Conformational Transition of KcsA Gating and the Mechanism of its pH-Dependence

Abstract

The bacterial K+ channel KcsA has served as a prototypical system to study the architecture and the mechanism of gating of ion channels in response to proper stimuli. KcsA is activated in response to a drop in intracellular pH through the protonation of several His/Glu residues and the subsequent rearrangement of the helical bundle controlling the inner-gate opening. A constitutively open mutant channel (KcsA-OM) has been crystallized in several conformational states, each capturing a distinct degree of the inner-gate opening. However, due the heterogeneity of the structures and the use of antibodies, no firm conclusion could be made on the physiological relevance of the open conformation. In addition, due to the elimination of several charged residues, the mechanism of pH gating could not be fully determined by this particular mutant.We have conducted both equilibrium and nonequilibrium driven MD simulations of the channel in membrane to probe the conformational variability of KcsA. When the pH-sensing residues are protonated in the simulations, the structure relaxes into an open conformation that resembles the crystal structure of KcsA-OM with second largest (23 A) opening. The degree of opening captured in the simulations is consistent with that measured with EPR spectroscopy in the full-length KcsA, representing better the native state of the channel. Interestingly, the opening and closure of the cytoplasmic gate seem to be controlled by the competition between the protein-lipid interactions and several salt bridges between the channel's subunits. In particular, the neutralization of Glu118/Glu120 at low pH allows their entrance into the membrane, which permits the transmembrane helices surrounding the inner gate to tilt more and results in the opening of the channel.