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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: 3596
TitleThe structure of the ESX-5 mycobacterial type VII secretion system membrane complex
Map data
SampleESX-5 T7SS membrane complex
SourceMycobacterium xenopi rivm700367 / / bacteria
Methodsingle particle reconstruction / negative staining / 13.1 Å resolution
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 / 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.
DateDeposition: Feb 17, 2017 / Header (metadata) release: Apr 5, 2017 / Map release: Apr 12, 2017 / Last update: Aug 2, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1
  • Imaged by UCSF Chimera
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  • 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

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Map

Fileemd_3596.map.gz (map file in CCP4 format, 131073 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
320 pix
2.18 Å/pix.
= 697.6 Å
320 pix
2.18 Å/pix.
= 697.6 Å
320 pix
2.18 Å/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
Contour Level:1 (by author), 1 (movie #1):
Minimum - Maximum-1.8008946 - 2.1403747
Average (Standard dev.)0.0028283054 (0.13325706)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions320320320
Origin000
Limit319319319
Spacing320320320
CellA=B=C: 697.60004 Å
α=β=γ: 90 deg.

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
start NX/NY/NZ
NX/NY/NZ
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 / Details: Solubilized in amphipol A8-35 / Number of components: 1
MassTheoretical: 1.8 MDa

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Component #1: protein, ESX-5 T7SS membrane complex

ProteinName: ESX-5 T7SS membrane complex / Details: Solubilized in amphipol A8-35 / Recombinant expression: No
MassTheoretical: 1.8 MDa
SourceSpecies: Mycobacterium xenopi rivm700367 / / bacteria / Strain: RIVM700367
Source (engineered)Expression System: Mycobacterium smegmatis str. mc2 155 / / bacteria
Vector: pMV

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: negative staining
Sample solutionSpecimen conc.: 0.1 mg/ml
Buffer solution: 20 mM Tris pH 8, 100 mM NaCl, 5 % (v/v) glycerol
pH: 8
StainingA drop of 2% (w/v) uranyl acetate was added to a carbon-coated grid with absorbed protein and blotted after 30 s.
VitrificationCryogen name: NONE

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Electron microscopy imaging

ImagingMicroscope: FEI TECNAI 20
Electron gunElectron source: LAB6 / Accelerating voltage: 200 kV / Electron dose: 20 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 50000 X (nominal), 44000 X (calibrated) / Cs: 1.2 mm / Imaging mode: BRIGHT FIELD / Defocus: 1400 - 5000 nm
Specimen HolderModel: SIDE ENTRY, EUCENTRIC
CameraDetector: FEI EAGLE (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 65

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C6 (6 fold cyclic) / Number of projections: 1418
3D reconstructionAlgorithm: BACK PROJECTION / Software: EMAN / Resolution: 13.1 Å / Resolution method: FSC 0.143 CUT-OFF

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