Research Article

Structure of a eukaryotic voltage-gated sodium channel at near-atomic resolution

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Science  03 Mar 2017:
Vol. 355, Issue 6328, eaal4326
DOI: 10.1126/science.aal4326

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Navigating regulated cell excitation

Voltage-gated sodium (Nav) channels respond to a change in voltage potential by allowing sodium ions to move into cells, thus initiating electrical signaling. Mutations in Nav channels cause neurological and cardiovascular disorders, making the channels important therapeutic targets. Shen et al. determined a high-resolution structure of a Nav channel from the American cockroach by electron microscopy. The structure affords insight into voltage sensing and ion permeability and provides a foundation for understanding function and disease mechanism of Nav and the related Cav ion channels.

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Structured Abstract


Voltage-gated sodium (Nav) channels are responsible for the generation and propagation of action potentials in excitable cells. They undergo voltage-dependent activation to initiate electrical signaling at millisecond scale and inactivate by both fast and slow mechanisms. The eukaryotic Nav channels comprise a pore-forming α subunit and auxiliary β subunits that facilitate membrane localization and modulate channel properties. The α subunit is a single polypeptide chain that folds to four homologous repeats (domains I to IV), each containing six transmembrane segments, S1 to S6. The S5 and S6 segments enclose the central pore domain, and their intervening sequences constitute the selectivity filter (SF). One residue at the corresponding SF locus in each repeat, Asp/Glu/Lys/Ala (DEKA), determines Na+ selectivity. The S1 to S4 segments in each repeat form a voltage-sensing domain (VSD), wherein S4 carries repetitively occurring positive residues essential for voltage sensing.

More than 1000 mutations have been identified in human Nav channels associated with various neurological and cardiovascular disorders. Nav channels represent important targets for multiple pharmaceutical drugs and natural toxins. The atomic structure of a eukaryotic Nav channel is required to reveal the molecular basis for ion selectivity, voltage-dependent activation and inactivation, and recognition of toxins, agonists, and antagonists.


The technological breakthrough of electron microscopy (EM) has offered unprecedented opportunity for structure elucidation of eukaryotic Nav channels, but the bottleneck exists in the generation of sufficient amounts of high-quality proteins. After extensive screening, we succeeded in obtaining homogeneous proteins of PaFPC1, a putative Nav channel from the American cockroach. Despite the lack of electrophysiological characterizations, PaFPC1 contains all the hallmarks of a Nav channel except for the fast inactivation motif. We designated the protein NavPaS.


The cryogenic EM (cryo-EM) structure of NavPaS was determined with an overall resolution of 3.8 Å, allowing side-chain assignment for the complete transmembrane fold, extracellular loops of the pore domain, the intact III-IV linker, and the carboxy-terminal domain (CTD). A poly-Ala backbone was modeled for an amino-terminal domain that is located below VSDI. In addition, 20 sugar moieties were built into seven glycosylation sites on the extracellular loops. Conserved disulfide bonds are observed within the extracellular loops and between P2II and S6II segments.

The asymmetric selectivity filter vestibule is constituted by the side chains of the signature DEKA residues and the carbonyl oxygens of the two preceding residues in each repeat. The closed pore domain has only one small fenestration constituted by the S6III and S6IV segments. The four VSDs exhibit distinct conformations, with the corresponding gating charges located at different heights relative to their respective charge transfer center. Extensive interactions are observed between the III-IV linker, CTD, VSDIV, S4-S5IV, S6IV, and S6III segments of NavPaS. Despite the sequence variations of the III-IV linker and the lack of Ile/Met/Phe/Thr motif, the structure provides an important clue to understanding the fast inactivation mechanism of Nav channels.


The structure of a single-chain eukaryotic Nav channel serves as the framework for elucidating function and disease mechanisms of Nav channels. It provides the molecular template for interpretation of a wealth of experimental observations accumulated over the past six decades. Structural comparison between the related NavPaS and Cav1.1 reveals conformational shifts that may shed light on the understanding of the electromechanical coupling mechanism of voltage-gated channels. Structure-guided protein engineering will facilitate future mechanistic investigations of Nav and Cav channels.

The cryo-EM structure of a eukaryotic Nav channel at 3.8-Å resolution.

(Left) The overall structure of NavPaS/PaFPC1, a putative Nav channel identified from American cockroach. The glycosyl moieties and disulfide bonds are shown as sticks and spheres, respectively. The structure is domain colored. (Right) The pore domain of NavPaS. The permeation path is illustrated by brown dots in the pore domain, and the corresponding pore radii along the conducting passage are tabulated on the right. The functional entities along the permeation path—including the selectivity filter, the central cavity, and the intracellular activation gate—are annotated.


Voltage-gated sodium (Nav) channels are responsible for the initiation and propagation of action potentials. They are associated with a variety of channelopathies and are targeted by multiple pharmaceutical drugs and natural toxins. Here, we report the cryogenic electron microscopy structure of a putative Nav channel from American cockroach (designated NavPaS) at 3.8 angstrom resolution. The voltage-sensing domains (VSDs) of the four repeats exhibit distinct conformations. The entrance to the asymmetric selectivity filter vestibule is guarded by heavily glycosylated and disulfide bond–stabilized extracellular loops. On the cytoplasmic side, a conserved amino-terminal domain is placed below VSDI, and a carboxy-terminal domain binds to the III-IV linker. The structure of NavPaS establishes an important foundation for understanding function and disease mechanism of Nav and related voltage-gated calcium channels.

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