The effects of magnesium on calcium inhibition of parathyroid adenylate cyclase.

cAMP has been shown to mediate the response of the parathyroid gland to a number of agonists and appears to take part in the regulation of this gland by divalent cations as well. We have studied the effects of the concentrations of free magnesium (Mg+2) and ionic calcium (Ca+2) on the kinetic properties of normal porcine parathyroid adenylate cyclase. In a previous study we obtained evidence for two calcium inhibition sites in this enzyme complex. In the present study we observed that the Mg+2 concentration influences the relative contribution of these sites to the overall calcium inhibition. At a high Mg+2 concentration (10 mM), the high affinity site contributes less than 50% of the total calcium inhibitable activity, whereas at a Mg+2 concentration in the low physiological range (0.5 mM), the high affinity site accounts for all the calcium inhibitable activity. Mg+2 was found to be a potent activator of porcine parathyroid adenylate cyclase, with a Ka of Mg of 0.8-2 mM. Ionic calcium at low concentrations (0.2-5 microM) acts as a competitive inhibitor with respect to Mg+2 activation. The calcium inhibition constant was estimated to be 2-3 microM. The Km for ATPMg-2 was 0.3 mM, which is similar to that found in other studies of adenylate cyclase activity in parathyroid tissue. The effects of Ca+2 on enzymatic activity with respect to the ATPMg-2 concentration showed noncompetitive inhibition. The calcium inhibition constant with respect to its effect on Vmax (KIv) was 3 microM; the calcium inhibition constant with respect to its effect on the binding of ATPMg-2 (KIs) was 10 microM. It is concluded from these results that the concentrations of intracellular Ca+2 reported to be present in parathyroid cells could inhibit adenylate cyclase activity. The mode of calcium inhibition that involves competition with magnesium would be particularly significant at low intracellular Mg+2 concentrations, and this phenomenon may account for the parathyroid secretory defect which is a characteristic feature of the magnesium-deficient state.