Alpha-bungarotoxin binding to a high molecular weight component from lower vertebrate brain identified on dodecyl sulfate protein-blots.
The binding of [125I]iodo-alpha-bungarotoxin [( 125]alpha-BuTX) to the dissociated alpha-subunit of Torpedo acetylcholine receptor (AChR) can be readily demonstrated in a modified 'protein-blot' analysis utilizing electrophoretically transferred, dissociated subunits immobilized onto positively charged nylon membranes which are then incubated directly with [125I]alpha-BuTX. We report here the use of the protein-blotting technique to detect the alpha-BuTX binding site present in the central nervous system of lower vertebrates and to characterize some of the physicochemical properties of the toxin binding site. High molecular weight (Mr greater than or equal to 200,000 and greater than or equal to 120,000) alpha-BuTX-binding components can be readily demonstrated in avian and fish brain extracts upon protein-blotting with [125I]alpha-BuTX following lithium dodecyl sulfate PAGE. Neither extensive reduction with dithiothreitol nor prior reduction followed by alkylation with iodoacetamide alter the mobility of the CNS-derived BuTX-binding sites. In contrast to our findings with Torpedo AChR or muscle AChR derived from a number of different species, no binding is observed in the molecular weight range of the alpha-subunit (Mr = 40,000) nor is any binding at any molecular weight observed in similar fractions prepared from adult, mammalian (rat, guinea pig) brain using this technique. These results demonstrate the existence in lower vertebrate brain of a BuTX binding site comparable in size to the AChR oligomeric complex of electric organ and muscle. They also suggest, however, striking structural differences between muscle AChR and the central neuronal BuTX-binding complex as well as a considerable difference between the neuronal BuTX-binding sites derived from lower and higher vertebrate brain.
D000818:Animals; D001921:Brain / Q000378:metabolism*; D002038:Bungarotoxins / Q000378:metabolism*; D055598:Chemical Phenomena; D002621:Chemistry; D004372:Ducks; D004557:Electric Organ / Q000378:metabolism; D004593:Electrophorus; D005399:Fishes; D006054:Goldfish; D006168:Guinea Pigs; D009132:Muscles / Q000378:metabolism; D051381:Rats; D011950:Receptors, Cholinergic / Q000378:metabolism*; D011978:Receptors, Nicotinic; D013045:Species Specificity; D014101:Torpedo; D064569:alpha7 Nicotinic Acetylcholine Receptor
D002038:Bungarotoxins; C579165:Chrna7 protein, rat; D011950:Receptors, Cholinergic; D011978:Receptors, Nicotinic; D064569:alpha7 Nicotinic Acetylcholine Receptor