Reticulocyte cytosol activator protein: its actions upon the beta adrenergic receptor and the N-proteins of adenylate cyclase.
Rat reticulocytes contain a cytosol activator protein (RCAP) that augments catecholamine-sensitive adenylate cyclase activity in reticulocyte membranes. A partially purified preparation of RCAP was obtained by Sephacryl S-200 chromatography and used to elucidate further its mechanism of action. The specific activity of the S-200 fraction to augment isoproterenol responsiveness is increased approximately 1100-fold over the starting material from 1.2 nmoles to 1300 nmoles cyclic AMP formed per milligram of RCAP. The molecular weight is approximately 20,000. In addition to its effects on catecholamine-responsive adenylate cyclase, RCAP is associated with significant increases in basal (0.9 +/- 0.2 to 1.5 +/- 0.4 nmol/mg; p less than 0.02), guanyl-5'-yl imidodiphosphate [Gpp(NH)p]; (3.9 +/- 0.9 to 4.4. +/- 1.1 nmol/mg; p less than 0.005) and fluoride (4.1 +/- 0.6 to 4.8 +/- 0.6 nmol/mg; p less than 0.005) associated activities. RCAP stimulates isoproterenol responsiveness in wild type S49 cell membranes but is inactive in the mutant line, cyc-. RCAP alters the characteristics of agonist binding to the beta-adrenergic receptor of reticulocyte and wild S49 cell membranes, causing a significant increase in the IC50 for isoproterenol. Direct assessment of Ns and Ni components of the adenylate cyclase complex demonstrates that RCAP inhibits cholera toxin-specific ADP-ribosylation of the 42K subunit of Ns and stimulates pertussis toxin-specific ADP ribosylation of the 39K subunit of Ni.
D000244:Adenosine Diphosphate / Q000378:metabolism; D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D002772:Cholera Toxin / Q000494:pharmacology; D004789:Enzyme Activation; D004910:Erythrocyte Membrane / Q000378:metabolism*; D066298:In Vitro Techniques; D007545:Isoproterenol / Q000378:metabolism; D011506:Proteins / Q000494:pharmacology*; D051381:Rats; D011943:Receptors, Adrenergic, beta / Q000187:drug effects* / Q000378:metabolism; D012156:Reticulocytes / Q000378:metabolism*
D011506:Proteins; D011943:Receptors, Adrenergic, beta; D000244:Adenosine Diphosphate; D002772:Cholera Toxin; D000262:Adenylyl Cyclases; D007545:Isoproterenol