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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Two atomic constraints unambiguously position the S4 segment relative to S1 and S2 segments in the closed state of Shaker K channel
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Two atomic constraints unambiguously position the S4 segment relative to S1 and S2 segments in the closed state of Shaker K channel

机译：在Shaker K通道处于关闭状态时，两个原子约束明确地将S4段相对于S1和S2段定位

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摘要

It is now well established that the voltage-sensing S4 segment in voltage-dependent ion channels undergoes a conformational change in response to varying membrane potential. However, the magnitude of the movement of S4 relative to the membrane and the rest of the protein remains controversial. Here, by using histidine scanning mutagenesis in the Shaker K channel, we identified mutants I241H (S1 segment) and I287H (S2 segment) that generate inward currents at hyperpolarized potentials, suggesting that these residues are part of a hydrophobic plug that separates the water-accessible crevices. Additional experiments with substituted cysteine residues showed that, at hyperpolarized potentials, both I241C and I287C can spontaneously form disulphide and metal bridges with R362C, the position of the first charge-carrying residue in S4. These results constrain unambiguously the closed-state positions of the S4 segment with respect to the S1 and S2 segments, which are known to undergo little or no movement during gating. To satisfy these constraints, the S4 segment must undergo an axial rotation of ≈ 180° and a transmembrane (vertical) movement of ≈ 6.5 A at the level of R362 in going from the open to the closed state of the channel, moving the gating charge across a focused electric field.
机译：现在已经很好地确定，电压依赖性离子通道中的电压感测S4片段响应于膜电位的变化而发生构象变化。然而，S4相对于膜和其余蛋白质的运动幅度仍然存在争议。在这里，通过在Shaker K通道中使用组氨酸扫描诱变，我们鉴定了突变体I241H（S1区段）和I287H（S2区段），它们在超极化电势下产生内向电流，表明这些残基是将水与水分离的疏水性塞子的一部分。可访问的缝隙。用取代的半胱氨酸残基进行的其他实验表明，在超极化电势下，I241C和I287C均可与R362C（S4中第一个载电荷残基的位置）自发形成二硫键和金属桥。这些结果明确地限制了S4段相对于S1段和S2段的关闭状态位置，已知这些位置在选通过程中几乎没有移动。为了满足这些约束，从通道的打开状态到关闭状态，S4段在R362的水平上必须经历≈180°的轴向旋转和≈6.5 A的跨膜（垂直）运动，从而使选通电荷移动跨过聚焦电场。

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