STATES OF ACTIVATION OF CHICK KIDNEY ADENYLATE CYCLASE INDUCED BY PARATHYROID HORMONE AND GUANYL NUCLEOTIDES

Several aspects of the activation of adenylate cyclase by guanosine 5′-triphosphate (GTP), 5′-guanylylimidodiphosphate (Gpp(NH)p) and bovine parathyroid hormone (bPTH) have been studied in chick kidney plasma membrane preparations. GTP (10−4mol/l), Gpp(NH)p (10−4mol/l) and bPTH (10 i.u./ml) activated adenylate cyclase without any significant time lag. However a 2 min delay was observed before the activity of the enzyme increased after the addition of bPTH (−6 → + 34) to incubations.The early (0–3 min) effects of GTP and Gpp(NH)p upon chick kidney adenylate cyclase activity were antagonized by the addition of the alternative guanyl nucleotide. After 5 min of incubation with kidney plasma membranes, Gpp(NH)p induced a stable state of activation of adenylate cyclase which was not reversible by subsequent addition of GTP. GTP did not induce an irreversible state of enzyme activation. In pre-incubation studies, GTP did not produce a persistent enzyme activation and did not modify the effect of Gpp(NH)p added subsequently at the incubation stage. Gpp(NH)p produced a stable state of activation of adenylate cyclase which was not inhibited by addition of GTP at the incubation stage.Bovine PTH (2–34) inhibited the effect of bPTH upon adenylate cyclase activity when the native hormone (10 i.u./ml) had been incubated with plasma membranes for up to 8 min before addition of the analogue (5 μg/ml). Incubation of plasma membranes with bPTH (2–34) for as little as 10 s prevented activation of adenylate cyclase by subsequent addition of bPTH. This pattern was confirmed in pre-incubation studies. After pre-incubation of kidney membranes with bPTH and bPTH (2–34), followed by washing, an acid extract of the membranes contained immunoreactive bPTH. Gpp(NH)p produced a greater increase in adenylate cyclase activity in membranes pre-incubated with bPTH or bPTH (2–34) than in membranes pre-incubated with buffer alone, suggesting that the hormone and analogue facilitated the interaction of Gpp(NH)p with adenylate cyclase.