Solution Structure and Function of the “Tandem Inactivation Domain” of the Neuronal A-type Potassium Channel Kv1.4

Ralph Wissmann,Wolfgang Bildl,Dominik Oliver,Michael Beyermann,Hans-Robert Kalbitzer,Detlef Bentrop,Bernd Fakler

doi:10.1074/jbc.m210191200

Abstract

Cumulative inactivation of voltage-gated (Kv) K(+) channels shapes the presynaptic action potential and determines timing and strength of synaptic transmission. Kv1.4 channels exhibit rapid "ball-and-chain"-type inactivation gating. Different from all other Kvalpha subunits, Kv1.4 harbors two inactivation domains at its N terminus. Here we report the solution structure and function of this "tandem inactivation domain" using NMR spectroscopy and patch clamp recordings. Inactivation domain 1 (ID1, residues 1-38) consists of a flexible N terminus anchored at a 5-turn helix, whereas ID2 (residues 40-50) is a 2.5-turn helix made up of small hydrophobic amino acids. Functional analysis suggests that only ID1 may work as a pore-occluding ball domain, whereas ID2 most likely acts as a "docking domain" that attaches ID1 to the cytoplasmic face of the channel. Deletion of ID2 slows inactivation considerably and largely impairs cumulative inactivation. Together, the concerted action of ID1 and ID2 may promote rapid inactivation of Kv1.4 that is crucial for the channel function in short term plasticity.

Highlights

Fast inactivation of voltage-gated Kϩ (Kv)1 channels is a major determinant for signal propagation and synaptic transmission in central nervous system neurons [1,2,3,4]
It remained obscure what the functional role of the second inactivation domains (ID) might be in intact channels and, how inactivation gating is realized by this tandem inactivation domain
NOEs between the ␣-proton of one residue and the amide (d␣N(i,iϩ3)) or ␤-proton (d␣␤(i,iϩ3)) of the third amino acid following were observed together with weak sequential contacts (d␣N(i,iϩ1)) and strong amide-amide NOEs (dNN(i,iϩ1)), a NOE pattern usually seen with ␣-helices

Summary

Introduction

Fast inactivation of voltage-gated Kϩ (Kv) channels is a major determinant for signal propagation and synaptic transmission in central nervous system neurons [1,2,3,4]. A-type Kϩ channels were shown to regulate orthograde and retrograde propagation of dendritic action potentials (APs) in CA1 pyramidal neurons and to dynamically control presynaptic Ca2ϩ inflow into hippocampal mossy fiber boutons [2, 3] In the latter, repetitive stimulation (frequencies of Ն10 Hz) led to a broadening of the APs because of cumulative inactivation of A-type Kϩ channels. The existence of the second ID became evident after the first 39 residues were deleted, and the resulting mutant channels did not lack inactivation but rather displayed rapid inactivation very similar to that observed with ID1 present at the N terminus [20, 21] It remained obscure what the functional role of the second ID might be in intact channels and, how inactivation gating is realized by this tandem inactivation domain. We analyze the solution structure and the function of this tandem inactivation domain of Kv1.4 channels (amino acids 1–75, Kv1.4N-(1–75)) using NMR spectroscopy together with patch clamp recordings in whole cell configuration and simulations of neuronal spiking behavior

Methods

Results

Discussion

Conclusion