Hormone receptor modulates the regulatory component of adenylyl cyclase by reducing its requirement for Mg2+ and enhancing its extent of activation by guanine nucleotides

Proc Natl Acad Sci U S A. 1982 Sep;79(17):5179-83. doi: 10.1073/pnas.79.17.5179.

Abstract

N-Ethylmaleimide treatment of rat liver plasma membranes results in an adenylyl cyclase (EC 4.6.1.1) system that shows no measurable cyclizing activity but retains both an active glucagon receptor and a receptor-sensitive regulatory component N as assessed by reconstitution into cyclase-negative (cyc-) membranes from S49 murine lymphoma. Treatment of such N-ethylmaleimide-treated membranes, termed C- liver membranes, with guanosine 5'-[gamma-thio]triphosphate (GTP[gamma S] ) and Mg2+, followed by the removal of GTP[gamma S] by washing, yields an activated N which upon mixing with cyc- S49 membranes reconstitutes the cyc- S49 membrane adenylyl cyclase in the absence of added GTP[gamma S]. It was found that GTP[gamma S] activation of the N at saturating concentrations of GTP[gamma S] is slow at low Mg2+ concentration and accelerated by increasing Mg2+ concentrations. Addition of glucagon during the activation results in a lowering of the Mg2+ requirement for full activation from 25 mM to around 10 muM and in concomitant increases in both the rate and the extent of N activation. In contrast to its dramatic effect on Mg2+ requirement, glucagon has little (less than 2-fold) effect on the GTP[gamma S] requirement of N activation. These experiments indicate that the glucagon receptor facilitates activation of N by: (i) decreasing the apparent Km of N for Mg2+, and (ii) increasing the extent of activation that can be elicited by saturating concentrations of guanine nucleotide. It is postulated that the mechanism by which Mg2+ and receptors facilitate N activation involves dissociation of n alpha activated ADP-ribosylatable subunits (with guanine nucleotide bound to them) from n beta non-ADP-ribosylatable subunits (with receptor and Mg2+ bound to them).

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adenylyl Cyclases / metabolism*
  • Animals
  • Cell Line
  • Cell Membrane / metabolism
  • Ethylmaleimide / pharmacology*
  • Glucagon / metabolism*
  • Glucagon / pharmacology
  • Guanosine 5'-O-(3-Thiotriphosphate)
  • Guanosine Triphosphate / analogs & derivatives*
  • Guanosine Triphosphate / pharmacology
  • Kinetics
  • Liver / metabolism*
  • Lymphoma / metabolism*
  • Magnesium / pharmacology*
  • Mice
  • Neoplasms, Experimental / metabolism
  • Receptors, Cell Surface / drug effects
  • Receptors, Cell Surface / metabolism*
  • Receptors, Glucagon
  • Thionucleotides / pharmacology*

Substances

  • Receptors, Cell Surface
  • Receptors, Glucagon
  • Thionucleotides
  • Guanosine 5'-O-(3-Thiotriphosphate)
  • Guanosine Triphosphate
  • Glucagon
  • Adenylyl Cyclases
  • Magnesium
  • Ethylmaleimide