Isolation and characterization of plasma membranes containing LH sensitive adenylate cyclase from a Leydig cell tumour

Abstract

A LH [luteinizing hormone] sensitive adenylate cyclase from a tumor Leydig cell was investigated. The plasma membranes, prepared by a 2 phase (dextran-polyethylene glycol) centrifugation method had the following properties: in the presence of LH plus p(NH)ppG (guanosine 5''-.beta.,.gamma.-imido triphosphate) or Fl ions, maximum adenylate cyclase activity was obtained in the plasma membranes with 4-6 mM Mg2+ plus 0.33 to 2 mM ATP. LH alone stimulated adenylate cyclase activity 2-fold when compared with basal activity and the time course of cAMP production was linear up to 45 min. With GTP (10-5 M) and GTP plus LH, adenylate cyclase activity was increased 3- and 6-fold, respectively, for up to 20 min and thereafter declined. In contrast p(NH)ppG (10-5 M) and p(NH)ppG plus LH increased adenylate cyclase activity 7- and 14-fold which was maintained for at least 45 min. Fl ions increased the enzyme activity linearly over 45 min .apprx. 18-fold. When GTP or p(NH)ppG were added alone there was a lag time of activation of .apprx. 10 min which was abolished by the addition of LH. GTP but not p(NH)ppG at concentrations > 10-4 inhibited basal and LH stimulated adenylate cyclase when compared with 10-5 M GTP. The tumor Leydig cell adenylate cyclase is thus essentially similar to other hormone sensitive somatic cells. It is feasible to prepare plasma membranes by a simple method from large quantities of pure Leydig cells.