Regulation of the PTH-receptor-cyclase system of canine kidney: effects of calcium, magnesium, and guanine nucleotides.

These studies examine the interactions of Ca2+, Mg2+, and guanine nucleotides with PTH binding to renal receptors and activation of adenylate cyclase in a purified preparation of basolateral renal cortical membranes. Inhibition of adenylate cyclase activity by increasing concentrations of CA2+ was more pronounced in the presence of PTH than on basal activity. This effect was not explained by an effect of Ca2+ on equilibrium PTH binding or binding affinity. The presence of PTH also decreased the requirements of the enzyme for Mg2+, consistent with the competitive nature of Ca2+ and Mg2+ interaction. Guanine nucleotides produced similar changes in the metal requirements for enzyme activity. PTH and guanine nucleotides, however, were not additive in this regard in spite of marked synergism in total enzyme activity. Additional studies were performed to further define the effects of guanine nucleotides on the receptor-cyclase system. GTP and Gpp(NH)p caused a dose-dependent decline in equilibrium PTH binding and a small increase in half-maximal displacement. PTH-stimulated adenylate cyclase activity was markedly increased by either nucleotide, and the half-maximal activity was decreased (Gpp(NH)p greater than GTP). This was partly explained by a marked effect of guanine nucleotides on the dissociation rate of receptor-bound PTH. These data suggest that Ca2+ inhibition of PTH-stimulated adenylate cyclase activity is not due to a decreased binding of PTH but to an effect of PTH on the metal requirements for adenylate cyclase activation at the nucleotide regulatory site.