A Localized Interaction Surface for Voltage-Sensing Domains on the Pore Domain of a K+ Channel

Abstract

Voltage-gated K+ channels contain a central pore domain and four surrounding voltage-sensing domains. How and where changes in the structure of the voltage-sensing domains couple to the pore domain so as to gate ion conduction is not understood. The crystal structure of KcsA, a bacterial K+ channel homologous to the pore domain of voltage-gated K+ channels, provides a starting point for addressing this question. Guided by this structure, we used tryptophan-scanning mutagenesis on the transmembrane shell of the pore domain in the Shaker voltage-gated K+ channel to localize potential protein-protein and protein-lipid interfaces. Some mutants cause only minor changes in gating and when mapped onto the KcsA structure cluster away from the interface between pore domain subunits. In contrast, mutants producing large changes in gating tend to cluster near this interface. These results imply that voltage-sensing domains interact with localized regions near the interface between adjacent pore domain subunits.

S4 Charges Move Close to Residues in the Pore Domain during Activation in a K Channel

“…The large motion of R362 suggested by our results is compatible with both a helical screw motion (Catterall 1986; Guy and Seetharamulu 1986; Glauner et al 1999; Keynes and Elinder 1999; Gandhi et al 2000) and a pure twisting motion (Papazian and Bezanilla 1997; Cha et al 1999; Glauner et al 1999) of S4. Our estimated location and motion is also compatible with the results of other studies (Li-Smerin et al 2000; Loots and Isacoff 2000). …”

Structural basis of two-stage voltage-dependent activation in K+channels

“…A variety of distance constraints based on experimental data were modeled as artificial distance-energy restraints in the form of a quadratic function with harmonic force constants of 5-20 kcal⅐[mol Å 2 ] Ϫ1 (7). Models were based on the following reasonable assumptions: (i) the central pore in voltagegated K ϩ channels is similar in structure to KcsA (1), which is consistent with the high degree of sequence similarity between Shaker and KcsA, with toxin-binding data, and with the generation of functional chimeric channels containing the Shaker voltage sensor and the KcsA pore (8,9); (ii) S1-S4 form transmembrane ␣-helices that are oriented roughly perpendicular to the plane of the membrane, which is consistent with alanine and tryptophan scanning mutagenesis of these segments in voltage-gated K ϩ channels (10-12); (iii) the voltage sensor and pore domains interact closely in the structure, which is supported by single-particle electron microscope images of Shaker channels at 25-Å resolution (13), by perturbation analysis of the pore domain (14), by distance measurements using lanthanide-based resonance energy transfer (15) and tethered quaternary ammonium pore blockers (16), and by electrostatic calculations (17).…”

The S1 helix critically regulates the finely tuned gating of Kv11.1 channels

“…Similar approaches that use perturbations from scanning mutagenesis to assess the role and interaction partners for various helices and regions of the voltage sensor domain have been applied to other voltage-gated potassium channels (52)(53)(54)(55), as well as to other helices of the voltage sensor domain in Kv11.1 channels (56). In this study perturbations to the equilibrium of activation or inactivation gating parameters are reported as changes in the Gibbs free energy (⌬⌬G) of the transition, compared with WT, whereas perturbations to the rates of activation, deactivation, onset of inactivation, or recovery from inactivation are reported as changes in lnk relative to WT (Ϫ⌬lnk).…”