title: Photoaffinity labeling of adenosine transporter in cardiac membranes with nitrobenzylthioinosine.
abstract: The kinetic and molecular properties of the adenosine transporter in guinea pig cardiac membranes were studied using nitrobenzylthioinosine (NBMPR), a potent and specific inhibitor of nucleoside transport. [3H]-NBMPR bound tightly but reversibly to guinea pig cardiac membranes (apparent dissociation constant 0.24 +/- 0.07 nM; maximum binding capacity 1.24 +/- 0.45 pmol of NBMPR bound/mg protein). Reversible high-affinity [3H]NBMPR binding was inhibited in an apparent competitive manner by adenosine (apparent inhibition constant 0.14 mM). L-N-phenylisopropyladenosine (L-PIA) had no effect on NBMPR binding. Exposure of cardiac membranes in the presence of [3H]-NBMPR and dithiothreitol, a free-radical scavenger, to ultraviolet light resulted in covalent incorporation of 3H into polypeptides of apparent molecular weight 66,000-50,000. Covalent attachment of [3H]NBMPR under equilibrium binding conditions was inhibited by adenosine, nitrobenzylthioguanosine , and dipyridamole but was unaffected by the adenosine receptor agonist L-PIA. These data suggest that the photolabeled molecular weight protein (apparent mol wt 66,000-50,000) is involved in adenosine permeation by guinea pig cardiac membranes.
mesh_terms: D000241:Adenosine / Q000378:metabolism*; D000345:Affinity Labels; D000818:Animals; D001692:Biological Transport; D005260:Female; D006168:Guinea Pigs; D008566:Membranes / Q000378:metabolism; D009206:Myocardium / Q000378:metabolism*; D010782:Photolysis; D013868:Thioinosine / Q000031:analogs & derivatives / Q000378:metabolism; D014316:Tritium
chemical_list: D000345:Affinity Labels; D014316:Tritium; D013868:Thioinosine; C001789:4-nitrobenzylthioinosine; D000241:Adenosine