Berdiev B K, Karlson K H, Jovov B, Ripoll P J, Morris R, Loffing-Cueni D, Halpin P, Stanton B A, Kleyman T R, Ismailov I I
Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, Alabama 35294-0005, USA.
Biophys J. 1998 Nov;75(5):2292-301. doi: 10.1016/S0006-3495(98)77673-8.
The molecular composition of a core conduction element formed by the alpha-subunit of cloned epithelial Na+ channels (ENaC) was studied in planar lipid bilayers. Two pairs of in vitro translated proteins were employed in combinatorial experiments: 1) wild-type (WT) and an N-terminally truncated alphaDeltaN-rENaC that displays accelerated kinetics (tauo = 32 +/- 13 ms, tauc = 42 +/- 11 ms), as compared with the WT channel (tauc1 = 18 +/- 8 ms, tauc2 = 252 +/- 31 ms, and tauo = 157 +/- 43 ms); and 2) WT and an amiloride binding mutant, alphaDelta278-283-rENaC. The channels that formed in a alphaWT:alphaDeltaN mixture fell into two groups: one with tauo and tauc that corresponded to those exhibited by the alphaDeltaN-rENaC alone, and another with a double-exponentially distributed closed time and a single-exponentially distributed open time that corresponded to the alphaWT-rENaC alone. Five channel subtypes with distinct sensitivities to amiloride were found in a 1alphaWT:1alphaDelta278-283 protein mixture. Statistical analyses of the distributions of channel phenotypes observed for either set of the WT:mutant combinations suggest a tetrameric organization of alpha-subunits as a minimal model for the core conduction element in ENaCs.
在平面脂质双分子层中研究了由克隆的上皮钠通道（ENaC）的α亚基形成的核心传导元件的分子组成。在组合实验中使用了两对体外翻译的蛋白质：1）野生型（WT）和N端截短的αDeltaN-rENaC，与WT通道（tauc1 = 18 +/- 8毫秒，tauc2 = 252 +/- 31毫秒，tauo = 157 +/- 43毫秒）相比，其显示出加速的动力学（tauo = 32 +/- 13毫秒，tauc = 42 +/- 11毫秒）；2）WT和氨氯地平结合突变体αDelta278-283-rENaC。在αWT:αDeltaN混合物中形成的通道分为两组：一组的tauo和tauc与单独的αDeltaN-rENaC表现出的相对应，另一组的关闭时间呈双指数分布且开放时间呈单指数分布，与单独的αWT-rENaC相对应。在1αWT:1αDelta278-283蛋白质混合物中发现了五种对氨氯地平敏感性不同的通道亚型。对WT:突变体组合的任一组所观察到的通道表型分布进行的统计分析表明，α亚基的四聚体组织是ENaCs中核心传导元件的最小模型。
