title: Do canrenone and 6,7-dihydroxylated derivatives compete with ouabain at the same site on Na,K-ATPase?
abstract: Canrenone is the major metabolic product of the synthetic steroids spironolactone and K+-canrenoate, used in antihypertensive therapy. Canrenone can competitively displace [3H]ouabain from Na,K-ATPase [Na+- and K+-activated, Mg2+-dependent adenosine triphosphatase (E.C.3.6.1.3)] and partially inhibit enzymatic activity. These features have led to a hypothesis that competition between canrenone and endogenous digitalis-like materials, suggested to be involved in etiology of essential hypertension, could underly the antihypertensive effect of canrenone. Surprisingly, three derivatives of canrenone (6 beta,7 alpha-,6 beta,7 beta-, and 6 alpha,7 alpha-dihydroxy-6,7-dihydrocanrenone) reportedly occur in normal human and rat urine. This paper characterizes the interactions with partially purified renal Na,K-ATPase of canrenone, the three 6,7-dihydroxylated derivatives, and one epoxide intermediate, synthesized from K+-canrenoate. Canrenone and all the 6,7-substituted derivatives partially inhibited Na,K-ATPase activity (39-45%), with approximately the same apparent affinity, 100-200 microM. Canrenone displaced [3H]ouabain in an apparently competitive fashion (Ki = 100-300 microM) but none of the tested derivatives significantly displaced ouabain even at very high concentrations. The ability to differentiate the ATPase inhibition and [3H]ouabain displacement by modification of the 6,7-double bond indicates that inhibition of ATPase activity does not occur from within the ouabain binding site. We suggest that neither canrenone nor the 6,7-derivatives bind to the ouabain site, but rather interact with it 'allosterically.' Our findings do not support the proposed mechanisms for the antihypertensive action of canrenone.
authors.affiliation: Department of Biochemistry, Weizmann Institute of Science, Rehovot, Israel.
authors.forename: D M
authors.initials: DM
authors.lastname: Tal
authors.forename: S J
authors.initials: SJ
authors.lastname: Karlish
mesh_terms: D000959:Antihypertensive Agents / Q000494:pharmacology; D001667:Binding, Competitive; D002192:Canrenone / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D010042:Ouabain / Q000378:metabolism*; D011245:Pregnadienes / Q000494:pharmacology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D000959:Antihypertensive Agents; D011245:Pregnadienes; D010042:Ouabain; D002192:Canrenone; D000254:Sodium-Potassium-Exchanging ATPase