Recovery from slow inactivation in K~+ channels is controlled by water molecules

摘要

当一个刺激造成一个离子通道的门被打开后， 这个通道将一直让离子通过，直到其选择性过 滤器关闭。之后有一个恢复阶段，涉及被关闭 的通道从失活状态向导通状态的构形转变。细 菌K*通道KcsA为研究该通道的慢速(C-型）失 活和从失活状态的恢复的发生机制提供了一个 极好的模型系统。在这篇文章中，作者通过一 系列长期分子动态模拟发现，选择性过滤器被 结合在该选择性过滤器后面的埋没的水分子 以空间立体方式锁定于失活构形。从以上模拟 推导出的一个动力模型，显示了释放埋没的水rn分子何以能延长恢复期的时间尺度，同时作 者还通过"湿"生物物理实验对该模型进行了 研究。%Application of a specific stimulus opens the intracellular gate of a K~+ channel (activation), yielding a transient period of ion conduction until the selectivity filter spontaneously undergoes a conformational change towards a non-conductive state (inactivation). Removal of the stimulus closes the gate and allows the selectivity filter to interconvert back to its conductive conformation (recovery). Given that the structural differences between the conductive and inactivated filter are very small, it is unclear why the recovery process can take up to several seconds. The bacterial K~+ channel KcsA from Streptomyces lividans can be used to help elucidate questions about channel inactivation and recovery at the atomic level. Although KcsA contains only a pore domain, without voltage-sensing machinery, it has the structural elements necessary for ion conduction, activation and inactivation. Here we reveal, by means of a series of long molecular dynamics simulations, how the selectivity filter is sterically locked in the inactive conformation by buried water molecules bound behind the selectivity filter. Potential of mean force calculations show how the recovery process is affected by the buried water molecules and the rebinding of an external K~+ ion. A kinetic model deduced from the simulations shows how releasing the buried water molecules can stretch the timescale of recovery to seconds. This leads to the prediction that reducing the occupancy of the buried water molecules by imposing a high osmotic stress should accelerate the rate of recovery, which was verified experimentally by measuring the recovery rate in the presence of a 2-molar sucrose concentration.