Abstract
Membrane-bound α-bungarotoxin-binding entities derived from rat brain are found to interact specifically with the affinity reagents maleimidobenzyltrimethylammonium (MBTA) and bromoacetylcholine (BAC), originally designed to label nicotinic acetylcholine receptors from electroplax and skeletal muscle. Following treatment of membranes with dithiothreitol, all specffic toxin binding sites are irreversibly blocked by reaction with MBTA or BAC. Affinity reagent labeling of dithiothreitol-reduced membranes is prevented (toxin binding sites are not blocked) by prior alkylation with N-ethylmaleimide, by prior oxidation with dithiobis(2-nitrobenzoic acid), or by incubation with neurotoxin. Reversibly associating cholinergic agonists and antagonists retard the rate of affinity reagent interaction with toxin receptors. The apparent rates of affinity reagent alkylation of toxin receptors, and the influences of other sulfhydryl/disulfide reagents on affinity labeling are comparable to those observed for reaction with nicotinic acetylcholine receptors in the periphery. The results provide further evidence that central nervous system α-bungarotoxin receptors share a remarkable number of biochemical properties with nicotinic receptors from the periphery.
ACKNOWLEDGMENTS The authors thank personnel in the Technical Information Division of the Lawrence Berkeley Laboratory for secretarial assistance, and express sincere gratitude to Dr. Mark G. McNamee, University of California at Davis, for supplying details of BAC synthesis prior to their publication, for his generous gifts of MBTA, and for valuable advice.
- Copyright © 1980 by The American Society for Pharmacology and Experimental Therapeutics
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