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- EMDB-0501: Cryo-EM structure of a human-cockroach hybrid Nav channel bound t... -

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Entry
Database: EMDB / ID: EMD-0501
TitleCryo-EM structure of a human-cockroach hybrid Nav channel bound to alpha-scorpion toxin AaH2.
Map dataSharpened and mask map used for model building.
Sample
  • Complex: NavPas-VSD4-AaH2
    • Protein or peptide: Alpha-mammal toxin AaH2
  • Protein or peptide: Sodium channel protein PaFPC1,Sodium channel protein type 9 subunit alpha,Sodium channel protein PaFPC1
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: (7E,21R,24S)-27-amino-24-hydroxy-18,24-dioxo-19,23,25-trioxa-24lambda~5~-phosphaheptacos-7-en-21-yl (9Z,12E)-octadeca-9,12-dienoate
  • Ligand: Digitonin
  • Ligand: 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE
  • Ligand: CHOLESTEROL HEMISUCCINATE
KeywordsSodium channel / scorpion toxin / electrical signaling / fast inactivation / MEMBRANE PROTEIN
Function / homology
Function and homology information


action potential propagation / detection of mechanical stimulus involved in sensory perception / membrane depolarization during action potential / cardiac muscle cell action potential involved in contraction / node of Ranvier / voltage-gated sodium channel complex / sodium channel inhibitor activity / voltage-gated monoatomic cation channel activity / Interaction between L1 and Ankyrins / voltage-gated sodium channel activity ...action potential propagation / detection of mechanical stimulus involved in sensory perception / membrane depolarization during action potential / cardiac muscle cell action potential involved in contraction / node of Ranvier / voltage-gated sodium channel complex / sodium channel inhibitor activity / voltage-gated monoatomic cation channel activity / Interaction between L1 and Ankyrins / voltage-gated sodium channel activity / detection of temperature stimulus involved in sensory perception of pain / Phase 0 - rapid depolarisation / behavioral response to pain / neuronal action potential / axon terminus / sensory perception of pain / sodium ion transmembrane transport / post-embryonic development / defense response / circadian rhythm / response to toxic substance / Sensory perception of sweet, bitter, and umami (glutamate) taste / toxin activity / inflammatory response / axon / extracellular region / plasma membrane
Similarity search - Function
Scorpion long chain toxin / LCN-type cysteine-stabilized alpha/beta (CS-alpha/beta) domain / LCN-type cysteine-stabilized alpha/beta (CS-alpha/beta) domain profile. / Scorpion long chain toxin/defensin / Scorpion toxin-like domain / Knottins / Knottin, scorpion toxin-like / Voltage-gated Na+ ion channel, cytoplasmic domain / Cytoplasmic domain of voltage-gated Na+ ion channel / Voltage-dependent L-type calcium channel, IQ-associated domain ...Scorpion long chain toxin / LCN-type cysteine-stabilized alpha/beta (CS-alpha/beta) domain / LCN-type cysteine-stabilized alpha/beta (CS-alpha/beta) domain profile. / Scorpion long chain toxin/defensin / Scorpion toxin-like domain / Knottins / Knottin, scorpion toxin-like / Voltage-gated Na+ ion channel, cytoplasmic domain / Cytoplasmic domain of voltage-gated Na+ ion channel / Voltage-dependent L-type calcium channel, IQ-associated domain / Voltage-dependent L-type calcium channel, IQ-associated / Sodium ion transport-associated / Voltage-gated sodium channel alpha subunit, inactivation gate / Sodium ion transport-associated / SCN5A-like, C-terminal IQ motif / Voltage gated sodium channel, alpha subunit / Knottin, scorpion toxin-like superfamily / Voltage-gated cation channel calcium and sodium / Short calmodulin-binding motif containing conserved Ile and Gln residues. / IQ motif, EF-hand binding site / Voltage-dependent channel domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Sodium channel protein PaFPC1 / Alpha-mammal toxin AaH2 / Sodium channel protein type 9 subunit alpha
Similarity search - Component
Biological speciesPeriplaneta americana (American cockroach) / Androctonus australis (Sahara scorpion)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.5 Å
AuthorsClairfeuille T / Rohou A
CitationJournal: Science / Year: 2019
Title: Structural basis of α-scorpion toxin action on Na channels.
Authors: Thomas Clairfeuille / Alexander Cloake / Daniel T Infield / José P Llongueras / Christopher P Arthur / Zhong Rong Li / Yuwen Jian / Marie-France Martin-Eauclaire / Pierre E Bougis / Claudio ...Authors: Thomas Clairfeuille / Alexander Cloake / Daniel T Infield / José P Llongueras / Christopher P Arthur / Zhong Rong Li / Yuwen Jian / Marie-France Martin-Eauclaire / Pierre E Bougis / Claudio Ciferri / Christopher A Ahern / Frank Bosmans / David H Hackos / Alexis Rohou / Jian Payandeh /
Abstract: Fast inactivation of voltage-gated sodium (Na) channels is essential for electrical signaling, but its mechanism remains poorly understood. Here we determined the structures of a eukaryotic Na ...Fast inactivation of voltage-gated sodium (Na) channels is essential for electrical signaling, but its mechanism remains poorly understood. Here we determined the structures of a eukaryotic Na channel alone and in complex with a lethal α-scorpion toxin, AaH2, by electron microscopy, both at 3.5-angstrom resolution. AaH2 wedges into voltage-sensing domain IV (VSD4) to impede fast activation by trapping a deactivated state in which gating charge interactions bridge to the acidic intracellular carboxyl-terminal domain. In the absence of AaH2, the S4 helix of VSD4 undergoes a ~13-angstrom translation to unlatch the intracellular fast-inactivation gating machinery. Highlighting the polypharmacology of α-scorpion toxins, AaH2 also targets an unanticipated receptor site on VSD1 and a pore glycan adjacent to VSD4. Overall, this work provides key insights into fast inactivation, electromechanical coupling, and pathogenic mutations in Na channels.
History
DepositionJan 28, 2019-
Header (metadata) releaseFeb 20, 2019-
Map releaseFeb 20, 2019-
UpdateMay 28, 2025-
Current statusMay 28, 2025Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4.6
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 4.6
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6nt4
  • Surface level: 4.6
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_0501.png

