Solubilization and characterization of σ-receptors from guinea pig brain membranes


Kavanaugh, M.P., Parker, Joel D., Bobker , D.H., Keana, J.F.W. and Weber, E. (1989) Solubilization and characterization of σ-receptors from guinea pig brain membranes. Journal of Neurochemistry , 53, (5), 1575-1580. (doi:10.1111/j.1471-4159.1989.tb08554.x).

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Description/Abstract

The σ-receptor, a distinct binding site in brain tissue that may mediate some of the psychotomimetic properties of benzomorphan opiates and phencyclidine, has been sol-ubilized using the ionic detergent sodium cholate. Binding assays were performed with the solubilized receptor using vacuum filtration over polyethyleneimine-treated glass fiber filters. The pharmacological specificity of the solubilized binding site for σ-receptor ligands is nearly identical to the membrane-bound form of the receptor, with the order of potencies for displacement of the selective σ-ligand [3H]di-o-tolylguanidine ([3H]DTG) closely correlated. The stereoselectivity for (+)-benzomorphan opiate enantiomers was retained by the solubilized receptor. The soluble receptor retained high affinity for binding of [3H]DTG (KD= 28 ± 0.5 nM) and (+)-[3H]3-(3-hydroxyphenyl)-N-(l-propyl)piperi-dine {(+)-[3H]3-PPP} (KD= 36 ± 2 nM). Photoaffinity labeling of the solubilized receptor by [3H]p-azido-DTG, a σ-selective photoaffinity label, resulted in labeling of a 29-kilodalton polypeptide identical in size to that labeled in intact membranes. Estimation of the Stokes radius of the [3H]DTG binding site was obtained by Sepharose CL-6B chromatography in the presence of 20 mM cholate and calculated to be 8.7 nm. This value was identical to the molecular size found for the binding sites of the σ-selective ligands (+)-[3H]3-PPP and (+)-[3H]SKF-10,047, supporting the hypothesis that all three ligands bind to the same macro-molecular complex.

Item Type: Article
ISSNs: 0022-3042 (print)
1471-4159 (electronic)
Subjects: Q Science > QH Natural history > QH301 Biology
Divisions: University Structure - Pre August 2011 > School of Biological Sciences
ePrint ID: 143689
Date Deposited: 21 Jul 2010 10:47
Last Modified: 27 Mar 2014 19:05
Publisher: LIPPINCOTT-RAVEN PUBL
URI: http://eprints.soton.ac.uk/id/eprint/143689

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