| Abstract: | Prostaglandin synthase (EC 1.14.99.1, 8,11,14-eicosatrienoate, hydrogen-donor:oxygen oxidoreductase), commonly referred to as cyclooxygenase, was inhibited irreversibly upon application of a fixed oxidative potential (+0.4 V versus saturated calomel electrode) in the presence of the aspirin metabolite gentisic acid (2,5-dihydroxybenzoic acid). Electrolyses were carried out at 0 degrees C in a phosphate buffer solution (pH 7.2, 0.1 M) using a carbon felt electrode. This electroinactivation process was time-dependent and pseudo first-order with respect to gentisic acid at concentrations up to 300 microM. These concentrations of gentisic acid are below those normally reported to be inhibitory. The enzyme was stable to this applied potential in the absence of gentisic acid. Similar treatment of apoenzyme (heme-removed) revealed no loss in catalytic activity after reconstitution to the holoenzyme. Oxyphenbutazone, a nonoxidizable competitive inhibitor of cyclolooxygenase, was observed to protect the enzyme from electrolytic inactivation mediated by gentisic acid. Radiolabeling studies indicated the covalent attachment of approximately 1 eq of gentisic acid/subunit of enzyme. These studies support the possible role of quinonoid intermediates in the observed anti-inflammatory action of salicylate derivatives. |
