K+ channel inactivation mediated by the concerted action of the cytoplasmic N- and C-terminal domains

Abstract

We have examined the molecular mechanism of rapid inactivation gating in a mouse Shal K+ channel (mKv4.1). The results showed that inactivation of these channels follows a complex time course that is well approximated by the sum of three exponential terms. Truncation of an amphipathic region at the N-terminus (residues 2-71) abolished the rapid phase of inactivation (r = 16 ms) and altered voltage-dependent gating. Surprisingly, these effects could be mimicked by deletions affecting the hydrophilic C-terminus. The sum of two exponential terms was sufficient to describe the inactivation of deletion mutants. In fact, the time constants corresponded closely to those of the intermediate and slow phases of inactivation observed with wild-type channels. Further analysis revealed that several basic amino acids at the N-terminus do not influence inactivation, but a positively charged domain at the C-terminus (amino acids 420-550) is necessary to support rapid inactivation. Thus, the amphipathic N-terminus and the hydrophilic C-terminus of mKv4.1 are essential determinants of inactivation gating and may interact with each other to maintain the N-terminal inactivation gate near the inner mouth of the channel. Furthermore, this inactivation gate may not behave like a simple open-channel blocker because channel blockade by internal tetraethylammonium was not associated with slower current decay and an elevated external K+ concentration retarded recovery from inactivation.

Modulation of the Voltage-gated Potassium Channel (Kv4.3) and the Auxiliary Protein (KChIP3) Interactions by the Current Activator NS5806

“…It has also been shown that the association of KChIPs at the N terminus of Kv4 channels impairs open-state inactivation (41). Electrophysiological studies support the idea that both the N and C termini of Kv4 channels interact to regulate the inactivation kinetics (40,42), opening the possibility that such interactions are modulated by binding of KChIPs at the N terminus (34,38). It is also proposed that the N terminus functions as a membrane transport control, which in the absence of KChIPs anchors Kv4 channels near the perinuclear region of the cell (43,44).…”

KChIP3 Rescues the Functional Expression of Shal Channel Tetramerization Mutants

“…Kv4 channel inactivation has been dissected in numerous biophysical studies into three separate processes: open state, closed state, and slow inactivation (15,20,21). Open state inactivation utilizes a pore block produced by the Kv4 cytoplasmic N terminus and is lost by deletion of the Kv4 N terminus or binding of KChIP proteins to the N terminus (16,22).…”

“…The N terminus of Kv4.2 is clearly involved in inactivation gating, since deletion of this region produces slowed inactivation. Moreover, experiments on the functional properties of the N terminus show that it can act as a pore-binding peptide (15)(16)(17). Therefore, the gating changes produced by KChIPs might depend on the binding and sequestering of the Kv4 N terminus, causing an alteration to a complex set of allosteric interactions that involve other structural elements of the channel (18).…”

Conformational changes in the C terminus of Shaker K + channel bound to the rat Kvβ2-subunit

“…It is known that in Shaker-like K ϩ channels, C termini are involved in a variety of processes that range from channel gating (10,11) and voltage sensitivity (4) to the binding of the membrane-associated guanylate kinases (12). They also participate in channel assembly in Kv 2.1 (13,14).…”