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Yorodumi- PDB-6p6w: Cryo-EM structure of voltage-gated sodium channel NavAb N49K/L109... -
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-Basic information
Entry | Database: PDB / ID: 6p6w | |||||||||||||||
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Title | Cryo-EM structure of voltage-gated sodium channel NavAb N49K/L109A/M116V/G94C/Q150C disulfide crosslinked mutant in the resting state | |||||||||||||||
Components | Fusion of Maltose-binding protein and voltage-gated sodium channel NavAb | |||||||||||||||
Keywords | membrane protein / metal transport / Ion channel / ion transport protein | |||||||||||||||
Function / homology | Function and homology information membrane depolarization during action potential / voltage-gated sodium channel complex / voltage-gated sodium channel activity / detection of maltose stimulus / maltose transport complex / carbohydrate transport / maltose binding / maltose transport / maltodextrin transmembrane transport / carbohydrate transmembrane transporter activity ...membrane depolarization during action potential / voltage-gated sodium channel complex / voltage-gated sodium channel activity / detection of maltose stimulus / maltose transport complex / carbohydrate transport / maltose binding / maltose transport / maltodextrin transmembrane transport / carbohydrate transmembrane transporter activity / ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing / ATP-binding cassette (ABC) transporter complex / cell chemotaxis / outer membrane-bounded periplasmic space / periplasmic space / DNA damage response / identical protein binding / membrane / metal ion binding Similarity search - Function | |||||||||||||||
Biological species | Escherichia coli K-12 (bacteria) Arcobacter butzleri (bacteria) | |||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4 Å | |||||||||||||||
Authors | Wisedchaisri, G. / Tonggu, L. / McCord, E. / Gamal El-Din, T.M. / Wang, L. / Zheng, N. / Catterall, W.A. | |||||||||||||||
Funding support | United States, 4items
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Citation | Journal: Cell / Year: 2019 Title: Resting-State Structure and Gating Mechanism of a Voltage-Gated Sodium Channel. Authors: Goragot Wisedchaisri / Lige Tonggu / Eedann McCord / Tamer M Gamal El-Din / Liguo Wang / Ning Zheng / William A Catterall / Abstract: Voltage-gated sodium (Na) channels initiate action potentials in nerve, muscle, and other electrically excitable cells. The structural basis of voltage gating is uncertain because the resting state ...Voltage-gated sodium (Na) channels initiate action potentials in nerve, muscle, and other electrically excitable cells. The structural basis of voltage gating is uncertain because the resting state exists only at deeply negative membrane potentials. To stabilize the resting conformation, we inserted voltage-shifting mutations and introduced a disulfide crosslink in the VS of the ancestral bacterial sodium channel NaAb. Here, we present a cryo-EM structure of the resting state and a complete voltage-dependent gating mechanism. The S4 segment of the VS is drawn intracellularly, with three gating charges passing through the transmembrane electric field. This movement forms an elbow connecting S4 to the S4-S5 linker, tightens the collar around the S6 activation gate, and prevents its opening. Our structure supports the classical "sliding helix" mechanism of voltage sensing and provides a complete gating mechanism for voltage sensor function, pore opening, and activation-gate closure based on high-resolution structures of a single sodium channel protein. | |||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6p6w.cif.gz | 190.9 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6p6w.ent.gz | 147.5 KB | Display | PDB format |
PDBx/mmJSON format | 6p6w.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6p6w_validation.pdf.gz | 716.6 KB | Display | wwPDB validaton report |
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Full document | 6p6w_full_validation.pdf.gz | 723.1 KB | Display | |
Data in XML | 6p6w_validation.xml.gz | 35.7 KB | Display | |
Data in CIF | 6p6w_validation.cif.gz | 52.4 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/p6/6p6w ftp://data.pdbj.org/pub/pdb/validation_reports/p6/6p6w | HTTPS FTP |
-Related structure data
Related structure data | 20265MC 6p6xC 6p6yC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 74072.797 Da / Num. of mol.: 4 Mutation: R4A, N49K, L109A, M116V, G94C, Q150C,R4A, N49K, L109A, M116V, G94C, Q150C Source method: isolated from a genetically manipulated source Source: (gene. exp.) Escherichia coli K-12 (bacteria), (gene. exp.) Arcobacter butzleri (strain RM4018) (bacteria) Gene: malE, BX06_05300, Abu_1752 / Strain: RM4018 / Production host: Trichoplusia ni (cabbage looper) References: UniProt: A0A028ANC5, UniProt: A8EVM5, UniProt: P0AEX9*PLUS |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Fusion of maltose-binding protein and voltage-gated sodium channel NavAb in the resting state Type: COMPLEX / Entity ID: all / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Value: 0.3 MDa / Experimental value: YES | ||||||||||||||||||||
Source (natural) | Organism: Arcobacter butzleri RM4018 (bacteria) | ||||||||||||||||||||
Source (recombinant) | Organism: Trichoplusia ni (cabbage looper) | ||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: C-flat-1.2/1.3 4C | ||||||||||||||||||||
Vitrification | Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 40 % / Chamber temperature: 298 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 103.5 K / Temperature (min): 90.8 K |
Image recording | Average exposure time: 8.6 sec. / Electron dose: 60 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 9 / Num. of real images: 5000 |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
Image scans | Width: 3838 / Height: 3710 / Movie frames/image: 43 / Used frames/image: 1-43 |
-Processing
Software | Name: PHENIX / Version: 1.14_3260: / Classification: refinement | ||||||||||||||||||||||||||||||||||||||||
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EM software |
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Image processing | Details: Movie frames were aligned and 2x binned to a pixel size off 1.056 A using MotionCor2. | ||||||||||||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 476000 Details: Auto-particle picking using RELION 3.0 beta and manual particle removals of bad particles. | ||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C4 (4 fold cyclic) | ||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 333899 / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: FLEXIBLE FIT / Space: REAL | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 3RVY Pdb chain-ID: A / Pdb chain residue range: 1001-1221 |