Potassium (K+) channels allow rapid, highly selective and finely controlled diffusion of potassium ions across cell membranes. Control over K+ flux occurs by channels rapidly switching between permissive and restrictive states in a process known as ‘gating’. It is conventionally thought that K+ channels gate by undergoing a conformational change. Conductive channels are thought to contain a wide inner pore mouth, whereas closed channels contain a constricted mouth, such that this constriction sterically occludes the flow of fully hydrated K+ ions.
In this study, we investigated the extent of this conformational change required for conduction and gating in the KIR family of potassium channels. By covalently linking adjacent subunits together, movement at the pore mouth could be constrained, locking the channel into its narrow or ‘closed’ state. Crosslinking was inspected by a variety of biophysical techniques, including X-ray crystallography, SDS-PAGE and native mass spectrometry. The functional capacity of ‘locked’ relative to ‘non-locked’ channels was then assessed by means of a bulk fluorometric liposomal flux assay.
Results indicated that channels with a limited narrow pore aperture were able to function as effectively as wildtype channels, suggesting that in KIR channels, gating is inconsistent with a conformational change in the pore. These experimental findings, supported by molecular dynamics simulations, suggest that K+ ions transit through the narrow pore mouth in a partially hydrated state. The study opens up new perspectives on the mechanisms underlying conduction and gating in K+ channels and particularly highlights the importance of variations in K+ hydration during conduction.