Department of Neurobiology, Harvard Medical School, Boston, MA 02115, USA.
J Gen Physiol. 2012 Nov;140(5):469-79. doi: 10.1085/jgp.201210850. Epub 2012 Oct 15.
HCN (hyperpolarization-activated cyclic nucleotide gated) pacemaker channels have an architecture similar to that of voltage-gated K(+) channels, but they open with the opposite voltage dependence. HCN channels use essentially the same positively charged voltage sensors and intracellular activation gates as K(+) channels, but apparently these two components are coupled differently. In this study, we examine the energetics of coupling between the voltage sensor and the pore by using cysteine mutant channels for which low concentrations of Cd(2+) ions freeze the open-closed gating machinery but still allow the sensors to move. We were able to lock mutant channels either into open or into closed states by the application of Cd(2+) and measure the effect on voltage sensor movement. Cd(2+) did not immobilize the gating charge, as expected for strict coupling, but rather it produced shifts in the voltage dependence of voltage sensor charge movement, consistent with its effect of confining transitions to either closed or open states. From the magnitude of the Cd(2+)-induced shifts, we estimate that each voltage sensor produces a roughly three- to sevenfold effect on the open-closed equilibrium, corresponding to a coupling energy of ∼1.3-2 kT per sensor. Such coupling is not only opposite in sign to the coupling in K(+) channels, but also much weaker.
HCN（超极化激活环核苷酸门控）起搏通道的结构与电压门控 K(+)通道相似，但它们的开启具有相反的电压依赖性。HCN 通道使用与 K(+)通道基本相同的带正电荷的电压传感器和细胞内激活门，但显然这两个组件的耦合方式不同。在这项研究中，我们通过使用半胱氨酸突变通道来研究电压传感器和孔之间的耦合能，对于这些通道，低浓度的 Cd(2+)离子可以冻结开-关门控机制，但仍允许传感器移动。我们能够通过施加 Cd(2+)将突变通道锁定在打开或关闭状态，并测量对电压传感器运动的影响。Cd(2+)没有固定门控电荷，这与严格的耦合一致，但它确实会改变电压传感器电荷运动的电压依赖性，这与其将跃迁限制在关闭或打开状态的效果一致。根据 Cd(2+)诱导的位移幅度，我们估计每个电压传感器对开-关平衡产生约三到七倍的影响，对应于每个传感器约 1.3-2 kT 的耦合能。这种耦合不仅与 K(+)通道中的耦合符号相反，而且也弱得多。
