Solubilization and characterization of the cholecystokininB binding site from pig cerebral cortex

Stephan H. Gut, Catherine D. Demoliou-Mason, John C. Hunter, John Hughes, Eric A. Barnard

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Cholecystokinin (CCK) binding sites were solubilized from pig cerebral cortical membranes with digitonin (2%, w/v) in the presence of Na+ (120 mM) and Mg2+ (5 mM). Scatchard plot transformation of equilibrium binding data obtained with 125I-CCK-8S gave an apparent dissociation constant (Kd) of 0.6 nM, comparable to that obtained in membranes in the presence of these cations. Hill coefficients close to unity suggested the presence of a single population of receptor sites. Competitive inhibition studies with pentagastrin, gastrin(1-17)S and the CCKA receptor antagonist L-364,718 indicated that the solubilized receptor sites were of the B-type (CCKB), with the same pharmacological profile as that observed in membranes. Optimal specific binding of 125I-CCK-8S to membrane-bound and solubilized receptors was obtained in the presence of divalent cations. Both the membrane-bound and the solubilized receptor activity were attenuated by guanylyl-imidodiphosphate (Gpp(NH)p) indicating that the brain CCKB receptors are coupled to G proteins.

Original languageEnglish
Pages (from-to)339-346
Number of pages8
JournalEuropean Journal of Pharmacology: Molecular Pharmacology
Volume172
Issue number4-5
DOIs
Publication statusPublished - 17 Oct 1989

Fingerprint

Cerebral Cortex
Swine
Binding Sites
Cholecystokinin
Membranes
Guanylyl Imidodiphosphate
Devazepide
Digitonin
Pentagastrin
Divalent Cations
G-Protein-Coupled Receptors
Cations
Pharmacology
Brain
Population

Keywords

  • (Pig)
  • Cation effects
  • Cholecystokinin (CCK) receptor solubilization
  • Guanine nucleotide effect

Cite this

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title = "Solubilization and characterization of the cholecystokininB binding site from pig cerebral cortex",
abstract = "Cholecystokinin (CCK) binding sites were solubilized from pig cerebral cortical membranes with digitonin (2{\%}, w/v) in the presence of Na+ (120 mM) and Mg2+ (5 mM). Scatchard plot transformation of equilibrium binding data obtained with 125I-CCK-8S gave an apparent dissociation constant (Kd) of 0.6 nM, comparable to that obtained in membranes in the presence of these cations. Hill coefficients close to unity suggested the presence of a single population of receptor sites. Competitive inhibition studies with pentagastrin, gastrin(1-17)S and the CCKA receptor antagonist L-364,718 indicated that the solubilized receptor sites were of the B-type (CCKB), with the same pharmacological profile as that observed in membranes. Optimal specific binding of 125I-CCK-8S to membrane-bound and solubilized receptors was obtained in the presence of divalent cations. Both the membrane-bound and the solubilized receptor activity were attenuated by guanylyl-imidodiphosphate (Gpp(NH)p) indicating that the brain CCKB receptors are coupled to G proteins.",
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Solubilization and characterization of the cholecystokininB binding site from pig cerebral cortex. / Gut, Stephan H.; Demoliou-Mason, Catherine D.; Hunter, John C.; Hughes, John; Barnard, Eric A.

In: European Journal of Pharmacology: Molecular Pharmacology, Vol. 172, No. 4-5, 17.10.1989, p. 339-346.

Research output: Contribution to journalArticle

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