Molecular Dynamics Simulations Describe the Mechanism of K Block in Bacterial Nav Channels

Abstract

Although extensive electrophysiological characterization is available for eukaryotic voltage-gated Na+ channels (Nav), no high-resolution structures of these channels are available. Crystal structures of several bacterial Nav channels have been published and molecular dynamics simulations of ion permeation through these channels are consistent with many electrophysiological properties of the eukaryotic channels. Unlike eukaryotic Nav channels, however, the bacterial Nav channels are strongly outwardly rectifying, and the mechanism of this rectification has not previously been described. We used step-wise pulling protocols to implement Jarzynski's Equality in non-equilibrium molecular dynamics simulations of ion permeation through the bacterial NavAb channel to obtain a mechanistic description of this outward rectification. Results of the simulations indicate that two or three extracellular K+ ions bind tightly at the same z-coordinate along the selectivity filter of NavAb and can effectively block the channel in the presence of modest voltages or concentration driving forces. The configuration with two potassium ions located at the same z-coordinate is also found in the two-dimensional potential of mean forces generated from umbrella sampling and weighted histogram analysis. In contrast to K+, three Na+ ions move through the selectivity filter together as a unit in a “knock-on” mechanism of permeation. Differences in the amount of work required to move three Na+ ions through the selectivity filter of NavAb compared to three K+ ions predict the large negative reversal potentials observed for bacterial Nav channels in instantaneous current-voltage plots. The results of the simulations suggest that the block of bacterial voltage-gated Na+ channels by extracellular K+ does not occur in eukaryotic voltage-gated Na+ channels because of differences in the amino acids present in the selectivity filters of the different channels.