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- EMDB-3596: The structure of the ESX-5 mycobacterial type VII secretion syste... -

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Basic information

Entry
Database: EMDB / ID: EMD-3596
TitleThe structure of the ESX-5 mycobacterial type VII secretion system membrane complex
Map data
Sample
  • Complex: ESX-5 T7SS membrane complex
Biological speciesMycobacterium xenopi RIVM700367 (bacteria)
Methodsingle particle reconstruction / negative staining / Resolution: 13.1 Å
AuthorsCiccarelli L / Marlovits TC
CitationJournal: Nat Microbiol / Year: 2017
Title: Structure of the mycobacterial ESX-5 type VII secretion system membrane complex by single-particle analysis.
Authors: Katherine S H Beckham / Luciano Ciccarelli / Catalin M Bunduc / Haydyn D T Mertens / Roy Ummels / Wolfgang Lugmayr / Julia Mayr / Mandy Rettel / Mikhail M Savitski / Dmitri I Svergun / ...Authors: Katherine S H Beckham / Luciano Ciccarelli / Catalin M Bunduc / Haydyn D T Mertens / Roy Ummels / Wolfgang Lugmayr / Julia Mayr / Mandy Rettel / Mikhail M Savitski / Dmitri I Svergun / Wilbert Bitter / Matthias Wilmanns / Thomas C Marlovits / Annabel H A Parret / Edith N G Houben /
Abstract: Mycobacteria are characterized by their impermeable outer membrane, which is rich in mycolic acids. To transport substrates across this complex cell envelope, mycobacteria rely on type VII (also ...Mycobacteria are characterized by their impermeable outer membrane, which is rich in mycolic acids. To transport substrates across this complex cell envelope, mycobacteria rely on type VII (also known as ESX) secretion systems. In Mycobacterium tuberculosis, these ESX systems are essential for growth and full virulence and therefore represent an attractive target for anti-tuberculosis drugs. However, the molecular details underlying type VII secretion are largely unknown, due to a lack of structural information. Here, we report the molecular architecture of the ESX-5 membrane complex from Mycobacterium xenopi determined at 13 Å resolution by electron microscopy. The four core proteins of the ESX-5 complex (EccB, EccC, EccD and EccE) assemble with equimolar stoichiometry into an oligomeric assembly that displays six-fold symmetry. This membrane-associated complex seems to be embedded exclusively in the inner membrane, which indicates that additional components are required to translocate substrates across the mycobacterial outer membrane. Furthermore, the extended cytosolic domains of the EccC ATPase, which interact with secretion effectors, are highly flexible, suggesting an as yet unseen mode of substrate interaction. Comparison of our results with known structures of other bacterial secretion systems demonstrates that the architecture of type VII secretion system is fundamentally different, suggesting an alternative secretion mechanism.
History
DepositionFeb 17, 2017-
Header (metadata) releaseApr 5, 2017-
Map releaseApr 12, 2017-
UpdateAug 2, 2017-
Current statusAug 2, 2017Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 1
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_3596.jpg

emd_3596.png

Downloads & links

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Map

FileDownload / File: emd_3596.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.18 Å/pix.
x 320 pix.
= 697.6 Å		2.18 Å/pix.
x 320 pix.
= 697.6 Å		2.18 Å/pix.
x 320 pix.
= 697.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.18 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1. / Movie #1: 1
Minimum - Maximum-1.8008946 - 2.1403747
Average (Standard dev.)0.0028283054 (±0.13325706)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 697.60004 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.182.182.18
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z697.600697.600697.600
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-1.8012.1400.003

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Supplemental data

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Sample components

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Entire : ESX-5 T7SS membrane complex

EntireName: ESX-5 T7SS membrane complex
Components
  • Complex: ESX-5 T7SS membrane complex

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Supramolecule #1: ESX-5 T7SS membrane complex

SupramoleculeName: ESX-5 T7SS membrane complex / type: complex / ID: 1 / Parent: 0 / Details: Solubilized in amphipol A8-35
Source (natural)Organism: Mycobacterium xenopi RIVM700367 (bacteria) / Strain: RIVM700367
Recombinant expressionOrganism: Mycobacterium smegmatis str. MC2 155 (bacteria) / Recombinant plasmid: pMV
Molecular weightTheoretical: 1.8 MDa

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.1 mg/mL
BufferpH: 8 / Details: 20 mM Tris pH 8, 100 mM NaCl, 5 % (v/v) glycerol
StainingType: NEGATIVE / Material: Uranyl Acetate
Details: A drop of 2% (w/v) uranyl acetate was added to a carbon-coated grid with absorbed protein and blotted after 30 s.
GridMaterial: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 5.0 nm / Pretreatment - Type: GLOW DISCHARGE

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Electron microscopy

MicroscopeFEI TECNAI 20
Image recordingFilm or detector model: FEI EAGLE (4k x 4k) / Number grids imaged: 1 / Number real images: 65 / Average electron dose: 20.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: LAB6
Electron opticsCalibrated magnification: 44000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 1.2 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 1.4 µm / Nominal magnification: 50000
Sample stageSpecimen holder model: SIDE ENTRY, EUCENTRIC

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Image processing

Particle selectionNumber selected: 1418
Details: A total of 1,418 particles were boxed out manually using e2boxer.py in the EMAN2 software package using a box size of 320 pixels.
CTF correctionSoftware - Name: EMAN (ver. 2)
Startup modelType of model: OTHER
Details: The initial 3D model was obtained using the EMAN2 initial-model generation program e2initialmodel.py without applying symmetry.
Final reconstructionApplied symmetry - Point group: C6 (6 fold cyclic) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 13.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: EMAN (ver. 2) / Number images used: 1418
Initial angle assignmentType: OTHER / Software - Name: EMAN (ver. 2)
Final angle assignmentType: OTHER / Software - Name: EMAN (ver. 2)
Final 3D classificationSoftware - Name: EMAN (ver. 2)

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