Mechanism for Selectivity-Inactivation Coupling in KcsA Potassium Channels

Potassium channels containing the GYG motif often diverge in their selectivity for monovalent cations, but the molecular basis is unknown. Using the prokaryotic potassium channel KcsA as our model, we have investigated the role of the interaction between glutamate 71 and aspartate 80, located behind the selectivity filter, in determining the selectivity of the channel as well as its influence on the conformation of the filter. In E71A KcsA channels, Na+ permeates at higher rates in both the presence and absence of K+, as seen with 86Rb+ and 22Na+ flux measurements. Single channel recordings indicate that Na+ “punches through” E71A KcsA channels at lower voltages than wild-type KcsA, and in the punchthrough regime the Na+-blocked current appears significantly larger. A crystal structure of E71A KcsA reveals that in contrast to what was seen for wild type KcsA, the selectivity filter does not collapse in the absence of K+, but instead assumes a “flipped” conformation. This flipped conformation is the same one observed in previous E71A KcsA structures in the presence of K+. The data reveal the importance of this E71-D80 interaction in both favoring inactivation and maintaining high K+ selectivity. We propose a molecular mechanism by which inactivation and K+ selectivity are linked, a mechanism that may also be at work in other channels containing the canonical GYG signature sequence.