Energetic Coupling Between Amino Acids in the pH-Sensing Region of the KcsA Channel

The bacterial potassium channel KcsA is gated by high concentrations of intracellular protons, allowing the channel to open at pH < 5.5. Replacing key ionizable residues from the N and C termini of KcsA with residues mimicking their protonated counterparts with respect to charge renders the channel open up to pH 9.0 (Thompson et al., 2008). We proposed that these residues function as the proton-binding sites. At neutral pH they form a complex network of inter- and intrasubunit salt bridges and hydrogen bonds near the bundle crossing, stabilizing the closed state. At acidic pH, these residues change their ionization state, thereby disrupting this network, favoring channel opening. To gain insight into the interactions that govern channel opening, we performed a thermodynamic analysis of the residues in the pH-sensing region. Individual mutations of most residues in this region result in modest shifts in the pH dependence of channel opening. However, pair-wise mutations of a subset of these amino acids show a large shift on the pH dependence of the channel opening suggesting these amino acids interact to open the channel with protons.