Properties of gonadotropin-stimulable adenylyl cyclase of the human corpus luteum: regulation of hormonal responsiveness by guanyl nucleotide and magnesium ion.

We studied the properties of a gonadotropin-responsive adenylyl cyclase in membrane preparations obtained from human corpus luteum and explored the nature of guanyl nucleotide and Mg ion involvement in activation of the enzyme. Maximal adenylyl cyclase activity required ATP concentrations of 1-2.5 mM, total MgCl2 concentrations of 8-10 mM, and 1 mM EDTA. Optimal hCG responsiveness, however, required lower (approximately 5 mM) MgCl2 concentrations. Both GTP and its hydrolysis-resistant analog guanyl 5'-yl imidodiphosphate [GMP-P(NH)P] increased enzyme activity, but the response to each nucleotide had distinct characteristics. The rate and the extent of activation were greater in the presence of GMP-P(NH)P than in that of GTP. Moreover, enzyme activation by GMP-P(NH)P was hysteretic in nature, requiring about 8 min to reach steady state velocity in the absence of hormonal stimuli. The slow rate of activation by GMP-P(NH)P was accelerated by either hCG or increases (3-10 mM) in the concentration of MgCl2. Thus, both gonadotropin and Mg ion are inherently antihysteretic in the human luteal adenylyl cyclase system. Basal and hCG stimulation were under the control of guanine nucleotides. Dose-response curves showed that the apparent activation constants for GTP and GMP-P(NH)P were 0.30 and 0.51 microM, respectively; these values did not shift after the addition of hCG. At a higher concentration of guanyl nucleotides (1000 microM), basal and hCG-stimulated activities were markedly reduced, suggesting bimodal regulation of the enzyme by the nucleotides. We also found that enzyme responsiveness to prostaglandin E2 was small and that, in contrast to a number of other nonprimate species, adenylyl cyclase from the human corpus luteum was not stimulated by isoproterenol. Taken together, these data support the usefulness of the cell-free model for studying the role of adenylyl cyclase in the regulation of luteal function in the human.