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- EMDB-10188: Structure of EccB3 dimer from the ESX-3 core complex -

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Basic information

Entry
Database: EMDB / ID: EMD-10188
TitleStructure of EccB3 dimer from the ESX-3 core complex
Map data
SampleEccB3 dimeric structure from ESX-3/Type VII secretion system:
ESX-3 secretion system protein EccB3
Function / homologyType VII secretion system EccB, repeat 3 domain / Type VII secretion system EccB / Type VII secretion system ESX-1, transport TM domain B / Hydrolases, Acting on acid anhydrides / hydrolase activity / integral component of membrane / ATP binding / plasma membrane / ESX-3 secretion system ATPase EccB3
Function and homology information
Biological speciesMycolicibacterium smegmatis MC2 155 (bacteria) / Mycobacterium smegmatis (strain ATCC 700084 / mc(2)155) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.6 Å
AuthorsRivera-Calzada A / Famelis N / Geibel S / Llorca O
Funding support Germany, Spain, 6 items
OrganizationGrant numberCountry
German Research FoundationElite Network of Bavaria (N-BM-2013-246), Bavarian State Ministry of Science and the Arts Germany
European Regional Development FundY2018/BIO4747 co-funded by the Autonomous Region of Madrid Spain
Spanish Ministry of Science, Innovation, and UniversitiesSAF2017-82632-P o-funded by the European Regional Development Fund Spain
European UnionHorizon 2020, iNEXT (PID2907) , Grant number 653706 Spain
European Regional Development FundP2018/NMT4443 co-funded by Autonomous Region of Madrid Spain
Bavarian State Ministry of Science and the Arts Funding OrganisationN-BM-2013-246 Germany
CitationJournal: Nature / Year: 2019
Title: Architecture of the ESX-3/Type VII secretion system
Authors: Famelis N / Rivera-Calzada A / Degliesposti G / Wingender M / Mietrach N / Skehel JM / Fernandez-Leiro R / Bottcher B / Schlosser A / Llorca O / Geibel S
Validation ReportPDB-ID: 6sgy

SummaryFull reportAbout validation report
History
DepositionAug 5, 2019-
Header (metadata) releaseSep 11, 2019-
Map releaseOct 9, 2019-
UpdateOct 9, 2019-
Current statusOct 9, 2019Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6sgy
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_10188.map.gz / Format: CCP4 / Size: 107.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 304 pix.
= 323.304 Å
1.06 Å/pix.
x 304 pix.
= 323.304 Å
1.06 Å/pix.
x 304 pix.
= 323.304 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.0635 Å
Density
Contour LevelBy AUTHOR: 0.03 / Movie #1: 0.03
Minimum - Maximum-0.048456762 - 0.0922983
Average (Standard dev.)0.000056797973 (±0.0018579063)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions304304304
Spacing304304304
CellA=B=C: 323.30402 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.06351.06351.0635
M x/y/z304304304
origin x/y/z0.0000.0000.000
length x/y/z323.304323.304323.304
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS304304304
D min/max/mean-0.0480.0920.000

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Supplemental data

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Sample components

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Entire EccB3 dimeric structure from ESX-3/Type VII secretion system

EntireName: EccB3 dimeric structure from ESX-3/Type VII secretion system
Details: The sample consists of four protein components, EccB3:EccC3:EccD3:EccE3 in a 1:1:2:1 stoichiometry Molecular weight of the complex without the amphipol micelle: 0.65 MDa
Number of components: 2

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Component #1: protein, EccB3 dimeric structure from ESX-3/Type VII secretion system

ProteinName: EccB3 dimeric structure from ESX-3/Type VII secretion system
Details: The sample consists of four protein components, EccB3:EccC3:EccD3:EccE3 in a 1:1:2:1 stoichiometry Molecular weight of the complex without the amphipol micelle: 0.65 MDa
Recombinant expression: No
MassTheoretical: 83 kDa
SourceSpecies: Mycolicibacterium smegmatis MC2 155 (bacteria)
Source (engineered)Expression System: Mycolicibacterium smegmatis MC2 155 (bacteria)

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Component #2: protein, ESX-3 secretion system protein EccB3

ProteinName: ESX-3 secretion system protein EccB3 / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 42.746754 kDa
SourceSpecies: Mycobacterium smegmatis (strain ATCC 700084 / mc(2)155) (bacteria)
Source (engineered)Expression System: Mycolicibacterium smegmatis MC2 155 (bacteria)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.3 mg/mL / Buffer solution: 30 mM Hepes pH 8.0, 150 mM NaCl / pH: 8
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 278 K / Humidity: 100 % / Details: Blotting time: 3s Blotting force: -10.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 50 e/Å2 / Illumination mode: SPOT SCAN
LensMagnification: 75000.0 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 1600.0 - 2600.0 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: FEI FALCON III (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 11903
Details: Images acquired as 55 frames movies at a calibrated magnification of 1.0635 Angs/px

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 55342 / Details: Images were collected in counting mode
3D reconstructionSoftware: RELION / Resolution: 4.6 Å / Resolution method: FSC 0.143 CUT-OFF
Details: Particles were extracted centred at extracellular density instead than in the centre of mass in order to improve the resolution in that distal region. Extra density not corresponding to the EccB3 dimer was subtracted
FSC plot (resolution estimation)

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Atomic model buiding

Modeling #1Target criteria: Map correlation coefficient / Refinement space: REAL
Details: Two copies of the periplasmic domain of a EccB3 homology model (based on pdb 3X3M) were fitted into the density and then morphing was performed using PHENIX Real-Space Refinement.
Input PDB model: 3X3M
Chain ID: A
Output model

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