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- PDB-6n4i: Structural basis of Nav1.7 inhibition by a gating-modifier spider... -

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Basic information

Entry
Database: PDB / ID: 6n4i
TitleStructural basis of Nav1.7 inhibition by a gating-modifier spider toxin
Components
  • Beta/omega-theraphotoxin-Tp2a
  • Nav1.7 VSD2-NavAb channel chimera protein
KeywordsMETAL TRANSPORT / sodium channel / toxin / gating-modifier / voltage-gated
Function / homologytoxin activity / lipid binding / extracellular region / Beta/omega-theraphotoxin-Tp2a
Function and homology information
Specimen sourceHomo sapiens (human)
Thrixopelma pruriens (green velvet)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / 3.541 Å resolution
AuthorsXu, H. / Koth, C.M. / Payandeh, J.
CitationJournal: Cell / Year: 2019
Title: Structural Basis of Nav1.7 Inhibition by a Gating-Modifier Spider Toxin.
Authors: Hui Xu / Tianbo Li / Alexis Rohou / Christopher P Arthur / Foteini Tzakoniati / Evera Wong / Alberto Estevez / Christine Kugel / Yvonne Franke / Jun Chen / Claudio Ciferri / David H Hackos / Christopher M Koth / Jian Payandeh
Abstract: Voltage-gated sodium (Nav) channels are targets of disease mutations, toxins, and therapeutic drugs. Despite recent advances, the structural basis of voltage sensing, electromechanical coupling, and ...Voltage-gated sodium (Nav) channels are targets of disease mutations, toxins, and therapeutic drugs. Despite recent advances, the structural basis of voltage sensing, electromechanical coupling, and toxin modulation remains ill-defined. Protoxin-II (ProTx2) from the Peruvian green velvet tarantula is an inhibitor cystine-knot peptide and selective antagonist of the human Nav1.7 channel. Here, we visualize ProTx2 in complex with voltage-sensor domain II (VSD2) from Nav1.7 using X-ray crystallography and cryoelectron microscopy. Membrane partitioning orients ProTx2 for unfettered access to VSD2, where ProTx2 interrogates distinct features of the Nav1.7 receptor site. ProTx2 positions two basic residues into the extracellular vestibule to antagonize S4 gating-charge movement through an electrostatic mechanism. ProTx2 has trapped activated and deactivated states of VSD2, revealing a remarkable ∼10 Å translation of the S4 helix, providing a structural framework for activation gating in voltage-gated ion channels. Finally, our results deliver key templates to design selective Nav channel antagonists.
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Nov 19, 2018 / Release: Jan 23, 2019
RevisionDateData content typeGroupCategoryItemProviderType
1.0Jan 23, 2019Structure modelrepositoryInitial release
1.1Feb 6, 2019Structure modelData collection / Database references / Structure summarycitation / struct_citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _struct.title

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Nav1.7 VSD2-NavAb channel chimera protein
B: Nav1.7 VSD2-NavAb channel chimera protein
C: Nav1.7 VSD2-NavAb channel chimera protein
D: Nav1.7 VSD2-NavAb channel chimera protein
E: Beta/omega-theraphotoxin-Tp2a
F: Beta/omega-theraphotoxin-Tp2a
G: Beta/omega-theraphotoxin-Tp2a
H: Beta/omega-theraphotoxin-Tp2a
hetero molecules


Theoretical massNumber of molelcules
Total (without water)160,66124
Polyers149,1738
Non-polymers11,48816
Water0
1


TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area (Å2)18970
ΔGint (kcal/M)-173
Surface area (Å2)52740
MethodPISA
Unit cell
γ
α
β
Length a, b, c (Å)221.220, 123.530, 123.990
Angle α, β, γ (deg.)90.00, 124.00, 90.00
Int Tables number5
Space group name H-MC 1 2 1

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Components

#1: Protein/peptide
Nav1.7 VSD2-NavAb channel chimera protein


Mass: 33453.512 Da / Num. of mol.: 4 / Source: (gene. exp.) Homo sapiens (human) / Production host: Trichoplusia ni (cabbage looper)
#2: Protein/peptide
Beta/omega-theraphotoxin-Tp2a / Beta/omega-TRTX-Tp2a / ProTx-II / PT-II / Protoxin-2 / ProTx2


