Effects of phospholipases C on the beta-receptor-adenylate cyclase system of chick erythrocyte membranes.
The beta-adrenergic receptor located in chick erythrocyte membranes was characterized using (-)-[3H]-dihydroalprenolol ([3H]-DHA) with rapid filtration techniques. The affinity of beta-adrenergic antagonist, (-)-propranolol, was approximately 100-fold higher than that of (+)-propranolol. Catecholamines were bound with the receptor in the following order, (-)-isoproterenol greater than (-)-norepinephrine greater than (-)-epinephrine, suggested the binding site to be beta 1-classification. When the membrane preparation was treated with phosphatidylcholine-hydrolyzing phospholipase C (PCase) of Clostridium perfringens or phosphatidylinositol-specific phospholipase C (PIase) of Bacillus thuringiensis, [3H]-DHA binding was decreased to the level of 66 or 86% of the control, respectively. The treatment with sphingomyelinase C (SMase) of Bacillus cereus, however, did not cause any appreciable reduction of [3H]-DHA binding. Throughout these experiments, the equilibrium dissociation constant (KD) of [3H]-DHA was not influenced by phospholipases C. The affinity of isoproterenol for beta-receptor was decreased in the absence of GTP, but not altered in the presence of GTP by PIase action. Treatment with PCase or SMase, however, did not affect the affinity of isoproterenol for beta-receptor. Treatment with PIase inhibited basal, isoproterenol-stimulated and forskolin-stimulated adenylate cyclase activities. On the other hand, PCase treatment inhibited only isoproterenol-stimulated adenylate cyclase activity, but not basal and forskolin-stimulated activities. These results suggest that membrane phospholipids, especially phosphatidylcholine (PC) and phosphatidylinositol (PI), are directly related to the receptor binding and that PI interacts with adenylate cyclase activity.
D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D001409:Bacillus cereus / Q000201:enzymology; D001413:Bacillus thuringiensis / Q000201:enzymology; D001667:Binding, Competitive; D002645:Chickens; D003016:Clostridium perfringens / Q000201:enzymology; D004082:Dihydroalprenolol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004837:Epinephrine / Q000378:metabolism; D004910:Erythrocyte Membrane / Q000201:enzymology*; D007545:Isoproterenol / Q000378:metabolism; D009638:Norepinephrine / Q000378:metabolism; D011433:Propranolol / Q000378:metabolism; D011943:Receptors, Adrenergic, beta / Q000378:metabolism*; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism*
D011943:Receptors, Adrenergic, beta; D004082:Dihydroalprenolol; D011433:Propranolol; D010738:Type C Phospholipases; D013108:Sphingomyelin Phosphodiesterase; D000262:Adenylyl Cyclases; D007545:Isoproterenol; D009638:Norepinephrine; D004837:Epinephrine