Pure beta-adrenergic receptors (beta-AR) isolated from guinea pig lung and pure guanine nucleotide binding regulatory protein (NS) of adenylate cyclase isolated from human erythrocytes have been inserted into phospholipid vesicles, resulting in the functional coupling of these two components. The reconstitution of receptor and NS interactions results in the establishment of a guanine nucleotide sensitive state of the receptor that binds agonists with high affinity. Competition curves of isoproterenol for labeled antagonist binding to vesicles containing both beta-AR and NS are biphasic and reveal two affinity states, one of high (approximately 2 nM) and the other of low affinity (approximately 300 nM). In the presence of guanine nucleotides, the competition curves become monophasic and are shifted to a single low-affinity state for the agonist similar to the situation observed in membrane preparations. In addition, the interactions of the receptor and NS lead to the induction of a GTPase activity in NS. The GTPase activity can be stimulated by beta-adrenergic agonists such as isoproterenol (2-5-fold) and is completely blocked by antagonists such as alprenolol in a stereoselective manner. The established hormone responsive activity retains the beta 2-adrenergic specificity conferred by the pure receptor, and similar extents of stimulation (up to 4-fold) are observed with pure receptor from frog erythrocytes, indicating a similar efficiency of coupling between receptors from different species and NS.(ABSTRACT TRUNCATED AT 250 WORDS)