Solubilization and characterization of the cholecystokininB binding site from pig cerebral cortex |
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| Affiliation: | 1. MRC Molecular Neurobiology Unit, MRC Center, Hills Road, Cambridge CB2 2QH, U.K.;2. Parke-Davis Research Unit, Addenbrookes Hospital Site, Hills Road, Cambridge CB2 2QB, U.K.;1. Utrecht School of Economics, Utrecht University, the Netherlands;2. Institute for Operations Research and Computational Finance, University of St. Gallen, Switzerland;3. Faculty of Business and Economics, Technische Universität Dresden, Germany;4. Queen’s Management School, Queen’s University Belfast, UK;5. USEK Business School, Holy Spirit University of Kaslik, Jounieh, Lebanon;1. Department of Business Administration I, Cartuja s/n, 18071, Business Management Faculty. University of Granada, Spain;2. Department of Business Administration, Marketing and Sociology, Campus Las Lagunillas, s / n, 23071, University of Jaen, Spain;1. Department of Mechanical Engineering, IIT (ISM) Dhanbad, India;2. Department of Mechanical Engineering, NIT Agartala, India;1. Department of Food Technology, Institute of Human Nutrition and Food Science, CAU University, Kiel, Schleswig Holstein, Germany;2. Department of Food Technology, Universitiy Putra, Serdang, Selangor, Malaysia |
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| 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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