Binding of (3H)prostaglandin E1 to putative receptors linked to adenylate cyclase of cultured cell clones.
A method for assessing the binding of 3H-labeled prostaglandin E1 ([3H]PGE1) to cell membranes has been developed and used to study the interaction of [3H]PGE1 with membranes from cultured mammalian cells. Receptor sites were identified by correlation of the potency of a series of compounds to compete for [3H]PGE1 binding sites and to stimulate adenylate cyclase activity, by correlation of rates of binding and change in enzyme activity, and by the correspondence of [3H]PGE1-binding activity with the presence or absence of PGE1-sensitive adenylate cyclase in several clones. In clone B82, a murine L-cell, [3H]PGE1 binds with an activation energy of 14 kcal/mol to a class of sites with an affinity of 0.5 X 10(8) M-1 and a capacity of 150 fmol/mg of protein. Concentration dependence of adenylate cyclase activation by PGE1 (KD =30 nM) and kinetic analysis of [3H]PGE1 binding (k1 = 4 X 10(6) liters/mol/min, k-1 0.15/min) verify this affinity. Concentration dependence and specificity of binding and activation of adenylate cyclases in neuroblastoma clone N4TG1 and N18TG2 substantiate the method. In several clones that lack PGE1-responsive adenylate cyclase, no specific [3H]PGE1 binding is detectable.
D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D004305:Dose-Response Relationship, Drug; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007739:L Cells / Q000378:metabolism; D051379:Mice; D009447:Neuroblastoma / Q000378:metabolism; D011453:Prostaglandins / Q000494:pharmacology; D011458:Prostaglandins E / Q000378:metabolism*; D011485:Protein Binding; D011956:Receptors, Cell Surface; D013696:Temperature; D014316:Tritium
D011453:Prostaglandins; D011458:Prostaglandins E; D011956:Receptors, Cell Surface; D014316:Tritium; D000262:Adenylyl Cyclases