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Characterization of Curaremimetic Neurotoxin Binding Sites on Cellular Membrane Fragments Derived from the Rat Pheochromocytoma PC 12
Author(s) -
Lukas Ronald J.
Publication year - 1986
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.1471-4159.1986.tb13087.x
Subject(s) - dithiothreitol , acetylcholine receptor , nicotinic agonist , neurotoxin , binding site , biochemistry , receptor , chemistry , tetranitromethane , nicotinic acetylcholine receptor , dissociation constant , biology , biophysics , microbiology and biotechnology , tyrosine , enzyme
Studies were conducted on the properties of 125 I‐labeled α‐bungarotoxin binding sites on cellular membrane fragments derived from the PC 12 rat pheochromocytoma. Two classes of specific toxin binding sites are present at approximately equal densities (50 fmol/mg of membrane protein) and are characterized by apparent dissociation constants of 3 and 60 n M . Nicotine and d ‐tubocurarine are among the most potent inhibitors of high‐affinity toxin binding. The affinity of high‐affinity toxin binding sites for nicotinic cholinergic agonists is reversibly or irreversibly decreased, respectively, on treatment with dithiothreitol or dithiothreitol and N ‐ethylmaleimide. The nicotinic receptor affinity reagent bromoacetylcholine irreversibly blocks high‐affinity toxin binding to PC 12 cell membranes that have been treated with dithiothreitol. Two polyclonal antisera raised against the nicotinic acetylcholine receptor from Electrophorus electricus inhibit high‐affinity toxin binding. These detailed studies confirm that curaremimetic neurotoxin binding sites on the PC 12 cell line are comparable to toxin binding sites from neural tissues and to nicotinic acetylcholine receptors from the periphery. Because toxin binding sites are recognized by anti‐nicotinic receptor antibodies, the possibility remains that they are functionally analogous to nicotinic receptors.