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- EMDB-16683: Cryo-EM structure of the CupE pilus from Pseudomonas aeruginosa -

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Basic information

Entry
Database: EMDB / ID: EMD-16683
TitleCryo-EM structure of the CupE pilus from Pseudomonas aeruginosa
Map data
Sample
  • Complex: CupE pilus
    • Protein or peptide: SCPU domain-containing protein
Function / homologySpore coat protein U / Spore Coat Protein U domain / Spore Coat Protein U domain / Spore coat protein U domain-containing protein
Function and homology information
Biological speciesPseudomonas aeruginosa (bacteria) / Pseudomonas aeruginosa PAO1 (bacteria)
Methodhelical reconstruction / cryo EM / Resolution: 3.5 Å
AuthorsBoehning J / Bharat TAM
Funding support United Kingdom, United States, 7 items
OrganizationGrant numberCountry
Wellcome Trust202231/Z/16/Z United Kingdom
Royal Society202231/Z/16/Z United Kingdom
The Vallee Foundation Inc. United States
Leverhulme Trust United Kingdom
The Lister Institute of Preventive Medicine United Kingdom
Medical Research Council (MRC, United Kingdom)MR/K501256/1 United Kingdom
Medical Research Council (MRC, United Kingdom)MR/N013468/1 United Kingdom
CitationJournal: PLoS Pathog / Year: 2023
Title: Architecture of the biofilm-associated archaic Chaperone-Usher pilus CupE from Pseudomonas aeruginosa.
Authors: Jan Böhning / Adrian W Dobbelstein / Nina Sulkowski / Kira Eilers / Andriko von Kügelgen / Abul K Tarafder / Sew-Yeu Peak-Chew / Mark Skehel / Vikram Alva / Alain Filloux / Tanmay A M Bharat /
Abstract: Chaperone-Usher Pathway (CUP) pili are major adhesins in Gram-negative bacteria, mediating bacterial adherence to biotic and abiotic surfaces. While classical CUP pili have been extensively ...Chaperone-Usher Pathway (CUP) pili are major adhesins in Gram-negative bacteria, mediating bacterial adherence to biotic and abiotic surfaces. While classical CUP pili have been extensively characterized, little is known about so-called archaic CUP pili, which are phylogenetically widespread and promote biofilm formation by several human pathogens. In this study, we present the electron cryomicroscopy structure of the archaic CupE pilus from the opportunistic human pathogen Pseudomonas aeruginosa. We show that CupE1 subunits within the pilus are arranged in a zigzag architecture, containing an N-terminal donor β-strand extending from each subunit into the next, where it is anchored by hydrophobic interactions, with comparatively weaker interactions at the rest of the inter-subunit interface. Imaging CupE pili on the surface of P. aeruginosa cells using electron cryotomography shows that CupE pili adopt variable curvatures in response to their environment, which might facilitate their role in promoting cellular attachment. Finally, bioinformatic analysis shows the widespread abundance of cupE genes in isolates of P. aeruginosa and the co-occurrence of cupE with other cup clusters, suggesting interdependence of cup pili in regulating bacterial adherence within biofilms. Taken together, our study provides insights into the architecture of archaic CUP pili, providing a structural basis for understanding their role in promoting cellular adhesion and biofilm formation in P. aeruginosa.
History
DepositionFeb 10, 2023-
Header (metadata) releaseMar 22, 2023-
Map releaseMar 22, 2023-
UpdateApr 26, 2023-
Current statusApr 26, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16683.png

Downloads & links

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Map

FileDownload / File: emd_16683.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.09 Å/pix.
x 280 pix.
= 305.76 Å		1.09 Å/pix.
x 280 pix.
= 305.76 Å		1.09 Å/pix.
x 280 pix.
= 305.76 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.092 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.03
Minimum - Maximum-0.097849645 - 0.11703034
Average (Standard dev.)9.119918e-05 (±0.0021411313)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 305.76 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_16683_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_16683_half_map_2.map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Sample components

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Entire : CupE pilus

EntireName: CupE pilus
Components
  • Complex: CupE pilus
    • Protein or peptide: SCPU domain-containing protein

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Supramolecule #1: CupE pilus

SupramoleculeName: CupE pilus / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Pseudomonas aeruginosa (bacteria) / Strain: PAO1 mutant / Location in cell: Cell surface

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Macromolecule #1: SCPU domain-containing protein

MacromoleculeName: SCPU domain-containing protein / type: protein_or_peptide / ID: 1 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa PAO1 (bacteria)
Molecular weightTheoretical: 16.127815 KDa
SequenceString:
AGTLIGQVGV QMVIGAGCTI INGSVSGGIN QWGTLDFGSH SDLTNVVDAQ TVGTSGNIQI QCSTGLTPSL TVNAGLHASG GQRYMQNTT TTSSTIAYNI YSDAARSALI QANTPVDISS VSTGTAVNIP LYGRVVPTGQ STPTPTAGTY TDTLLVTIAW

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7.4 / Details: Phosphate-buffered saline
GridModel: Quantifoil R2/2 / Material: COPPER/RHODIUM / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 20 sec. / Details: 15 mA
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 81000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Average electron dose: 46.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Segment selectionNumber selected: 3766858 / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: NOT APPLICABLE
Final reconstructionApplied symmetry - Helical parameters - Δz: 33.04 Å
Applied symmetry - Helical parameters - Δ&Phi: -145.55 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 274457
FSC plot (resolution estimation)

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Atomic model buiding 1

DetailsInitial local fitting of a homology model was done using ChimeraX, and the structure manually re-built in Coot.
RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-8cio:
Cryo-EM structure of the CupE pilus from Pseudomonas aeruginosa

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