Studies of binding of parathyroid hormone to a detergent-dispersed preparation from bovine kidney cortex plasma membranes.
Bovine kidney plasma membranes containing parathyroid hormone-sensitive adenylate cyclase activity were dispersed with 1% Triton X-100 and centrifuged at 150,000 X g for 2 h. Approximately 40% of the total membrane protein was extracted by this procedure. The extraction greatly reduces the fluoride-stimulated and the parathyroid hormone-sensitive adenylate cyclase activity of the membranes and yields a supernatnat which binds biologically active, tritiated parathyroid hormone. Hormone binding is stable for up to 15 h and has a linear dependence on protein concentration in the extract. Binding of the labeled hormone at concentrations of 5 to 10 nM is inhibited by preincubation with unlabeled min, and displays a dependence on temperature, time, and pH. Binding specificity is maximal at physiological pH, being inhibited by only the native hormone or its synthetic 1-34 NH2-terminal, biologically active fragment. Binding increases dramatically at pH 6.0, but is nonspecific in character. Half-maximal inhibition of the binding was achieved at 3.2 X 10(-7) M concentrations of the native hormone and 5.0 X 10(-7) M concentrations of the synthetic 1-34 NH2-terminal fragment. Calcium does not inhibit either total or specific binding. Inhibition, kinetic, and pH dependence data suggest that the extracted component(s) represent the parathyroid hormone binding protein(s) formerly identified in particulate membrane preparations.
D000818:Animals; D001667:Binding, Competitive; D002118:Calcium / Q000494:pharmacology; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007328:Insulin / Q000494:pharmacology; D007672:Kidney Cortex / Q000378:metabolism*; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism*; D010281:Parathyroid Hormone / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D011092:Polyethylene Glycols / Q000494:pharmacology*; D011485:Protein Binding; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology*; D012995:Solubility
D007328:Insulin; D008565:Membrane Proteins; D010281:Parathyroid Hormone; D010446:Peptide Fragments; D000644:Quaternary Ammonium Compounds; D011092:Polyethylene Glycols; D002118:Calcium