Structural basis for the coupling between activation and inactivation gates in K+ channels

Abstract

Switching between conductive and non-conductive states is central to the function of ion channels. In potassium channels, inactivation gating occurs by two distinct molecular mechanisms: N-type inactivation (a rapid autoinhibitory process in which an N-terminal particle blocks conduction by binding to the open pore) and C-type inactivation (originating from conformational transitions at the selectivity filter). In the first of two papers, Eduardo Perozo and co-workers solve the X-ray crystal structure of the K+ channel KcsA in an 'open-inactivated' conformation together with a series of crystal structures of channels that are 'trapped' in a set of partially open conformations. In the second paper, the authors identify the underlying mechanism by which movements in the inner gate of this channel trigger conformational changes at the selectivity filter, leading to the non-conductive C-type inactivated state. K+ channels can convert between conductive and non-conductive forms through mechanisms that range from flicker transitions (which occur in microseconds) to C-type inactivation (which occurs on millisecond to second timescales). Here, the mechanisms are revealed through which movements of the inner gate of the K+ channel KcsA trigger conformational changes at the selectivity filter, leading to the non-conductive C-type inactivated state. The coupled interplay between activation and inactivation gating is a functional hallmark of K+ channels1,2. This coupling has been experimentally demonstrated through ion interaction effects3,4 and cysteine accessibility1, and is associated with a well defined boundary of energetically coupled residues2. The structure of the K+ channel KcsA in its fully open conformation, in addition to four other partial channel openings, richly illustrates the structural basis of activation–inactivation gating5. Here, we identify the mechanistic principles by which movements on the inner bundle gate trigger conformational changes at the selectivity filter, leading to the non-conductive C-type inactivated state. Analysis of a series of KcsA open structures suggests that, as a consequence of the hinge-bending and rotation of the TM2 helix, the aromatic ring of Phe 103 tilts towards residues Thr 74 and Thr 75 in the pore-helix and towards Ile 100 in the neighbouring subunit. This allows the network of hydrogen bonds among residues Trp 67, Glu 71 and Asp 80 to destabilize the selectivity filter6,7, allowing entry to its non-conductive conformation. Mutations at position 103 have a size-dependent effect on gating kinetics: small side-chain substitutions F103A and F103C severely impair inactivation kinetics, whereas larger side chains such as F103W have more subtle effects. This suggests that the allosteric coupling between the inner helical bundle and the selectivity filter might rely on straightforward mechanical deformation propagated through a network of steric contacts. Average interactions calculated from molecular dynamics simulations show favourable open-state interaction-energies between Phe 103 and the surrounding residues. We probed similar interactions in the Shaker K+ channel where inactivation was impaired in the mutant I470A. We propose that side-chain rearrangements at position 103 mechanically couple activation and inactivation in KcsA and a variety of other K+ channels.