Mechanism of glucagon stimulation of adenylate cyclase in the presence of GDP in rat liver plasma membranes.

In the present study on the specific role of GDP in regulation of the hepatic glucagon-sensitive adenylate cyclase system, two approaches were employed to eliminate the influence of the conversion of GDP during incubation: One was a time study during only the initial period of incubation with ATP as substrate and the other was a study using adenyl-5’-yl imidodiphosphate, a nonphosphorylating analog of ATP, as substrate. With ATP as substrate, GDP caused transient inhibition (lag) of the rate of accumulation of cyclic AMP in the first 30 s of incubation, whereas GTP caused transient stimulation (burst). Glucagon dose-dependently increased the “steady state” rate (1 to 3 min) to a similar extent in the presence of GDP and of GTP with concomitant loss of the lag with GDP and the burst with GTP. The effect of GTP resembled that of guanyl5’-yl imidodiphosphate in the early period of incubation but became qualitatively indistinguishable from that of GDP after establishment of a steady state, irrespective of the concentration of glucagon or guanine nucleotide. With adenyl-5’-yl imidodiphosphate as substrate, glucagon again increased the enzyme activity in the presence of GTP and of guanyl-5’-yl imidodiphosphate, whereas in the presence of GDP glucagon stimulation was markedly reduced. The reduced response of the enzyme to glucagon in the presence of GDP was completely restored to the same level as in the presence of GTP by the further addition of one of the nucleoside triphosphates (ITP, XTP, CTP, or UTP). These results show that GDP (inhibitor of the enzyme) and GTP (stimulator) may be in equilibrium at or near the guanine nucleotide sites when ATP is present as substrate and that the glucagon stimulation of the enzyme in the presence of GDP requires the formation of GTP by transfer of a terminal phosphate of ATP to GDP. The results suggest that glucagon may shift the equilibrium between GDP and GTP toward GTP by enhancing the formation of GTP, resulting in a high activity state of the enzyme.