Downloads & links

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Map

FileDownload / File: emd_0501.map.gz / Format: CCP4 / Size: 59.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened and mask map used for model building.
Projections & slices

Image control

Size
Brightness
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
1 Å/pix.
x 250 pix.
= 250. Å		1 Å/pix.
x 250 pix.
= 250. Å		1 Å/pix.
x 250 pix.
= 250. Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 4.6 / Movie #1: 4.6
Minimum - Maximum-14.935369 - 26.928422999999999
Average (Standard dev.)0.044959076 (±0.87032884)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions250250250
Spacing250250250
CellA=B=C: 250.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z111
M x/y/z250250250
origin x/y/z0.0000.0000.000
length x/y/z250.000250.000250.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS250250250
D min/max/mean-14.93526.9280.045

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Supplemental data

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Mask #1

Fileemd_0501_msk_1.map
Projections & Slices
AxesZYX

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Additional map: Unsharpened and unmodified map straight out of cisTEM...

Fileemd_0501_additional.map
AnnotationUnsharpened and unmodified map straight out of cisTEM last local refinement round.
Projections & Slices
AxesZYX

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Histogram

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Half map: Halfmap 1

Fileemd_0501_half_map_1.map
AnnotationHalfmap 1
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Histogram

Histogram (log scale)

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Half map: Halfmap 2

Fileemd_0501_half_map_2.map
AnnotationHalfmap 2
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Sample components

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Entire : NavPas-VSD4-AaH2

EntireName: NavPas-VSD4-AaH2
Components
  • Complex: NavPas-VSD4-AaH2
    • Protein or peptide: Alpha-mammal toxin AaH2
  • Protein or peptide: Sodium channel protein PaFPC1,Sodium channel protein type 9 subunit alpha,Sodium channel protein PaFPC1
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: (7E,21R,24S)-27-amino-24-hydroxy-18,24-dioxo-19,23,25-trioxa-24lambda~5~-phosphaheptacos-7-en-21-yl (9Z,12E)-octadeca-9,12-dienoate
  • Ligand: Digitonin
  • Ligand: 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE
  • Ligand: CHOLESTEROL HEMISUCCINATE

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Supramolecule #1: NavPas-VSD4-AaH2

SupramoleculeName: NavPas-VSD4-AaH2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #2
Source (natural)Organism: Periplaneta americana (American cockroach)
Molecular weightTheoretical: 155 KDa

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Macromolecule #1: Sodium channel protein PaFPC1,Sodium channel protein type 9 subun...

