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- EMDB-23037: MicroED structure of mVDAC -

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Basic information

Entry
Database: EMDB / ID: EMD-23037
TitleMicroED structure of mVDAC
Map data2Fo-Fc map of mVDAC determined by MicroED
Sample
  • Complex: Murine voltage-dependent anion channel mutant
    • Protein or peptide: Voltage-dependent anion-selective channel protein 1
KeywordsChannel / mammalian / voltage dependent / TRANSPORT PROTEIN
Function / homology
Function and homology information


Pyruvate metabolism / negative regulation of calcium import into the mitochondrion / positive regulation of parkin-mediated stimulation of mitophagy in response to mitochondrial depolarization / voltage-gated monoatomic anion channel activity / PINK1-PRKN Mediated Mitophagy / neuron-neuron synaptic transmission / monoatomic anion channel activity / mitochondrial calcium ion transmembrane transport / acetyl-CoA biosynthetic process from pyruvate / ceramide binding ...Pyruvate metabolism / negative regulation of calcium import into the mitochondrion / positive regulation of parkin-mediated stimulation of mitophagy in response to mitochondrial depolarization / voltage-gated monoatomic anion channel activity / PINK1-PRKN Mediated Mitophagy / neuron-neuron synaptic transmission / monoatomic anion channel activity / mitochondrial calcium ion transmembrane transport / acetyl-CoA biosynthetic process from pyruvate / ceramide binding / regulation of mitophagy / mitochondrial permeability transition pore complex / phosphatidylcholine binding / Ub-specific processing proteases / oxysterol binding / monoatomic anion transport / cholesterol binding / porin activity / mitochondrial nucleoid / negative regulation of reactive oxygen species metabolic process / behavioral fear response / presynaptic active zone membrane / epithelial cell differentiation / learning / mitochondrial membrane / postsynaptic density membrane / synaptic vesicle / myelin sheath / chemical synaptic transmission / mitochondrial inner membrane / mitochondrial outer membrane / transmembrane transporter binding / membrane raft / nucleotide binding / apoptotic process / protein-containing complex binding / negative regulation of apoptotic process / protein kinase binding / protein-containing complex / mitochondrion / membrane / identical protein binding / plasma membrane
Similarity search - Function
Eukaryotic mitochondrial porin signature. / Porin, eukaryotic type / Eukaryotic porin/Tom40 / Eukaryotic porin / Porin domain superfamily
Similarity search - Domain/homology
Voltage-dependent anion-selective channel protein 1
Similarity search - Component
Biological speciesMus musculus (house mouse)
Methodelectron crystallography / cryo EM / Resolution: 3.12 Å
AuthorsMartynowycz MW / Khan F
Funding support United States, 3 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)P41GM136508 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM135175 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2020
Title: MicroED structure of lipid-embedded mammalian mitochondrial voltage-dependent anion channel.
Authors: Michael W Martynowycz / Farha Khan / Johan Hattne / Jeff Abramson / Tamir Gonen /
Abstract: A structure of the murine voltage-dependent anion channel (VDAC) was determined by microcrystal electron diffraction (MicroED). Microcrystals of an essential mutant of VDAC grew in a viscous bicelle ...A structure of the murine voltage-dependent anion channel (VDAC) was determined by microcrystal electron diffraction (MicroED). Microcrystals of an essential mutant of VDAC grew in a viscous bicelle suspension, making it unsuitable for conventional X-ray crystallography. Thin, plate-like crystals were identified using scanning-electron microscopy (SEM). Crystals were milled into thin lamellae using a focused-ion beam (FIB). MicroED data were collected from three crystal lamellae and merged for completeness. The refined structure revealed unmodeled densities between protein monomers, indicative of lipids that likely mediate contacts between the proteins in the crystal. This body of work demonstrates the effectiveness of milling membrane protein microcrystals grown in viscous media using a focused ion beam for subsequent structure determination by MicroED. This approach is well suited for samples that are intractable by X-ray crystallography. To our knowledge, the presented structure is a previously undescribed mutant of the membrane protein VDAC, crystallized in a lipid bicelle matrix and solved by MicroED.
History
DepositionNov 25, 2020-
Header (metadata) releaseDec 23, 2020-
Map releaseDec 23, 2020-
UpdateMar 6, 2024-
Current statusMar 6, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.96644
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.96644
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7kuh
  • Surface level: 0.96644
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7kuh
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23037.png

