Asymmetric Mutations in Selectivity Filter of K+ Channel Pore Generate C-type Inactivation

Abstract

It is well established that activation of Kv channels results from voltage-dependent conformational changes within their voltage sensing domains (VSD), which are transferred to the intracellular activation gate. Some Kv channels, such as Kv.1.3 undergo a subsequent inactivation, which is caused by conformational rearrangements inside the selectivity filter. This process has been extensively studied but the molecular causes for the so-called C-type inactivation are not fully understood. In order to test whether asymmetries in the selectivity filter are part of C-type inactivation we constructed two-tandem concatemers from small viral K+ channel pores. The latter are good model systems for the pore module of complex K+ channels. Here we describe a mutation within the selectivity filter that introduces C-type-like inactivation in this channel. By using the K+ channel blocker Ba2+ in the bath to mimic the voltage dependency of a Kv channel we can reconstruct an outward current that is undistinguishable from the gating kinetics of Kv channels: depolarization-induced dissociation of Ba2+, which mimics the action of the VSD in Kv channels, leads to a fast channel activation followed by a pronounced and fast voltage dependent inactivation. Interestingly, the introduced mutation (G48C) is located at the first glycine of the TVGYG signature sequence e.g. at a position within the selectivity filter considered to be of particular importance for C-type inactivation in other K+ channels. We show that strength and voltage dependency of this C-type inactivation can be modulated by the number of mutations within the concatemers and by the type of amino acid substitution. All this renders these Kcv-concatemers an outstanding model system for the investigation of C-type inactivation and for understanding the role of asymmetry in the selectivity filter in this process.