Reconstitution of catecholamine-stimulated adenylate cyclase activity using three purified proteins

β-Adrenergic receptors, the GTP-binding regulatory protein that stimulates adenylate cyclase (G(s)), and adenylate cyclase were each purified and reconstituted into unilamellar vesicles composed of phosphatidylethanolamine and phosphatidylserine (3:2, w/w). The molar ratio of receptor:G(s):adenylate cyclase was estimated to be about 1:10:1. Adenylate cyclase activity in the vesicles was stimulated up to 2.6-fold by β-adrenergic agonists. Stimulation was dependent on the presence of guanine nucleotide, displayed appropriate β-adrenergic selectivity and stereoselectivity for agonists, and was blocked appropriately by β-adrenergic antagonists. Therefore, while additional proteins may modulate adenylate cyclase activity in native membranes, these results show that these three proteins are sufficient for the expression of hormone-stimulated adenylate cyclase.