Scanning the intracellular S6 activation gate in the shaker K+ channel.
Scanning the intracellular S6 activation gate in the shaker K+ channel.
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DOI：
10.1085/jgp.20028569
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发表时间：
2002-06
影响因子：
3.8
通讯作者：
Swartz, Kenton J
中科院分区：
文献类型：
--
作者：
Hackos, David H;Chang, Tsg-Hui;Swartz, Kenton J
在Kv通道中，激活门被认为位于离子传导孔的细胞内入口附近。尽管S6片段的COOH末端与gate结构的形成有关，但位于gate遮挡部分的残基仍未确定。我们在Shaker Kv通道中使用诱变扫描方法，将S6门区（T469-Y485）中的每个残基突变为丙氨酸、色氨酸和天冬氨酸，以确定对突变不敏感的位置，并找到破坏门的突变体。大多数突变体以电压依赖的方式打开，并在负电压下有效关闭，这表明栅极结构既可以在打开时支持离子通量，也可以在关闭时阻止离子通量。我们发现了几个突变通道，其中宏观离子电流非常小或无法检测到，而一个突变通道在负电压下显示本构电流。对这三种类型的取代的综合研究支持了S6的胞内部分形成激活门的观点，并确定了V478和F481作为封闭状态下孔隙封闭的候选物质。
In Kv channels, an activation gate is thought to be located near the intracellular entrance to the ion conduction pore. Although the COOH terminus of the S6 segment has been implicated in forming the gate structure, the residues positioned at the occluding part of the gate remain undetermined. We use a mutagenic scanning approach in the Shaker Kv channel, mutating each residue in the S6 gate region (T469-Y485) to alanine, tryptophan, and aspartate to identify positions that are insensitive to mutation and to find mutants that disrupt the gate. Most mutants open in a steeply voltage-dependent manner and close effectively at negative voltages, indicating that the gate structure can both support ion flux when open and prevent it when closed. We find several mutant channels where macroscopic ionic currents are either very small or undetectable, and one mutant that displays constitutive currents at negative voltages. Collective examination of the three types of substitutions support the notion that the intracellular portion of S6 forms an activation gate and identifies V478 and F481 as candidates for occlusion of the pore in the closed state.