Hormonal sensitivity of adenylate cyclase incorporated in proteoliposomes

Proteoliposomes containing adenylate cyclase (AC) from rabbit heart ventricles were obtained using a novel reconstitution procedure from solubilized state. The enzyme preparation can be stimulated by 5'guanylylimidodiphosphate (GppNHp) and NaF, but not by isoproterenol. Hormonal responsiveness of AC is restored by either isoproterenol trapped by the proteoliposomes during the reconstitution or pretreatment of proteoliposomes with alamethicin. GTP in the presence of alamethicin decreases the affinity of beta-adrenoceptors to the agonist, thus confirming that the properties of reconstituted AC system do not differ from the native one. It is demonstrated that the degree of AC activation by isoproterenol depends strongly on the beta-adrenoceptors content in the proteoliposomes, which in turn can be changed artificially in the process of reconstitution. The described reconstitution technique might be a useful tool for investigating the role of component stoichiometry in the functioning of hormone-regulated AC-system.