Journal: 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
Deposition
Feb 17, 2017
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Header (metadata) release
Apr 5, 2017
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Map release
Apr 12, 2017
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Update
Aug 2, 2017
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Current status
Aug 2, 2017
Processing site: PDBe / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
pH: 8 / Details: 20 mM Tris pH 8, 100 mM NaCl, 5 % (v/v) glycerol
Staining
Type: 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.
Grid
Material: 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
Microscope
FEI TECNAI 20
Image recording
Film or detector model: FEI EAGLE (4k x 4k) / Number grids imaged: 1 / Number real images: 65 / Average electron dose: 20.0 e/Å2
Electron beam
Acceleration voltage: 200 kV / Electron source: LAB6
Number 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 correction
Software - Name: EMAN (ver. 2)
Startup model
Type of model: OTHER Details: The initial 3D model was obtained using the EMAN2 initial-model generation program e2initialmodel.py without applying symmetry.
Final reconstruction
Applied 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 assignment
Type: OTHER / Software - Name: EMAN (ver. 2)
Final angle assignment
Type: OTHER / Software - Name: EMAN (ver. 2)
Final 3D classification
Software - Name: EMAN (ver. 2)
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