Recovery from Slow Inactivation in K(+) Channels is Controlled by Water Molecules

The bacterial K(+) channel KcsA 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(1–7). Available X-ray structures of KcsA provide an atomic view of the four most important functional states in which the intracellular gate is either closed or open, and the selectivity filter is either conductive or inactivated(8–10). 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 toward 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). In this manuscript, a series of long molecular dynamics (MD) simulations reveal 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 waters and the rebinding of an external K(+) ion. A kinetic model deduced from the simulations shows how releasing the buried waters can stretch the timescale of recovery to seconds. This leads to the prediction that reducing the occupancy of the buried waters 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 2M sucrose.