Department of Pharmacology and Physiology, Université de Montréal, Montréal, QC H3C 3J7, Canada.
Department of Pharmacology and Physiology, Université de Montréal, Montréal, QC H3C 3J7, Canada;
Proc Natl Acad Sci U S A. 2018 Jul 17;115(29):E6751-E6759. doi: 10.1073/pnas.1719105115. Epub 2018 Jun 29.
The S4-S5 linker physically links voltage sensor and pore domain in voltage-gated ion channels and is essential for electromechanical coupling between both domains. Little dynamic information is available on the movement of the cytosolic S4-S5 linker due to lack of a direct electrical or optical readout. To understand the movements of the gating machinery during activation and inactivation, we incorporated fluorescent unnatural amino acids at four positions along the linker of the Shaker K channel. Using two-color voltage-clamp fluorometry, we compared S4-S5 linker movements with charge displacement, S4 movement, and pore opening. We found that the proximal S4-S5 linker moves with the S4 helix throughout the gating process, whereas the distal portion undergoes a separate motion related to late gating transitions. Both pore and S4-S5 linker undergo rearrangements during C-type inactivation. In presence of accelerated C-type inactivation, the energetic coupling between movement of the distal S4-S5 linker and pore opening disappears.
S4-S5 连接段在电压门控离子通道中将电压传感器和孔域物理连接起来，对于这两个域之间的机电耦联是必不可少的。由于缺乏直接的电或光读出，因此关于细胞溶质 S4-S5 连接段的运动，仅有很少的动态信息。为了理解在激活和失活过程中门控机制的运动，我们在 Shaker K 通道的连接段的四个位置掺入了荧光非天然氨基酸。使用双色电压钳荧光法，我们将 S4-S5 连接段的运动与电荷位移、S4 运动和孔道开放进行了比较。我们发现，在整个门控过程中，近端 S4-S5 连接段与 S4 螺旋一起移动，而远端部分则发生与晚期门控转变相关的单独运动。在 C 型失活过程中，孔道和 S4-S5 连接段都发生了重排。在加速的 C 型失活存在的情况下，远端 S4-S5 连接段的运动与孔道开放之间的能量偶联消失了。