Downloads & links

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Map

FileDownload / File: emd_23037.map.gz / Format: CCP4 / Size: 2.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation2Fo-Fc map of mVDAC determined by MicroED
Voxel sizeX: 0.77266 Å / Y: 0.77613 Å / Z: 0.77267 Å
Density
Contour LevelBy AUTHOR: 0.96644 / Movie #1: 0.96644
Minimum - Maximum-4.105768 - 6.4787436
Average (Standard dev.)-0.00658367 (±0.9664383)
SymmetrySpace group: 5
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin4-15-19
Dimensions848581
Spacing1287590
CellA: 98.9 Å / B: 58.21 Å / C: 69.54 Å
α: 90.0 ° / β: 99.444 ° / γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.772656250.776133333333330.77266666666667
M x/y/z1287590
origin x/y/z0.0000.0000.000
length x/y/z98.90058.21069.540
α/β/γ90.00099.44490.000
start NX/NY/NZ-194-15
NX/NY/NZ818485
MAP C/R/S321
start NC/NR/NS-154-19
NC/NR/NS858481
D min/max/mean-4.1066.479-0.007

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Supplemental data

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Sample components

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Entire : Murine voltage-dependent anion channel mutant

EntireName: Murine voltage-dependent anion channel mutant
Components
  • Complex: Murine voltage-dependent anion channel mutant
    • Protein or peptide: Voltage-dependent anion-selective channel protein 1

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Supramolecule #1: Murine voltage-dependent anion channel mutant

SupramoleculeName: Murine voltage-dependent anion channel mutant / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 32.87 KDa

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Macromolecule #1: Voltage-dependent anion-selective channel protein 1

MacromoleculeName: Voltage-dependent anion-selective channel protein 1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 32.195879 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MRGSHHHHHH GSMAVPPTYA DLGESARDVF TKGYGFGLIK LDLKTKSENG LEFTSSGSAN TETTKVNGSL ETKYRWTEYG LTFTEKWNT DNTLGTEITV EDQLARGLKL TFDSSFSPNT GKKNAKIKTG YKREHINLGC DVDFDIAGPS IRGALVLGYE G WLAGYQMN ...String:
MRGSHHHHHH GSMAVPPTYA DLGESARDVF TKGYGFGLIK LDLKTKSENG LEFTSSGSAN TETTKVNGSL ETKYRWTEYG LTFTEKWNT DNTLGTEITV EDQLARGLKL TFDSSFSPNT GKKNAKIKTG YKREHINLGC DVDFDIAGPS IRGALVLGYE G WLAGYQMN FETSKSRVTQ SNFAVGYKTD EFQLHTNVND GTEFGGSIYQ KVNKKLETAV NLAWTAGNSN TRFGIAAKYQ VD PDACFSA KVNNSSLIGL GYTQTLKPGI KLTLSALLDG KNVNAGGHKL GLGLEFQA

UniProtKB: Voltage-dependent anion-selective channel protein 1

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron crystallography
Aggregation state3D array

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Sample preparation

Concentration12 mg/mL
BufferpH: 8.5
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Support film - Film thickness: 12 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 277 K / Instrument: LEICA EM GP
DetailsMonodisperse crystal embedded in lipid bicelles

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: DIFFRACTION / Cs: 2.7 mm / Nominal defocus max: 0.0 µm / Nominal defocus min: 0.0 µm / Camera length: 3000 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN / Tilt angle: -60.0, 68.0
TemperatureMin: 77.0 K / Max: 90.0 K
Image recordingFilm or detector model: FEI CETA (4k x 4k) / Digitization - Dimensions - Width: 2048 pixel / Digitization - Dimensions - Height: 2048 pixel / Number grids imaged: 2 / Number real images: 150 / Number diffraction images: 180 / Average exposure time: 5.0 sec. / Average electron dose: 0.01 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Crystallography statisticsNumber intensities measured: 32723 / Number structure factors: 5410 / Fourier space coverage: 76.29 / R sym: 0.22 / R merge: 0.47 / Overall phase error: 25 / Overall phase residual: 25 / Phase error rejection criteria: none used / High resolution: 3.12 Å / Shell - Shell ID: 1 / Shell - High resolution: 3.232 Å / Shell - Low resolution: 3.12 Å / Shell - Number structure factors: 565 / Shell - Phase residual: 27 / Shell - Fourier space coverage: 57.5 / Shell - Multiplicity: 6.2
Molecular replacementSoftware - Name: PHENIX / Software - details: Phaser
Symmetry determination software listSoftware - details: XDS
Final reconstructionAlgorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.12 Å / Resolution method: DIFFRACTION PATTERN/LAYERLINES / Software - Name: PHENIX
Merging software listSoftware - Name: AIMLESS
DetailsCetaD 2x binned

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Atomic model buiding 1

Initial modelPDB ID:

3emn


Chain - Source name: PDB / Chain - Initial model type: experimental model
DetailsStandard refinement with electron scattering factors
RefinementSpace: RECIPROCAL / Protocol: RIGID BODY FIT / Overall B value: 90 / Target criteria: Maximum liklihood
Output model

PDB-7kuh:
MicroED structure of mVDAC

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