Mass: 3839.687 Da / Num. of mol.: 4 / Source: (gene. exp.) Thrixopelma pruriens (green velvet) / Production host: synthetic construct (others) / References: UniProt: P83476
#3: Chemical
ChemComp-6OU / [(2~{R})-1-[2-azanylethoxy(oxidanyl)phosphoryl]oxy-3-hexadecanoyloxy-propan-2-yl] (~{Z})-octadec-9-enoate


Mass: 717.996 Da / Num. of mol.: 16 / Formula: C39H76NO8P

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 4.71 / Density percent sol: 73.87 %
Crystal growTemp: 292 K / Method: vapor diffusion, sitting drop / pH: 7
Details: 2.3-2.6M Ammonium sulfate, 100 mM HEPES, pH 7.0; 30% sucrose for cryo
PH range: 4.6-7.5

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Data collection

DiffractionMean temperature: 80 kelvins / Serial crystal experiment: N
SourceSource: SYNCHROTRON / Type: APS BEAMLINE 17-ID / Synchrotron site: APS / Beamline: 17-ID / Wavelength: 1
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Collection date: Nov 21, 2016
RadiationDiffraction protocol: SINGLE WAVELENGTH / Monochromatic or laue m l: M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionD resolution high: 3.5 Å / D resolution low: 50 Å / Number obs: 479752 / CC half: 0.999 / Rmerge I obs: 0.115 / NetI over sigmaI: 10.43 / Redundancy: 13.7 % / Percent possible obs: 99.3
Reflection shellRmerge I obs: 0.76 / Highest resolution: 3.5 Å / Lowest resolution: 3.59 Å / Number unique obs: 35190 / CC half: 0.756 / Redundancy: 13.7 % / Percent possible all: 99.6

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Processing

Software
NameVersionClassification
PHENIX(dev_2747: ???)refinement
XDSdata reduction
XSCALEdata scaling
PHENIXphasing
RefineMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5EK0
Overall SU ML: 0.4 / Cross valid method: FREE R-VALUE / Sigma F: 1.37 / Overall phase error: 35.23
Solvent computationSolvent shrinkage radii: 0.9 Å / Solvent vdw probe radii: 1.11 Å
Least-squares processR factor R free: 0.2942 / R factor R work: 0.2745 / R factor obs: 0.2755 / Highest resolution: 3.541 Å / Lowest resolution: 36.892 Å / Number reflection R free: 1373 / Number reflection obs: 27544 / Percent reflection R free: 4.98 / Percent reflection obs: 81.47
Refine hist #LASTHighest resolution: 3.541 Å / Lowest resolution: 36.892 Å
Number of atoms included #LASTProtein: 8312 / Nucleic acid: 0 / Ligand: 317 / Solvent: 0 / Total: 8629
Refine LS restraints
Refine IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0058822
X-RAY DIFFRACTIONf_angle_d0.85411969
X-RAY DIFFRACTIONf_dihedral_angle_d16.6495071
X-RAY DIFFRACTIONf_chiral_restr0.0471374
X-RAY DIFFRACTIONf_plane_restr0.0051396
Refine LS shell

Refine ID: X-RAY DIFFRACTION

Highest resolutionR factor R freeR factor R workLowest resolutionNumber reflection R freeNumber reflection R workPercent reflection obs
3.54090.33720.35683.66732032210.00
3.66730.29720.31013.814048114636.00
3.81400.30940.29103.9874118231772.00
3.98740.30720.28344.1974139310297.00
4.19740.28870.26274.45991663194100.00
4.45990.29710.24764.80361963192100.00
4.80360.27480.24485.28571943194100.00
5.28570.29050.31216.04771433237100.00
6.04770.31890.34007.60851633221100.00
7.60850.28670.250636.8936186324699.00
Refine TLSMethod: refined / L11: 0.7162 deg.2 / L12: -0.2847 deg.2 / L13: -0.0014 deg.2 / L22: 0.7802 deg.2 / L23: -0.0588 deg.2 / L33: 0.7156 deg.2 / S11: -0.1976 Å deg. / S12: -0.0779 Å deg. / S13: -0.1558 Å deg. / S21: -0.0204 Å deg. / S22: 0.2219 Å deg. / S23: -0.0585 Å deg. / S31: -0.1775 Å deg. / S32: 0.1176 Å deg. / S33: 0.0079 Å deg. / T11: 0.1316 Å2 / T12: -0.027 Å2 / T13: -0.0514 Å2 / T22: 0.2093 Å2 / T23: -0.0129 Å2 / T33: 0.1567 Å2 / Origin x: 99.0797 Å / Origin y: 248.6897 Å / Origin z: 216.8423 Å
Refine TLS groupSelection details: all

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