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KvAP-Based Model of the Pore Region of Shaker Potassium Channel Is Consistent with Cadmium- and Ligand-Binding Experiments

Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, Canada

Correspondence: Address reprint requests to Boris S. Zhorov, Dept. of Biochemistry and Biomedical Sciences, McMaster University, 1200 Main St. W., Hamilton, ON, L8N 3Z5 Canada. Tel.: 905-525-9140 ext. 22049; Fax: 905-522-9033; E-mail: zhorov{at}mcmaster.ca.

Potassium channels play fundamental roles in excitable cells. X-ray structures of bacterial potassium channels show that the pore-lining inner helices obstruct the cytoplasmic entrance to the closed channel KcsA, but diverge in widely open channels MthK and KvAP, suggesting a gating-hinge role for a conserved Gly in the inner helix. A different location of the gating hinge and a narrower open pore were proposed for voltage-gated Shaker potassium channels that have the Pro-473-Val-Pro motif. Two major observations back the proposal: cadmium ions lock mutant Val-476-Cys in the open state by bridging Cys-476 and His-486 in adjacent helices, and cadmium blocks the locked-open double mutant Val-474-Cys/Val-476-Cys by binding to Cys-474 residues. Here we used molecular modeling to show that the open Shaker should be as wide as KvAP to accommodate an open-channel blocker, correolide. We further built KvAP-, MthK-, and KcsA-based models of the Shaker mutants and Monte-Carlo-minimized them with constraints Cys-476—Cd²⁺—His-486. The latter were consistent with the KvAP-based model, causing a small-bend N-terminal to the Pro-473-Val-Pro motif. The constraints significantly distorted the MthK-based structure, making it similar to KvAP. The KcsA structure resisted the constraints. Two Cd²⁺ ions easily block the locked-open KvAP-based model at Cys-474 residues, whereas constraining a single cadmium ion to four Cys-474 caused large conformational changes and electrostatic imbalance. Although mutual disposition of the voltage-sensor and pore domains in the KvAP x-ray structure is currently disputed, our results suggest that the pore-region domain retains a nativelike conformation in the crystal.

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