A proper transmembrane Ca2+ gradient is essential for the stimulation of adenylate cyclase activity by stimulatory...

Stimulatory GTP-binding Protein (Gs) and adenylate cyclase prepared from bovine brain cortices were co-reconstituted into asolectin vesicles with or without 1000-fold transmembrane Ca2+ gradient. The results showed that both basal activity and Gs-stimulated activity of adenylate cyclase were highest in proteoliposomes with a transmembrane Ca2+ gradient similar to physiological condition (1 microM Ca2+ outside and 1 mM Ca2+ inside) and lowest when the transmembrane Ca2+ gradient was in the inverse direction. Such a difference could be diminished following dissipation of the transmembrane Ca2+ gradient by A23187. Comparable conformational changes of Gs in proteoliposomes were also observed when Gs was labeled with the fluorescence probe, acrylodan. These results may indicate that a proper transmembrane Ca2+ gradient is essential not only for higher adenylate cyclase activity but also for its stimulation by Gs.