Inhibitors of glucagon inactivation: Effect on glucagon-receptor interactions and glucagon-stimulated adenylate cyclase activity in liver cell membranes

https://doi.org/10.1016/0304-4165(74)90242-6Get rights and content

Abstract

The specific binding of 125I-labeled glucagon to liver cell microsomal membranes is accompanied by a rapid loss in the biological activity of the hormone present in the medium. Inhibitors of glucagon inactivation have been identified and the effects of such compounds on hormone-binding and hormone-stimulated adenylate cyclase activity have been examined.

  • 1.

    1. Compounds which inhibit inactivation of glucagon by microsomal membranes include: (a) peptide hormones of low molecular weight which have little or no tertiary structure, such as adrenocorticotropic hormone; (b) peptide antibiotics, such as bacitracin; (c) bile salts and synthetic detergents, such as sodium deoxycholate and Triton X-100; (d) polyene antibiotics, such as filipin; and (e) metal-complexing agents, such as 1,10-phenantroline.

  • 2.

    2.Under appropriate conditions, inhibitors of glucagon inactivation cause a significant (20–80%) increase in glucagon-binding to liver microsomal membranes when subsaturating concentrations of hormone are used. The ability of the various inhibitors to suppress inactivation correlates well with their ability to enhance hormone binding. The effects of the inhibitors on binding result from an enhanced affinity of the glucagon-membrane interaction, and not from the exposure of new glucagon-binding sites.

  • 3.

    3. Adrenocorticotropic hormone and bacitracin increase the apparent affinity of adenylate cyclase of liver membranes for glucagon, but do not affect the maximal activity of the enzyme. Such effects are not apparent with other inhibitors tested, probably because some of these compounds perturb unfavorably the catalytic properties of adenylate cyclase.

  • 4.

    4. When inactivation of inhibited the glucagon-membrane interaction obeys kinetics of simple, dissociable processes. In this case the rate constant of glucagon-membrane association (106 M−1 · −1) and of dissociayion (2 · 10−4 s−1) can be measured independently, and the dissociation constant (2 · 10−10 M) determined from these rate constants is similar to that (2.5 · 10−10 M) calculated separately from equilibrium data.

  • 5.

    5. Selective inhibition of the glucagon-inactivating components of liver cell membranes allows more meaningful kinetic characterization of the hormone-receptor interaction in these membranes and may also aid in further studies designed to isolate and purify the receptors.

References (35)

  • M. Rodbell et al.

    J. Biol. Chem.

    (1971)
  • I.D. Goldfine et al.

    J. Biol. Chem.

    (1972)
  • V. Tomasi et al.

    Biochim. Biophys. Acta

    (1970)
  • D.M. Neville

    Biochim. Biophys. Acta

    (1968)
  • W.B. Gratzer et al.

    Biochem. Biophys. Res. Commun.

    (1967)
  • P. Cuatrecasas et al.

    Biochem. Biophys. Res. Commun.

    (1971)
  • P. Cuatrecasas

    J. Biol. Chem.

    (1971)
  • M. Rodbell

    J. Biol. Chem.

    (1967)
  • A.A. White et al.

    Anal. Biochem.

    (1971)
  • J.H. Exton et al.

    J. Biol. Chem.

    (1971)
  • M. Rodbell et al.

    J. Biol. Chem.

    (1971)
  • S.L. Pohl et al.

    J. Biol. Chem.

    (1972)
  • S. Kakiuchi et al.

    J. Biol. Chem.

    (1964)
  • A.A. Iodice

    Arch. Biochim. Biophys.

    (1967)
  • J.K. McDonald et al.

    J. Biol. Chem.

    (1969)
  • A.I. Cohen et al.

    J. Biol. Chem.

    (1961)
  • P. Freychet et al.

    J. Biol. Chem.

    (1972)
  • Cited by (88)

    View all citing articles on Scopus

    Present address: Unite 30 I.N.S.E.R.M., 149 rue de Sevres, 75015 Paris, France.

    View full text