SIZE-EXCLUSION CHROMATOGRAPHIC RECONSTITUTION OF THE BOVINE BRAIN BENZODIAZEPINE RECEPTOR - EFFECTS OF LIPID ENVIRONMENT ON THE BINDING CHARACTERISTICS
Gt. Viel et al., SIZE-EXCLUSION CHROMATOGRAPHIC RECONSTITUTION OF THE BOVINE BRAIN BENZODIAZEPINE RECEPTOR - EFFECTS OF LIPID ENVIRONMENT ON THE BINDING CHARACTERISTICS, Journal of chromatography, 776(1), 1997, pp. 101-107
Citations number
25
Categorie Soggetti
Chemistry Analytical","Biochemical Research Methods
The benzodiazepine receptor from calf brain was solubilized with sodiu
m deoxycholate (2 mg/ml) in the presence of 0.5 M KCl and protease inh
ibitors, and bound flunitrazepam with an equilibrium dissociation cons
tant (K-d) of 2.7+/-1.2 nM and with 0.40+/-0.04 pmol binding sites per
mg protein (B-max). Up to 60% of the benzodiazepine binding sites (av
erage 25%) could be reconstituted in lipid vesicles, upon size-exclusi
on chromatography of protein-detergent-lipid mixtures on Sephadex G-50
Medium for detergent depletion. The flunitrazepam affinity for the re
constituted receptor varied with the lipid composition (K-d 1.4-4 nM),
Freezing and thawing increased the size of the small proteoliposomes
obtained by chromatographic reconstitution and, on the average, double
d the number of operative flunitrazepam binding sites. When the proteo
liposomes were stored at -20 degrees C or -80 degrees C or in lyophili
zed state, the receptor retained its benzodiazepine binding affinity a
nd B-max over a period of 2 months. (C) 1997 Elsevier Science B.V.