Mechanism of potassium-channel selectivity revealed by Na+ and Li+ binding sites within the KcsA pore

Abstract

Potassium channels allow K+ ions to easily diffuse through their pores while effectively preventing smaller Na+ ions from permeation. The ability to discriminate between these two similar and abundant ions is vital for these proteins to control electrical and chemical activity in all organisms. This selection process occurs at the narrow selectivity filter that contains structurally identified K+ binding-sites. Selectivity is thought to arise because smaller ions such as Na+ do not bind to these K+ sites in a thermodynamically favorable way. Using the model K+ channel KcsA, we examined how intracellular Na+ and Li+ interact with the pore and the permeant ions using electrophysiology, molecular dynamics simulations, and X-ray crystallography. Our results suggest that these small cations have a binding site within the K+ selectivity filter, albeit different from the K+ sites. We propose that selective permeation from the intracellular side is achieved mainly by a large energy barrier blocking filter entry for Na+ and Li+ in the presence of K+, and not by a difference of binding affinity between ions inside the selectivity filter.

On the selective ion binding hypothesis for potassium channels

“…[22][23][24]. These studies showed that the cage-of-oxygen sites provide selective binding for K þ over Na þ , by up to 5 kcal∕mol, reinforcing the view that K þ channels exclude Na þ ions due to differing thermodynamic stabilities in the crystallographic sites.…”

“…Perhaps higher barriers for Na þ may arise due to the different multi-ion configurations for Na þ or K þ -Na þ mixtures. We previously showed, for the case of ion entry from the intracellular side, that the presence of Na þ , with different (by ∼1 Å) binding site locations, disrupts the optimized knock-on conduction mechanism of KcsA, leading to large permeation barriers (24). We suggest that such an elevated barrier for Na þ may also exist for entry from the external solution.…”

“…This planar arrangement of carbonyls has been shown to yield close packing around Na þ , whereas the cage arrangement leads to a seemingly unfavorable partial-coordination by the eight ligands (e.g., refs. 24,27). We demonstrate here that such planar sites exist between all crystallographic K þ sites, and that each exhibits favorable binding for Na þ .…”

“…ions actually bind (23)(24)(25)(26)(27)(28)(29)(30), owing to the preference for the smaller Na þ ion to bind to fewer ligands (31,32). We have previously shown that the S4 and S3 K þ sites are separated by a planeof-carbonyl oxygen atoms site for Na þ ions, where Na þ ions have been proposed to cause slow blocking from the intracellular side (24) (labeled B34 in Fig.…”

Protein interactions central to stabilizing the K + channel selectivity filter in a four-sited configuration for selective K + permeation

“…Although the availability of a number of high-resolution structures of K þ channels has greatly aided our understanding of the molecular details of ion binding in this highly conserved structure (3)(4)(5)(6), the underlying mechanism of K þ selectivity remains elusive (7)(8)(9)(10)(11)(12). The structural studies of K þ channels seem to favor the classical snug-fit model to account for K þ over Na þ selectivity (13,14), while computational studies on K þ channel selectivity invoke a number of different concepts in explaining K þ selectivity (15)(16)(17)(18)(19)(20)(21)(22)(23)(24).…”