A proper transmembrane Ca gradient is essential for the stimulation of adenylate cyclase activity by stimulatory GTP-binding protein

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 Ca gradient. The results showed that both basal activity and Gs-stimulated activity of adenylate cyclase were highest in proteoliposomes with a transmembrane Ca gradient similar to physiological condition (1 μM Ca outside and 1 mM Ca inside) and lowest when the transmembrane Ca gradient was in the inverse direction. Such a difference could be diminished following dissipation of the transmembrane Ca 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 Ca gradient is essential not only for higher adenylate cyclase activity but also for its stimulation by Gs. © 1994 Plenum Publishing Corporation.