MacromoleculeName: Sodium channel protein PaFPC1,Sodium channel protein type 9 subunit alpha,Sodium channel protein PaFPC1
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Periplaneta americana (American cockroach)
Molecular weightTheoretical: 178.763656 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: WSHPQFEKGG GSGGGSGGSA WSHPQFEKGG SGGDYKDDDD KGGSGGDYKD DDDKMADNSP LIREERQRLF RPYTRAMLTA PSAQPAKEN GKTEENKDNS RDKGRGANKD RDGSAHPDQA LEQGSRLPAR MRNIFPAELA STPLEDFDPF YKNKKTFVVV T KAGDIFRF ...String:
WSHPQFEKGG GSGGGSGGSA WSHPQFEKGG SGGDYKDDDD KGGSGGDYKD DDDKMADNSP LIREERQRLF RPYTRAMLTA PSAQPAKEN GKTEENKDNS RDKGRGANKD RDGSAHPDQA LEQGSRLPAR MRNIFPAELA STPLEDFDPF YKNKKTFVVV T KAGDIFRF SGEKSLWMLD PFTPIRRVAI STMVQPIFSY FIMITILIHC IFMIMPATQT TYILELVFLS IYTIEVVVKV LA RGFILHP FAYLRDPWNW LDFLVTLIGY ITLVVDLGHL YALRAFRVLR SWRTVTIVPG WRTIVDALSL SITSLKDLVL LLL FSLSVF ALIGLQLFMG NLKHKCVKHF PADGSWGNFT DERWFNYTSN SSHWYIPDDW IEYPLCGNSS GAGMCPPGYT CLQG YGGNP NYGYTSFDTF GWAFLSVFRL VTLDYWEDLY QLALRSAGPW HILFFIIVVF YGTFCFLNFI LAVVVMSYTH MVKRA DEEK AAERELKKEK KAASVANNTA NGQEQTTIEM NGDEAVVIDN NDQAARQQSD PETPAPSVTQ RLTDFLCVWD CCVPWQ KLQ GAIGAVVLSP FFELFIAVII VLNITFMALD HHDMNIEFER ILRTGNYIFT SIYIVEAVLK IIALSPKFYF KDSWNVF DF IIVVFAILEL GLEGVQGLSV FRSFRLLRVF RLAKFWPTLN NFMSVMTKSY GAFVNVMYVM FLLLFIFAII GMQLFGMN Y IDNMERFPDG DLPRWNFTDF LHSFMIVFRA LCGEWIESMW DCMLVGDWSC IPFFVAVFFV GNLVILNLLI ALLLNNYGS FCTSPTSDEE DSKDEDALAQ IVRIFKRFKP NLNAVKLSPM KPDSEDIVES QEIQGNNIAD AEDVLAGEFP PDCCCNAFYK CFPSRPARD SSVQRMWSNI RRVCFLLAKN KYFQKFVTAV LVITSVLLAL EDIYLPQRPV LVNITLYVDY VLTAFFVIEM I IMLFAVGF KKYFTSKWYW LDFIVVVAYL LNFVLMCAGI EALQTLRLLR VFRLFRPLSK VNGMQVVTST LVEAVPHIFN VI LVGIFFW LVFAIMGVQL FAGKFYKCVD ENSTVLSHEI TMDRNDCLHE NYTWENSPMN FDHVGNAYLS LLQVATFKGW LQI MNDAID SREVHKQPIR ETNIYMYLYF IFFIVFGSFF ILKLFVCILI DIFRQQRRKA EGLSATDSRT QLIYRRAVMR TMSA KPVKR IPKPGNKIQG CIFDLVTNQA FDISIMVLIC LNMVTMMVEK EGQSQHMTEV LYWINVVFII LFTGECVLKL ISLRH YYFT VGWNIFDFVV VIISIVGMFL ADLIETYFVS PTLFRVIRLA RIGRILRLVK GAKGIRLLLL ALRKALRTLF NVSFLL FVI MFVYAVFGME FFMHIRDAGA IDDVYNFKTF GQSIILLFQL ATSAGWDGVY FAIANEEDCR APDHELGYPG NCGSRAL GI AYLVSYLIIT CLVVINMYAA VILDYVLEVY EDSKEGLTDD DYDMFFEVWQ QFDPEATQYI RYDQLSELLE ALQPPLQV Q KPNKYKILSM NIPICKDDHI FYKDVLEALV KDVFSRRG

