Sulfhydryl groups of aldosterone receptors from swine kidney

Abstract Specific [ 3 H]aldosterone binding activity in swine kidney cytosol was inactivated by pretreatment of the cytosol with monoiodoacetamide (pH 8.5), N-ethylmaleimide (pH 7.0), or 5,5′-dithiobis(2-nitrobenzoate) (pH 7.5). Dithiothreitol restored the specific binding activity inactivated by the nitrobenzoate, but not that inactivated with ethylmaleimide. Incubation of the cytosol with aldosterone prior to pretreatment with ethylmaleimide protected the receptors from inactivation. The rank order of steroids for the protection was: aldosterone > hydrocortisone ≧ dexamethazone = progesterone > triamcinolone > estradiol. The initial velocity of the specific hormone binding could be determined by the binding reaction for 60 sec at 30°. Double reciprocal plots of the initial velocity versus the hormone concentration with or without the nitrobenoate showed a typical pattern of competition between the hormone and the inactivator. The results indicated the presence of functional sulfhydryl groups on the hormone binding sites of aldosterone receptors.