Photoaffinity labeling of the solubilized, partially purified muscarinic acetylcholine receptor from porcine atria by p-azidoatropine methyl iodide.
The synthesis of a tritiated photoaffinity analogue of the muscarinic antagonist atropine, [3H]-p-azidoatropine methyl iodide is described. The compound appeared to bind to a single class of sites in membrane-bound, solubilized, and partially purified preparations of muscarinic receptor from porcine atria with a dissociation constant (determined by competition vs. [3H]-L-quinuclidinyl benzilate) of about 1.0 X 10(-7) M. This value was in agreement with the apparent dissociation constant (8.5 X 10(-8)M) determined by measuring the concentration dependence of covalent incorporation into a partially purified receptor preparation. Competition experiments indicated that the specific covalent labeling could be blocked by the muscarinic agonist carbamylcholine and the antagonists L-quinuclidinyl benzilate and atropine. An apparent molecular weight of 75 000 +/- 5000 was found for specifically labeled peptide(s) in a solubilized, partially purified receptor preparation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
D000345:Affinity Labels / Q000138:chemical synthesis*; D000818:Animals; D001286:Atropine Derivatives / Q000138:chemical synthesis* / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006325:Heart Atria / Q000378:metabolism; D007202:Indicators and Reagents; D007700:Kinetics; D008970:Molecular Weight; D009206:Myocardium / Q000378:metabolism*; D010777:Photochemistry; D011976:Receptors, Muscarinic / Q000302:isolation & purification / Q000378:metabolism*; D013552:Swine
D000345:Affinity Labels; D001286:Atropine Derivatives; D007202:Indicators and Reagents; D011976:Receptors, Muscarinic; C042562:4-azidoatropine methiodide