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- PDB-8ehr: Cryo-EM reconstruction of the CFA/I bacterial adhesion pili -

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Basic information

Entry
Database: PDB / ID: 8ehr
TitleCryo-EM reconstruction of the CFA/I bacterial adhesion pili
ComponentsCFA/I fimbrial subunit B
KeywordsCELL ADHESION / enterotoxigenic / adhesion pili / superelastic / helical reconstruction
Function / homologyFimbrial major subunit, CS1-type / CS1 type fimbrial major subunit / pilus / CFA/I fimbrial subunit B
Function and homology information
Biological speciesEscherichia coli (E. coli)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsDoran, M.H. / Bullitt, E.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)1R21AI156236-01 United States
CitationJournal: Structure / Year: 2023
Title: Three structural solutions for bacterial adhesion pilus stability and superelasticity.
Authors: Matthew H Doran / Joseph L Baker / Tobias Dahlberg / Magnus Andersson / Esther Bullitt /
Abstract: Bacterial adhesion pili are key virulence factors that mediate host-pathogen interactions in diverse epithelial environments. Deploying a multimodal approach, we probed the structural basis ...Bacterial adhesion pili are key virulence factors that mediate host-pathogen interactions in diverse epithelial environments. Deploying a multimodal approach, we probed the structural basis underpinning the biophysical properties of pili originating from enterotoxigenic (ETEC) and uropathogenic bacteria. Using cryo-electron microscopy we solved the structures of three vaccine target pili from ETEC bacteria, CFA/I, CS17, and CS20. Pairing these and previous pilus structures with force spectroscopy and steered molecular dynamics simulations, we find a strong correlation between subunit-subunit interaction energies and the force required for pilus unwinding, irrespective of genetic similarity. Pili integrate three structural solutions for stabilizing their assemblies: layer-to-layer interactions, N-terminal interactions to distant subunits, and extended loop interactions from adjacent subunits. Tuning of these structural solutions alters the biophysical properties of pili and promotes the superelastic behavior that is essential for sustained bacterial attachment.
History
DepositionSep 14, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 22, 2023Provider: repository / Type: Initial release
Revision 1.1Apr 12, 2023Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed ..._citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.2May 17, 2023Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: CFA/I fimbrial subunit B
C: CFA/I fimbrial subunit B
D: CFA/I fimbrial subunit B
E: CFA/I fimbrial subunit B
B: CFA/I fimbrial subunit B
F: CFA/I fimbrial subunit B
G: CFA/I fimbrial subunit B


Theoretical massNumber of molelcules
Total (without water)104,7827
Polymers104,7827
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, Forms filament under cryo-conditions
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
CFA/I fimbrial subunit B / CFA/I antigen / CFA/I pilin / Colonization factor antigen I subunit B


Mass: 14968.839 Da / Num. of mol.: 7
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Gene: cfaB / Production host: Escherichia coli (E. coli) / Strain (production host): BL21-AI(pMAM2-cfaB ) / References: UniProt: P0CK93

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: HELICAL ARRAY / 3D reconstruction method: helical reconstruction

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Sample preparation

ComponentName: CFA/I bacterial adhesion pili / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Escherichia coli (E. coli) / Strain: BL21-AI
Source (recombinant)Organism: Escherichia coli (E. coli) / Strain: BL21-AI(pMAM2-cfaB )
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 15 mA with the Pelco EasiGlow Machine / Grid material: GOLD / Grid mesh size: 400 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 283 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 53.7 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of real images: 4809

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Processing

SoftwareName: PHENIX / Version: 1.18_3861: / Classification: refinement
EM software
IDNameVersionCategory
1RELION3.1.1particle selection
2Leginonimage acquisition
4CTFFIND4CTF correction
7PHENIX1.18model fitting
8UCSF Chimera1.14model fitting
10RELION3.1.1initial Euler assignment
11RELION3.1.1final Euler assignment
13RELION3.1.13D reconstruction
14PHENIX1.18model refinement
15ISOLDE1.4model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Helical symmertyAngular rotation/subunit: 113.37 ° / Axial rise/subunit: 8.505 Å / Axial symmetry: C1
Particle selectionNum. of particles selected: 6094726
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 325069 / Symmetry type: HELICAL
Atomic model buildingProtocol: OTHER / Space: REAL
Atomic model buildingPDB-ID: 6NRV

6nrv

Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0027455
ELECTRON MICROSCOPYf_angle_d0.81610199
ELECTRON MICROSCOPYf_dihedral_angle_d4.3891057
ELECTRON MICROSCOPYf_chiral_restr0.0451302
ELECTRON MICROSCOPYf_plane_restr0.0041295

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