increased by about 350- and 50-fold, respectively. The wild-type enzyme and K228G mutant enzyme were treated with ethylacetimidate. Acetimidylation of the wild-type enzyme increased the activity about 10-fold, but the mutant enzyme was little affected. These results confirm that Lys-228 residue plays an important role in the activity of the enzyme through forming the hydrogen bond with adenosine ribose of NAD+.