Studies on renal aldosterone-binding proteins.

AH-Aldosterone-ma :romolecular complexes were isolated from the nuclear and :ytosol fractions of rat kidneys by precipitation with (NHS&iOc and by chromatography on Sephadex G-50 and G-75. l’hese complexes were recovered after injection of 3H-aldostN crone into adrenalectomized rats and after incubation of la bel’ed steroid with renal nuclear extracts and cytosol fract. ons in vitro. The nuclear aldosterone binding substances we*e identified as proteins by the following criteria: (a) chemical analysis of Tris-CaClz extracts of purified nuclear fractions; (b) dissociation of the complexes by wide spectrum proteolytic enzymes but not by nucleases; and (c) loss of 3H-aldosterone-binding activity when the complexes were treated with the sulfhydryl reagents, p-hydroxymercuribenzoate and Z-(2aminoethyl)-2-thiopseudourea dihydrochloride. The cytosol aldosterone-binding substances were identified as proteins by the following criteria: (a) dissociation of the complexes by wide spectrum proteolytic enzymes but not by nucleases, lipase, or phospholipase; (b) loss of 3H-aldosterone-binding activity on treatment with p-hydroxymercuribenzoate. Estradiol-17/3,6cr-methylprednisolone, deoxycorticosterone, and 9a-fluorocortisol impaired the formation of the 3H-aldosterone-macromolecular complexes in the nuclear and cytosol fractions in direct proportion to their potencies in the regulation of renal tubular transport of sodium. In the in vifro system, spirolactone inhibited the formation of the 3H-aldosterone complex at concentration ratios that inhibit the action of aldosterone in the rat in uivo. The isomer of aldosterone, 17-isoaldosterone, was not bound to the proteins in nuclear extracts, and at a lo-fold concentration ratio did not inhibit the formation of 3H-aldosterone complexes in nuclear extracts. In addition, 17-isoaldosterone (1.4 x lOA M) had no effect on active sodium transport in the isolated toad bladder system and at 1 pg/150 g of body weight had no effect on the urinary Na+: K+ concentration ratios in the