标题：Time-resolved photolabeling by the noncompetitive blocker chlorpromazine of the acetylcholine receptor in its transiently open and closed ion channel conformations
期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：1984
卷号：81
期号：6
页码：1897-1901
DOI：10.1073/pnas.81.6.1897
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：A rapid-mixing photolabeling apparatus is developed to resolve the kinetics of association of the noncompetitive channel blocker [3H]chlorpromazine (CPZ) with the membrane-bound acetylcholine (AcCho) receptor from Torpedo marmorata and to photolabel its subunits in the 100-milli-seconds to seconds time range. Rapid mixing of AcCho and [3H]CPZ with the receptor followed by brief (less than 20 msec) UV irradiation results in the selective labeling of the four chains of the AcCho receptor, according to a rapid bimolecular association process close to diffusion-controlled. Rapid association is not observed with the competitive antagonists d-tubocurarine or flaxedil or the snake venom alpha-toxins. Its initial rate increases with agonist concentration, with maxima of 0.6 for carbamoylcholine and 0.2 for phenyltrimethylammonium taking 1 for AcCho, with apparent dissociation constants of 30 microM, 400 microM, and 300 microM for AcCho, carbamoylcholine, and phenyltrimethylammonium, respectively, and with sigmoid shape (Hill coefficients of 1.1-1.3). Under conditions in which the receptor "desensitizes" and the ionic channel closes (preincubation with AcCho), rapid [3H]CPZ association decreases in parallel. It is concluded that the agonist-dependent rapid association of [3H]CPZ takes place at the level of a site common to all five subunits, which lies within the ion channel and becomes accessible when the channel opens.
