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Glycoprotein as a Constituent of Purified γ‐Aminobutyric Acid/Benzodiazepine Receptor Complex: Structures and Physiological Roles of Its Carbohydrate Chain
Author(s) -
Kuriyama Kinya,
Taguchi Junichi
Publication year - 1987
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1987.tb05753.x
Subject(s) - muscimol , chemistry , biochemistry , receptor , gabaa receptor
The effect of treatments with various enzymes and chemically modifying agents on [ 3 H]muscimol binding to a purified γ‐aminobutyric acid (GABA)/benzodiazepine receptor complex from the bovine cerebral cortex was examined. Treatments with pronase, trypsin, guanidine hydro‐chloride, and urea significantly decreased the binding of [ 3 H]muscimol, but dithiothreitol, N ‐ethylmaleimide, reduced glutathione, oxidized glutathione, cysteine, and cys‐tine had no significant effect. These results indicate that the GABA receptor indeed consists of protein, but SH and S‐S‐groups in the protein are not involved in the exhibition of the binding activity. On the other hand, column chromatography using concanavalin A‐Sepharose eluted protein having [H]muscimol binding activity and staining of glycoprotein using an electrophoresed slab gel indicated the existence of two bands originating from the subunits of the GABA/benzodiazepine receptor complex. Furthermore, treatments with various glycosidases such as glyco‐peptidase A, β‐galactosidase, and α‐mannosidase significantly increased the binding of [ 3 H]muscimol. These results strongly suggest that GABA/benzodiazepine receptor complex is a glycoprotein and that its carbohydrate chain may be a hybrid type. Treatment with β‐galactosidase resulted in the disappearance of the low‐affinity site for [ 3 H]muscimol binding and in an increase of B max of the high‐affinity site, without changing the K D value. These results suggest that the carbohydrate chain in the receptor complex may have a role in exhibiting the low‐affinity binding site for GABA. The observation that the enhancement of [ 3 H]muscimol binding by treatments with β‐galactosidase and glycopeptidase A were much higher than that with α‐mannosidase may also indicate a special importance of the β‐galactosyl residue in the inhibition of GABA receptor binding activity. Furthermore, the observation that the activation of high‐affinity [ 3 H]muscimol binding by benzodiazepines disappeared following β‐galactosidase treatment suggests that the carbohydrate chain in the receptor complex may also be involved in the functional coupling between the GABA receptor and the benzodiazepine receptor.