Molecular determinants of gating at the potassium-channel selectivity filter

Abstract

We show that in the potassium channel KcsA, proton-dependent activation is followed by an inactivation process similar to C-type inactivation, and this process is suppressed by an E71A mutation in the pore helix. EPR spectroscopy demonstrates that the inner gate opens maximally at low pH regardless of the magnitude of the single-channel-open probability, implying that stationary gating originates mostly from rearrangements at the selectivity filter. Two E71A crystal structures obtained at 2.5 A reveal large structural excursions of the selectivity filter during ion conduction and provide a glimpse of the range of conformations available to this region of the channel during gating. These data establish a mechanistic basis for the role of the selectivity filter during channel activation and inactivation.

Voltage Gating at the Selectivity Filter of the Ca2+ Release-activated Ca2+ Channel Induced by Mutation of the Orai1 Protein

“…We hypothesize the presence of a gating charge at the selectivity filter of the CRAC channel. A similar phenomenon is observed in the KcsA K ϩ channel, where the reorientation of the Glu 71 residue in the electric field induces destabilization of the inactivated conformation of the channel pore (39,40). Our results, however, suggest that the gating charge assumes a position that stabilizes the open channel conformation (destabilizes the closed conformation) at negative voltages.…”

Probing Conformational Changes during the Gating Cycle of a Potassium Channel in Lipid Bilayers

“…Under steady-state conditions, KcsA exhibits a low open probability (P O ) because most of the time it resides in a C-type inactivated state (12,(33)(34)(35). It was previously shown that anionic phospholipids may increase P O , providing a more favorable environment for the open state of KcsA (36)(37)(38).…”

“…Upon activation at acidic pH, KcsA mediates a transient K þ current that is characterized by a fast activation time course followed by a slow C-type inactivation process (12). This time course is well described by a successive and coordinate opening and closing of activation and inactivation gates within the conduction pathway of the KcsA channel ( Fig.…”

“…Despite these advancements, a direct structural visualization of the entire KcsA gating cycle, including the conformation with an open activation and closed inactivation gate (open-conductive conformation) in a lipidic environment has remained challenging due to the very short lifetime of this channel state and the propensity of the selectivity filter to rapidly collapse when the activation gate is opened (12)(13)(14)24,25). Using solid-state NMR spectroscopy, we previously identified a lipid-KcsA interaction site at the turret region, which connects the outer transmembrane helix of KcsA (transmembrane helix 1 (TM1)) and the pore helix behind the selectivity filter of the KcsA channel.…”

Principles underlying energetic coupling along an allosteric communication trajectory of a voltage-activated K + channel

“…Such conformational transitions regulate the flow of potassium ions across the membrane (6-9), a process underlying many fundamental biological processes, in particular the generation of nerve and muscle action potentials (10). These conformational changes, moreover, play a fundamental role in mediating coupling between the voltage-sensor, activation gate and selectivity filter elements of Kv channels (6)(7)(8)(9)(11)(12)(13)(14).Amenable to rapid and highly accurate functional characterization without the need for protein purification, the Kv channel represents an excellent model for studying the features underlying allosteric communication networks in proteins. Recent systematic energy perturbation analysis of the pore domain of the archetypal Shaker Kv channel (15) revealed that gating-sensitive positionsi.e., those positions where mutation dramatically affects the closedopen equilibrium of the channel-cluster around the channel activation gate and near the selectivity filter functional elements, when mapped onto the closed channel pore structure (Fig.…”