The role of zinc in alcohol dehydrogenase. V. The effect of metal-binding agents on thestructure of the yeast alcohol dehydrogenase molecule.

The role of zinc in the catalytic action of yeast alcohol dehydrogenase has been studied through the kinetics of the inhibition of activity by chelating agents (I), particularly l,lO-phenanthroline (2-4). In aqueous solution the complexes of 1, lo-phenanthroline with Zn++ ions exhibit characteristic absorption spectra, as do the complexes of this agent with the zinc atoms of yeast alcohol dehydrogenase and of other zinc metalloenzymes (5). The molecular stoichiometry of the enzyme-inhibitor complexes, deduced from spectrophotometric measurements, is in agreement with that inferred from kinetic data (6). Whereas the catalytic activity of the enzyme can thus be decreased by the localized attack of a chelating agent on a component of an “active site,” alterations in the protein structure may have similar functional consequences, of course. Chelating agents are here shown to induce changes in enzymatic activity both through such local action and through subsequent alterations of the macromolecular structure of yeast alcohol dehydrogenase. As a function of the time of exposure to chelating agents and of their concentration, the apoenzyme, molecular weight 151,000, dissociates into four equal subunits, molecular weight 36,000, while the four zinc atoms are removed concomitantly. Thus, a direct correlation between the enzymatic activity, the zinc content, and the protein structure of the enzyme can be shown to exist. A preliminary report has been made (7).