Mechanisms of Fast Blockade and Gating of the MthK Channel Pore by Divalent Cations

Abstract

K+ channels can undergo inactivation, a process in which K+ conduction decays over time, contributing to tuning of electrical excitabilty in nerve and muscle. Although K+ inactivation is associated with conformational changes in the channel pore, these are likely to be preceded by interactions between the pore and surrounding ions. Here we use a series of divalent metal cations to systematically probe these interactions in MthK, a K+ channel of known structure, by electrophysiological methods and molecular dynamics simulations. Mg2+, Ca2+, and Sr2+ each yielded roughly equivalent levels of fast blockade, with similar voltage dependences (delta ∼0.2). In contrast, Ca2+ and Sr2+ were found to substantially enhanced voltage-dependent inactivation, whereas Mg2+ did not. The inactivation enhanced by Ca2+ or Sr2+ was not consistent with a simple voltage-dependent slow blockade, because increasing either [Ca2+] or [Sr2+ ] to 20 mM yielded little additional inhibition compared with the level produced at 5 mM, over the same voltage range. The differential effects of Ca2+/Sr2+ vs. Mg2+ on inactivation suggests that the slightly larger divalent cations enhance inactivation through a site that is separate from the one responsible for fast blockade. Molecular dynamics simulations and potential of mean force calculations suggest that Mg2+, Ca2+, and Sr2+ each can access a site in the wide cavity of the MthK pore (Scav) whereas Ca2+ or Sr2+ (but not Mg2+) are likely to further access a site at the entry to the selectivity filter (S5). We hypothesize that a divalent cation at S5 can affect a redistribution of K+ ions in the selectivity filter, which may in turn inhibit the conduction cycle and enhance inactivation.