BASIC MECHANISMS OF ION CHANNEL FUNCTION

该提案寻求继续就基本机制开展工作，通过这些机制 离子通道蛋白的工作原理，以及它们如何在 门控时尚与其潜在的分子结构有关。这个 本研究的重点是三种不同的离子通道：高密度的离子通道。 哺乳动物肌肉电导Ca~(2+)激活K~+通道A型K~+ 果蝇Shaker基因编码的通道和电压门控Cl- 从鱼雷加利福尼亚电风琴的频道。我们将调查 Ca~(2+)激活的K~+通道传导孔重构为平面 Ba2+离子阻断和相互作用对双层膜的影响 与多肽轮藻毒素，以及这一点的定点突变体 多肽。我们将尝试纯化和重建Shaker K+通道 在杆状病毒感染的昆虫细胞系中高水平表达后， 作为对此进行蛋白质水平生化分析的首次尝试 频道。最后，我们将完成对鱼雷氯-的提纯 频道为了启动对一位杰出成员的分子分析 电压依赖性阴离子通道家族。对……的普遍兴趣 该项目位于细胞内离子通道蛋白的中心位置 电信号的过程。研究人员现在才刚刚开始 一瞥这些重要蛋白质的分子结构，这就是 项目将为这一努力做出贡献。

This proposal seeks to continue work on the fundamental mechanisms by which ion channel proteins work, and on how their ability to transport ions in a gated fashion is related to their underlying molecular structures. The focus of this research is on three different ion channels: the high- conductance Ca2+-activated K+ channel from mammalian muscle, the A-type K+ channel encoded by the Shaker gene of Drosophila, and the voltage-gated Cl- channel from the electric organ of Torpedo californica. We will probe the conduction pore of the Ca2+-activated K+ channel reconstituted into planar bilayer membranes by examining blocking by Ba2+ ion and by the interaction with the peptide charybdotoxin, and of site-directed mutants of this peptide. We will attempt to purify and reconstitute the Shaker K+ channel after high-level expression in the baculovirus-infected insect cell line, as a first attempt to carry out protein-level biochemical analysis of this channel. Finally, we will complete the purification of the Torpedo Cl- channel in order to launch a molecular analysis of a prominent member of the family of voltage-dependent anion channels. The general interest in this project lies in the central place of ion channel proteins in the process of electrical signalling. Researchers are now just beginning to glimpse the molecular architecture of these important proteins, and this project will contribute to that effort.