ACTIVITY OF 11-BETA-HYDROXYSTEROID DEHYDROGENASE IN TOAD BLADDER - EFFECTS OF 11-DEHYDROCORTICOSTERONE

11Beta-hydroxysteroid dehydrogenase (11beta-OHSD) transforms circulating glucocorticoids to their ''biologically inert'' 11-dehydro derivatives. Isoforms of 11beta-OHSD with different cofactor requirements and biochemical properties [Michaelis constant (K(m)) and maximal velocity (V(max))] exist in the kidney. Since epithelial cells derived from the toad bladder also contain this enzyme, we wished to further characterize its properties in prepared cell homogenates. 11Beta-OHSD from toad bladder demonstrated a clear preference for NAD+ over NADP+ as a cofactor similar to that observed in renal cortical collecting duct (CCD) cells. Furthermore, 11beta-OHSD had a rapid onset of action. The apparent K(m) for corticosterone was 16.3 x 10(-8) M, a value comparable to that observed for enzyme from CCD, and a V(max) of 4.8 x 10(-12) mol. mg protein-1 min-1. The end product, 11-dehydrocorticosterone (compound A), influenced enzyme activity; it increased 11beta-OHSD activity at corticosterone concentrations below the apparent K(m) for the enzyme and inhibited 11beta-OHSD activity at corticosterone concentrations above the K(m) for the enzyme. The inhibitory effects of compound A appeared noncompetitive with an apparent equilibrium constant (K(i)) of 2.8 X 10(-7) M. Consistent with its inhibitory action on 11beta-OHSD, compound A (10(-6) M) enhanced the short-circuit current response to corticosterone (10(-7) M) in the intact toad bladder (experimental 2.03 +/- 0.33 vs. control 1.40 +/- 0.17 times above baseline; n = 7, P < 0.01). Thus 11beta-OHSD in toad bladder resembles the isoform found in CCD, and compound A may be biologically important as a regulator of 11beta-OHSD.