|Entry||Database: PDB / ID: 6gv9|
|Title||Structure of the type IV pilus from enterohemorrhagic Escherichia coli (EHEC)|
|Components||Prepilin peptidase-dependent protein D|
|Keywords||PROTEIN FIBRIL / TYPE IV PILI EHEC T4P CELL ADHESION hemorrhagic coli pilus (HCP)|
|Function / homology||Prokaryotic N-terminal methylation site / Prokaryotic N-terminal methylation motif / Prokaryotic N-terminal methylation site. / Prepilin peptidase-dependent pilin|
Function and homology information
|Specimen source||Escherichia coli O157:H7 (bacteria)|
|Method||ELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 8 Å|
|Authors||Bardiaux, B. / Amorim, G.C. / Luna-Rico, A. / Zheng, W. / Guilvout, I. / Jollivet, C. / Nilges, M. / Egelman, E. / Francetic, O. / Izadi-Pruneyre, N.|
|Funding support||France , 1件 |
|Citation||Journal: Structure / Year: 2019|
Title: Structure and Assembly of the Enterohemorrhagic Escherichia coli Type 4 Pilus.
Authors: Benjamin Bardiaux / Gisele Cardoso de Amorim / Areli Luna Rico / Weili Zheng / Ingrid Guilvout / Camille Jollivet / Michael Nilges / Edward H Egelman / Nadia Izadi-Pruneyre / Olivera Francetic /
Abstract: Bacterial type 4a pili are dynamic surface filaments that promote bacterial adherence, motility, and macromolecular transport. Their genes are highly conserved among enterobacteria and their ...Bacterial type 4a pili are dynamic surface filaments that promote bacterial adherence, motility, and macromolecular transport. Their genes are highly conserved among enterobacteria and their expression in enterohemorrhagic Escherichia coli (EHEC) promotes adhesion to intestinal epithelia and pro-inflammatory signaling. To define the molecular basis of EHEC pilus assembly, we determined the structure of the periplasmic domain of its major subunit PpdD (PpdDp), a prototype of an enterobacterial pilin subfamily containing two disulfide bonds. The structure of PpdDp, determined by NMR, was then docked into the density envelope of purified EHEC pili obtained by cryoelectron microscopy (cryo-EM). Cryo-EM reconstruction of EHEC pili at ∼8 Å resolution revealed extremely high pilus flexibility correlating with a large extended region of the pilin stem. Systematic mutagenesis combined with functional and interaction analyses identified charged residues essential for pilus assembly. Structural information on exposed regions and interfaces between EHEC pilins is relevant for vaccine and drug discovery.
SummaryFull reportAbout validation report
|Date||Deposition: Jun 20, 2018 / Release: May 15, 2019Array|
|Structure viewer||Molecule: |
Downloads & links
A: Prepilin peptidase-dependent protein D
B: Prepilin peptidase-dependent protein D
C: Prepilin peptidase-dependent protein D
D: Prepilin peptidase-dependent protein D
E: Prepilin peptidase-dependent protein D
F: Prepilin peptidase-dependent protein D
G: Prepilin peptidase-dependent protein D
H: Prepilin peptidase-dependent protein D
I: Prepilin peptidase-dependent protein D
J: Prepilin peptidase-dependent protein D
K: Prepilin peptidase-dependent protein D
L: Prepilin peptidase-dependent protein D
M: Prepilin peptidase-dependent protein D
N: Prepilin peptidase-dependent protein D
Mass: 14883.821 Da / Num. of mol.: 14
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli O157:H7 (bacteria) / Gene: ECs0112 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q8X974
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: FILAMENT / 3D reconstruction method: helical reconstruction|
|Component||Name: EHEC type IV pili / Type: COMPLEX / Entity ID: 1 / Source: RECOMBINANT|
|Molecular weight||Experimental value: NO|
|Source (natural)||Organism: Escherichia coli O157:H7 (bacteria)|
|Source (recombinant)||Organism: Escherichia coli BL21(DE3) (bacteria)|
|Buffer solution||pH: 7.4|
|Specimen||Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Grid material: COPPER / Grid mesh size: 300 divisions/in.|
|Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 297 K|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Microscopy||Model: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELDBright-field microscopy|
|Image recording||Average exposure time: 2 sec. / Electron dose: 1.4 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1|
|CTF correction||Type: PHASE FLIPPING AND AMPLITUDE CORRECTION|
|Helical symmerty||Angular rotation/subunit: 96 ° / Axial rise/subunit: 11.2 Å / Axial symmetry: C1|
|3D reconstruction||Resolution: 8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 25669 / Symmetry type: HELICAL|
|Atomic model building||Protocol: OTHER / Space: REAL|
Details: Soluble domain structure and homology model of TM segment fitted as rigid-bodies. Linker region built ab initio. Real-space refinement after symmetrization.
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