Potassium channel selectivity filter dynamics revealed by single-molecule FRET

Potassium (K) channels exhibit exquisite selectivity for conduction of K ⁺ ions over other cations, particularly Na ⁺ . High-resolution structures reveal an archetypal selectivity filter (SF) conformation in which dehydrated K ⁺ ions, but not Na ⁺ ions, are perfectly coordinated. Using single-molecule FRET (smFRET), we show that the SF-forming loop (SF-loop) in KirBac1.1 transitions between constrained and dilated conformations as a function of ion concentration. The constrained conformation, essential for selective K ⁺ permeability, is stabilized by K ⁺ but not Na ⁺ ions. Mutations that render channels nonselective result in dilated and dynamically unstable conformations, independent of the permeant ion. Further, while wild-type KirBac1.1 channels are K ⁺ selective in physiological conditions, Na ⁺ permeates in the absence of K ⁺ . Moreover, whereas K ⁺ gradients preferentially support ⁸⁶ Rb ⁺ fluxes, Na ⁺ gradients preferentially support ²² Na ⁺ fluxes. This suggests differential ion selectivity in constrained versus dilated states, potentially providing a structural basis for this anomalous mole fraction effect.