UniProtKB: Sodium channel protein PaFPC1, Sodium channel protein type 9 subunit alpha, Sodium channel protein PaFPC1

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Macromolecule #2: Alpha-mammal toxin AaH2

MacromoleculeName: Alpha-mammal toxin AaH2 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Androctonus australis (Sahara scorpion)
Molecular weightTheoretical: 7.260175 KDa
SequenceString:
VKDGYIVDDV NCTYFCGRNA YCNEECTKLK GESGYCQWAS PYGNACYCYK LPDHVRTKGP GRCH(NH2)

UniProtKB: Alpha-mammal toxin AaH2

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Macromolecule #4: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 4 / Number of copies: 5 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #5: (7E,21R,24S)-27-amino-24-hydroxy-18,24-dioxo-19,23,25-trioxa-24la...

MacromoleculeName: (7E,21R,24S)-27-amino-24-hydroxy-18,24-dioxo-19,23,25-trioxa-24lambda~5~-phosphaheptacos-7-en-21-yl (9Z,12E)-octadeca-9,12-dienoate
type: ligand / ID: 5 / Number of copies: 5 / Formula: 76F
Molecular weightTheoretical: 742.018 Da
Chemical component information

ChemComp-76F:
(7E,21R,24S)-27-amino-24-hydroxy-18,24-dioxo-19,23,25-trioxa-24lambda~5~-phosphaheptacos-7-en-21-yl (9Z,12E)-octadeca-9,12-dienoate

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Macromolecule #6: Digitonin

MacromoleculeName: Digitonin / type: ligand / ID: 6 / Number of copies: 1 / Formula: AJP
Molecular weightTheoretical: 1.229312 KDa
Chemical component information

ChemComp-AJP:
Digitonin / detergent*YM

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Macromolecule #7: 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE

MacromoleculeName: 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE / type: ligand / ID: 7 / Number of copies: 1 / Formula: LHG
Molecular weightTheoretical: 722.97 Da
Chemical component information

ChemComp-LHG:
1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE / phospholipid*YM

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Macromolecule #8: CHOLESTEROL HEMISUCCINATE

MacromoleculeName: CHOLESTEROL HEMISUCCINATE / type: ligand / ID: 8 / Number of copies: 1 / Formula: Y01
Molecular weightTheoretical: 486.726 Da
Chemical component information

ChemComp-Y01:
CHOLESTEROL HEMISUCCINATE

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1.5 mg/mL
BufferpH: 7.5
GridModel: C-flat-2/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: PLASMA CLEANING
Details: Grids were plasma etched using the Solarus plasma cleaner (Gatan) in the hydrogen-oxygen setting. Grids were etched for 4 minutes on each side to remove burrs from hole edges. The grids were ...Details: Grids were plasma etched using the Solarus plasma cleaner (Gatan) in the hydrogen-oxygen setting. Grids were etched for 4 minutes on each side to remove burrs from hole edges. The grids were then coated on both sides with 5nm of Au/Pd which was plasma deposited using the Leica ACE600 (Leica).
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 14886 / Average exposure time: 10.0 sec. / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionDetails: Automated picking using a soft-edged disc as a template yielded 842,024 candidate coordinates. 2D classification led to the selection of 504,252 particles for further analysis.
CTF correctionSoftware - Name: cisTEM / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: EMDB MAP
EMDB ID:

6698


Details: The startup map was low-pass filtered to 40A and resolution-limited automatic refinement in cisTEM was used (beginning at 20A)
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cisTEM
Details: No information from spatial frequencies higher than 1/5A were ever used during refinement.
Number images used: 251714
Initial angle assignmentType: OTHER / Software - Name: cisTEM
Final angle assignmentType: OTHER / Software - Name: cisTEM
Final 3D classificationSoftware - Name: cisTEM
FSC plot (resolution estimation)

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