title: Solubilization and characterization of muscarinic receptors from bovine brain.
abstract: This study compared the capacity of different detergents to solubilize the muscarinic cholinergic receptor (mAChR) from bovine brain, evaluated various procedures for the measurement of [3H]-L-quinuclidinyl benzilate [( 3H]-L-QNB) binding to solubilized receptors, and examined some physical and chemical characteristics of the soluble material. An active form of the mAChR was solubilized using digitonin (1%), Triton X-100 (0.5%), and a digitonin-cholate mixture (1%, 0.1%). Values of maximal binding (Bmax) were 2.01, 0.47, and 0.68 pmoles/mg protein, respectively. Comparison of equilibrium dialysis, charcoal adsorption, and polyethylene glycol precipitation indicated that these methods differ in their estimation of Bmax. A decrease in [3H]-L-QNB binding to digitonin solubilized receptors occurred upon dilution or incubation at 37 degrees. The half-life at 37 degrees C was 25 min., but was increased by glycerol. Antagonist binding to digitonin solubilized receptors was saturable and of high affinity. Agonist binding had Hill coefficients less than 1 and was increased by micromolar concentrations of cupric ions.
mesh_terms: D000818:Animals; D001923:Brain Chemistry; D002217:Carbachol / Q000378:metabolism; D002417:Cattle; D002540:Cerebral Cortex / Q000032:analysis; D019826:Cholic Acid; D002793:Cholic Acids; D002846:Chromatography, Affinity; D003300:Copper / Q000494:pharmacology; D004072:Digitonin; D017830:Octoxynol; D011092:Polyethylene Glycols; D011699:Putamen / Q000032:analysis; D011813:Quinuclidinyl Benzilate / Q000378:metabolism; D011976:Receptors, Muscarinic / Q000378:metabolism; D012995:Solubility; D013696:Temperature
chemical_list: D002793:Cholic Acids; D011976:Receptors, Muscarinic; D011092:Polyethylene Glycols; D011813:Quinuclidinyl Benzilate; D003300:Copper; D002217:Carbachol; D017830:Octoxynol; D019826:Cholic Acid; D004072:Digitonin