Isoproterenol plus guanylyl imidodiphosphate (Gpp(NH)p) activate frog erythrocyte adenylate cyclase to a level much higher than the sum of the activities produced by the catecholamine and the synthetic nucleotide tested separately. Propranolol, the beta-receptor blocking agent, failed to inhibit activity when added after the enzyme system had been preincubated with both isoproterenol and Gpp(NH)p. However, if propranolol was added after only one of the two components had been added, it inhibited the effect of isoproterenol. Production of the propranolol-resistant state by treatment with isoproterenol and Gpp(NH)p did not require the presence of the productive substrate (MgATP). The activated propranolol-resistant state persisted following various treatments of the enzyme preparation including extensive washings of the membranes; considerable activity was retained even after sonication or treatment with the detergent Lubrol-PX, treatments which caused inactivation of the native enzyme. Extensive dilution of the membranes following pretreatment with isoproterenol and Gpp(NH)p did not significantly reduce to the activity of the enzyme. Readdition of isoproterenol after dilution caused some inhibition of adenylate cyclase activity, indicating apparently that the beta-receptor has not become inaccessible. In contrast, preincubation with isoproterenol alone failed to render the enzyme system refractive to propranolol, and dilution readily reduced the activity to negligibly low values. Preincubation with Gpp(NH)p alone also produced a persistent active state but the activity was much lower than that obtained throught the combined action of isoproterenol and Gpp(NH)p. The findings suggest that the hormone may be required only to facilitate the initial interaction of the enzyme with Gpp(NH)p. The differences, in this respect, between Gpp(NH)p and the more labile natural nucleotide, GTP, are discussed.