Zinc inhibition of cAMP signaling

Zn2+ is required as either a catalytic or structural component for a large number of enzymes and thus contributes to a variety of important biological processes. We report here that low micromolar concentrations of Zn2+ inhibited hormone- or forskolin-stimulated cAMP production in N18TG2 neuroblastoma cells. Similarly, low concentrations inhibited hormone- and forskolin-stimulated adenylyl cyclase (AC) activity in membrane preparations and did so primarily by altering the V-max of the enzyme. Zn2+ also inhibited recombinant isoforms, indicating that this reflects a direct interaction with the enzyme. The IC50 for Zn2+ inhibition was similar to1-2 muM with a Hill coefficient of 1.33. The dose-response curve for Zn2+ inhibition was identical for AC1, AC5, and AC6 as well as for the C441R mutant of AC5 whose defect appears to be in one of the catalytic metal binding sites. However, AC2 displayed a distinct dose-response curve. These data in combination with the findings that Zn2+ inhibition was not competitive with Mg2+ or Mg2+/ATP suggest that the inhibitory Zn2+ binding site is distinct from the metal binding sites involved in catalysis. The prestimulated enzyme was found to be less susceptible to Zn2+ inhibition, suggesting that the ability of Zn2+ to inhibit AC could be significantly influenced by the coincidence timing of the input signals to the enzyme.