Demonstration and characterization of two classes of cardiac glycoside binding sites to rat heart membrane preparations using quantitative computer modeling.
Cardiac glycoside binding to rat heart membrane preparations was measured by rapid filtration technique. The binding data were analyzed using quantitative computer analysis. The experimental results using [3H]-ouabain as the labeled ligand were consistent with a model in which cardiac glycoside specific binding occurs at two independent classes of sites. The high affinity sites were characterized by a dissociation constants of 40 nM, 50 nM, and 61 nM for ouabain, digoxin and digitoxin, respectively, with a binding capacity of 1.3 pmoles/mg protein. The lower affinity sites for ouabain were characterized by dissociation constants of 2.3 microM, 67 nM and 71 nM for ouabain, digoxin and digitoxin, respectively, with a binding capacity of 3 pmoles/mg protein. Potassium ions inhibit [3H]-ouabain binding in a dose dependent manner with an IC50 of 500 microM. Quantitative computer modelling indicated that potassium inhibits ouabain binding at both binding sites.
Lutz
R A
RA
Laboratory of Theoretical and Physical Biology, National Institute of Child Health and Human Development, Bethesda, MD 20892.
Lichtstein
D
D
Xu
H
H
Rodbard
D
D
D000818:Animals; D001667:Binding, Competitive; D002301:Cardiac Glycosides / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D004074:Digitoxin / Q000378:metabolism; D004077:Digoxin / Q000378:metabolism; D008297:Male; D009206:Myocardium / Q000378:metabolism*; D010042:Ouabain / Q000378:metabolism; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D011955:Receptors, Drug / Q000187:drug effects / Q000378:metabolism*; D000254:Sodium-Potassium-Exchanging ATPase; D012984:Software
D002301:Cardiac Glycosides; D011955:Receptors, Drug; C028996:cardiac glycoside receptors; D010042:Ouabain; D004077:Digoxin; D004074:Digitoxin; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium