Cryo-EM structure of human voltage-gated sodium channel Na v 1.6

Figures

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  Fig. 1.. Cryo-EM structure of human Nav1.6-β1. (A) Electrophysiological properties of Nav1.6 alone or co-expressed with the auxiliary subunits β1 and FHF2B in HEK293T cells. Shown here are the voltage-dependent activation and inactivation curves. Please refer to Methods and SI Appendix, Table S1 for details. (B) Cryo-EM map of the Nav1.6 complex comprising the α1 (domain colored) and β1 (light salmon) subunits. The cytosolic III-IV linker is colored chocolate. Sugar moieties and lipids are colored gray and orange, respectively. The same color scheme is applied throughout the manuscript. (C) Overall structure of the Nav1.6 complex in a Side view (Left) and a cytoplasmic view (Right). Sugar moieties are shown as sticks, and the IFM motif within the cytosolic III-IV linker is shown as spheres.

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  Fig. 2.. ECLI is sandwiched by β1 and the glycan attached to Asn326. (A) ECLs are the least conserved structural elements among Nav subtypes. The conservation scores (calculated by ConSurf) of human Navs are color-coded and presented on the Nav1.6 structure. The ECLI and ECLIII region are shown as surface. Two interfaces, ECLI-β1 and ECLI-glycan, are highlighted in squares. (B) Extensive interactions between ECLI and β1. Residues forming the hydrogen bond network, indicated by red, dashed lines, at the interface of ECLI and β1 are shown. (C) Interactions between ECLI and Asn326-glycan. Two aromatic residues, Trp302 and Phe288, are involved in the carbohydrate–aromatic interactions. The density in the Left panel is contoured at 5.2 σ. (D) Multiple sequence alignment of ECLI in human Nav subtypes. The alignment, calculated in Clustal X, is shaded in different colors based on sequence conservation and the chemical properties of the residues. The conserved glycosylation site (Asn326 in Nav1.6) is indicated. (E) Schematic illustration of the chemical assembly of the branched N-GlcNAc2Man7 glycan. Related key residues are labeled in blue ovals. (F) Density variations of the corresponding glycan…

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  Fig. 3.. Unique ligand density in the inactivated structure of Nav1.6. (A) An unidentified density bound to the IV-I fenestration of the PD. A Side view and a Top view of the PD are shown. A putative PE is shown as gray ball-and-sticks in the semitransparent gray density. The density-colored pale yellow may belong to a molecule whose identity is unknown. (B) The intracellular gate of Nav1.6 is penetrated by a GDN (green spheres). The permeation path of Nav1.6 is calculated in HOLE, and the pore radii are superimposed with that of human Nav1.2 (green) and Nav1.7 (pink) (43). (C) VSDIV exhibits a slightly different conformation in different Nav channels. Structures of VSDIV from Nav1.6 (cyan), Nav1.2 (green), and Nav1.7 (pink) are superimposed relative to the overall structure. The PDB codes for Nav1.2 and Nav1.7 are 6J8E and 7W9K, respectively. Two perpendicular views are shown. (D) Distinct coordination of GC residues in VSDIV of Nav1.7 (Left) and Nav1.6 (Right). The GC residues on S4IV and their coordinating residues on S1IV-S3IV are shown as sticks, and the potential hydrogen bonds are shown as red-dashed lines.

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  Fig. 4.. Structural mapping of disease-related mutations in human Nav1.6. (A) Mapping of the disease-associated mutations on the structure of Nav1.6 in two perpendicular views. The Cα atoms of the disease-related residues are shown as spheres and colored for different types of disease. DEE13 (purple); BFIS5 (blue); MYOCL2 (red). (B) Mapping of the disease-related mutations in each domain of human Nav1.6.