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- PDB-6hs7: Type VI membrane complex -

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Basic information

Entry
Database: PDB / ID: 6hs7
TitleType VI membrane complex
Components
  • ImcF-like family protein
  • Type VI secretion system protein VasD
KeywordsMEMBRANE PROTEIN / Membrane complex / tether
Function / homology
Function and homology information


membrane => GO:0016020
Similarity search - Function
IcmF-related / Intracellular multiplication and human macrophage-killing / Type VI secretion system IcmF, C-terminal / Type VI secretion system, lipoprotein SciN / Type VI secretion protein SciN-like superfamily / Type VI secretion protein IcmF C2-like domain / Type VI secretion lipoprotein, VasD, EvfM, TssJ, VC_A0113 / Prokaryotic membrane lipoprotein lipid attachment site profile.
Similarity search - Domain/homology
Putative type VI secretion protein / Type VI secretion system protein VasD / ImcF-like family protein
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.6 Å
AuthorsRapisarda, C. / Fronzes, R.
Funding support France, 1items
OrganizationGrant numberCountry
France
CitationJournal: EMBO J / Year: 2019
Title: and high-resolution cryo-EM structure of a bacterial type VI secretion system membrane complex.
Authors: Chiara Rapisarda / Yassine Cherrak / Romain Kooger / Victoria Schmidt / Riccardo Pellarin / Laureen Logger / Eric Cascales / Martin Pilhofer / Eric Durand / Rémi Fronzes /
Abstract: Bacteria have evolved macromolecular machineries that secrete effectors and toxins to survive and thrive in diverse environments. The type VI secretion system (T6SS) is a contractile machine that is ...Bacteria have evolved macromolecular machineries that secrete effectors and toxins to survive and thrive in diverse environments. The type VI secretion system (T6SS) is a contractile machine that is related to phages. It is composed of a phage tail-like structure inserted in the bacterial cell envelope by a membrane complex (MC) comprising the TssJ, TssL and TssM proteins. We previously reported the low-resolution negative-stain electron microscopy structure of the enteroaggregative MC and proposed a rotational 5-fold symmetry with a TssJ:TssL:TssM stoichiometry of 2:2:2. Here, cryo-electron tomography analyses of the T6SS MC confirm the 5-fold symmetry and identify the regions of the structure that insert into the bacterial membranes. A high-resolution model obtained by single-particle cryo-electron microscopy highlights new features: five additional copies of TssJ, yielding a TssJ:TssL:TssM stoichiometry of 3:2:2, an 11-residue loop in TssM, protruding inside the lumen of the MC and constituting a functionally important periplasmic gate, and hinge regions. Based on these data, we propose an updated model on MC structure and dynamics during T6SS assembly and function.
History
DepositionSep 28, 2018Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 27, 2019Provider: repository / Type: Initial release
Revision 1.1May 15, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
A: ImcF-like family protein
B: ImcF-like family protein
C: ImcF-like family protein
D: ImcF-like family protein
E: ImcF-like family protein
F: Type VI secretion system protein VasD
G: Type VI secretion system protein VasD
H: Type VI secretion system protein VasD
I: Type VI secretion system protein VasD
J: Type VI secretion system protein VasD
K: Type VI secretion system protein VasD
L: Type VI secretion system protein VasD
M: Type VI secretion system protein VasD
N: Type VI secretion system protein VasD
O: Type VI secretion system protein VasD
P: Type VI secretion system protein VasD
Q: Type VI secretion system protein VasD
R: Type VI secretion system protein VasD
S: Type VI secretion system protein VasD
T: Type VI secretion system protein VasD
a: ImcF-like family protein
b: ImcF-like family protein
c: ImcF-like family protein
d: ImcF-like family protein
e: ImcF-like family protein


Theoretical massNumber of molelcules
Total (without water)1,577,25225
Polymers1,577,25225
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area63280 Å2
ΔGint-199 kcal/mol
Surface area316140 Å2
MethodPISA

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Components

#1: Protein
ImcF-like family protein


Mass: 127255.367 Da / Num. of mol.: 10
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: EC90111_2427 / Production host: Escherichia coli (E. coli) / References: UniProt: I2W7L4
#2: Protein
Type VI secretion system protein VasD


Mass: 20313.236 Da / Num. of mol.: 15
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: vasD, ECDEC6A_3651 / Production host: Escherichia coli (E. coli) / References: UniProt: H4UNW1, UniProt: A0A377LAC1*PLUS

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Membrane complex of the type VI secretion system (TssM and TssJ)
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Escherichia coli (E. coli)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
SpecimenConc.: 0.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: C-flat-2/2
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 120000 X / Nominal defocus max: 2 nm / Nominal defocus min: 0.5 nm / Calibrated defocus max: 3 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 120 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

EM software
IDNameVersionCategory
1RELION2.1particle selection
2EPUimage acquisition
4GctfCTF correction
10cryoSPARCinitial Euler assignment
11RELIONfinal Euler assignment
13RELION3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C5 (5 fold cyclic)
3D reconstructionResolution: 4.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 36828 / Symmetry type: POINT

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