Monte Carlo-energy minimization of correolide in the Kv1.3 channel: possible role of potassium ion in ligand-receptor interactions

BACKGROUND: Correolide, a nortriterpene isolated from the Costa Rican tree Spachea correa, is a novel immunosuppressant, which blocks Kv1.3 channels in human T lymphocytes. Earlier mutational studies suggest that correolide binds in the channel pore. Correolide has several nucleophilic groups, but the pore-lining helices in Kv1.3 are predominantly hydrophobic raising questions about the nature of correolide-channel interactions. RESULTS: We employed the method of Monte Carlo (MC) with energy minimization to search for optimal complexes of correolide in Kv1.2-based models of the open Kv1.3 with potassium binding sites 2/4 or 1/3/5 loaded with K+ ions. The energy was MC-minimized from many randomly generated starting positions and orientations of the ligand. In all the predicted low-energy complexes, oxygen atoms of correolide chelate a K+ ion. Correolide-sensing residues known from mutational analysis along with the ligand-bound K+ ion provide major contributions to the ligand-binding energy. Deficiency of K+ ions in the selectivity filter of C-type inactivated Kv1.3 would stabilize K+-bound correolide in the inner pore. CONCLUSION: Our study explains the paradox that cationic and nucleophilic ligands bind to the same region in the inner pore of K+ channels and suggests that a K+ ion is an important determinant of the correolide receptor and possibly receptors of other nucleophilic blockers of the inner pore of K+ channels

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

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—Cd(2+)—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 Cd(2+) 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