Mechanism for selectivity-inactivation coupling in KcsA potassium channels

Structures of the prokaryotic K⁺ channel, KcsA, highlight the role of the selectivity filter carbonyls from the GYG signature sequence in determining a highly selective pore, but channels displaying this sequence vary widely in their cation selectivity. Furthermore, variable selectivity can be found within the same channel during a process called C-type inactivation. We investigated the mechanism for changes in selectivity associated with inactivation in a model K⁺ channel, KcsA. We found that E71A, a noninactivating KcsA mutant in which a hydrogen-bond behind the selectivity filter is disrupted, also displays decreased K⁺ selectivity. In E71A channels, Na⁺ permeates at higher rates as seen with ⁸⁶Rb⁺ and ²²Na⁺ flux measurements and analysis of intracellular Na⁺ block. Crystal structures of E71A reveal that the selectivity filter no longer assumes the "collapsed," presumed inactivated, conformation in low K⁺, but a "flipped" conformation, that is also observed in high K⁺, high Na⁺, and even Na⁺ only conditions. The data reveal the importance of the 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.