Solubilization of the acetylcholine receptor protein from Loligo opalescens without detergents
1974; Wiley; Volume: 48; Issue: 1 Linguagem: Inglês
10.1016/0014-5793(74)81054-9
ISSN1873-3468
Autores Tópico(s)Nicotinic Acetylcholine Receptors Study
ResumoWe [I] recently showed that the acetylcholine (ACh) receptor isolated from the optic ganglia of squid (Lo&o opalescens) is similar to the ACh nicotine receptor of electric organs of Electrophonrs electricus [2], Torpedo marmorata [3] and of vertebrate skeletal muscle [4]. The assay for the receptor in crude extracts of optic ganglia was based on the ability of the receptor site to bind [I4 C] decamethonium, [I4 C] -dtubocurarine and [ ’ ” I] o-bungarotoxin and for the capacity of the antagonists gallamine, d-tubocurarine and a-bungarotoxin to displace [ r4 C] decamethonium binding [l] . It is generally believed [ 51 that the ACh receptor is strongly membrane bound and that it can be solubilized only by treatment of membrane extracts containing the receptor with mild detergents such as Triton X-100 [6], sodium deoxycholate [7] or lubrol-WX [3,8]. Several laboratories [7,9-121 have succeeded in purifying the detergent solubilized receptor and have determined its molecular weight using different physical methods. In our attempts to isolate membrane fragments rich in ACh receptors from optic ganglia of Loligo opalescens [l] we observed that the receptor protein could be solubilized without using detergents. In this paper we show that in salt solution (0.1 M NaC 1, 10 mM phosphate buffer, pH 7.8) the [r2’ I]obungarotoxin labeled receptor protein of Loligo opalescens has a mol. wt. of 240 000+20 000 when determined by gel filtration on Bio-Gel A-5m or by sucrose gradient centrifugation. Sodium dodecyl sulphate disc gel electrophoresis indicates that the
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