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- EMDB-8188: aerolysin post-prepore and quasipore cryo-EM maps. -

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Basic information

Entry
Database: EMDB / ID: EMD-8188
Titleaerolysin post-prepore and quasipore cryo-EM maps.
Map dataaerolysin post-prepore and quasipore
Sample
  • Complex: aerolysin post-prepore and quasipore
    • Protein or peptide: Aerolysin
Function / homology
Function and homology information


toxin activity / host cell plasma membrane / extracellular region / identical protein binding / membrane
Similarity search - Function
Aerolysin / Aerolysin toxin / Aerolysin toxin / Aerolysin/Pertussis toxin domain / Aerolysin/Pertussis toxin (APT), N-terminal domain superfamily / Aerolysin/Pertussis toxin (APT) domain / Aerolysin/haemolysin toxin, conserved site / Aerolysin type toxins signature. / C-type lectin fold
Similarity search - Domain/homology
Biological speciesAeromonas hydrophila (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.46 Å
AuthorsIacovache I / Zuber B
CitationJournal: Nat Commun / Year: 2016
Title: Cryo-EM structure of aerolysin variants reveals a novel protein fold and the pore-formation process.
Authors: Ioan Iacovache / Sacha De Carlo / Nuria Cirauqui / Matteo Dal Peraro / F Gisou van der Goot / Benoît Zuber /
Abstract: Owing to their pathogenical role and unique ability to exist both as soluble proteins and transmembrane complexes, pore-forming toxins (PFTs) have been a focus of microbiologists and structural ...Owing to their pathogenical role and unique ability to exist both as soluble proteins and transmembrane complexes, pore-forming toxins (PFTs) have been a focus of microbiologists and structural biologists for decades. PFTs are generally secreted as water-soluble monomers and subsequently bind the membrane of target cells. Then, they assemble into circular oligomers, which undergo conformational changes that allow membrane insertion leading to pore formation and potentially cell death. Aerolysin, produced by the human pathogen Aeromonas hydrophila, is the founding member of a major PFT family found throughout all kingdoms of life. We report cryo-electron microscopy structures of three conformational intermediates and of the final aerolysin pore, jointly providing insight into the conformational changes that allow pore formation. Moreover, the structures reveal a protein fold consisting of two concentric β-barrels, tightly kept together by hydrophobic interactions. This fold suggests a basis for the prion-like ultrastability of aerolysin pore and its stoichiometry.
History
DepositionMay 17, 2016-
Header (metadata) releaseJul 13, 2016-
Map releaseJul 13, 2016-
UpdateNov 21, 2018-
Current statusNov 21, 2018Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.125
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.125
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-5jzw
  • Surface level: 0.125
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_8188.png

Downloads & links

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Map

FileDownload / File: emd_8188.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationaerolysin post-prepore and quasipore
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.34 Å/pix.
x 200 pix.
= 268. Å		1.34 Å/pix.
x 200 pix.
= 268. Å		1.34 Å/pix.
x 200 pix.
= 268. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.34 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.125 / Movie #1: 0.125
Minimum - Maximum-0.50525826 - 0.750208
Average (Standard dev.)0.0033729975 (±0.031512413)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 268.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.341.341.34
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z268.000268.000268.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ329329329
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.5050.7500.003

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Supplemental data

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Sample components

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Entire : aerolysin post-prepore and quasipore

EntireName: aerolysin post-prepore and quasipore
Components
  • Complex: aerolysin post-prepore and quasipore
    • Protein or peptide: Aerolysin

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Supramolecule #1: aerolysin post-prepore and quasipore

SupramoleculeName: aerolysin post-prepore and quasipore / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Details: Aerolysin oligomer arrested in post-prepore and quasipore states by the K246C/E258C mutation.
Source (natural)Organism: Aeromonas hydrophila (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria) / Recombinant plasmid: PET22b
Molecular weightTheoretical: 350 KDa

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Macromolecule #1: Aerolysin

MacromoleculeName: Aerolysin / type: protein_or_peptide / ID: 1 / Number of copies: 14 / Enantiomer: LEVO
Source (natural)Organism: Aeromonas hydrophila (bacteria)
Molecular weightTheoretical: 47.133145 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: AEPVYPDQLR LFSLGQGVCG DKYRPVNREE AQSVKSNIVG MMGQWQISGL ANGWVIMGPG YNGEIKPGTA SNTWCYPTNP VTGEIPTLS ALDIPDGDEV DVQWRLVHDS ANFIKPTSYL AHYLGYAWVG GNHSQYVGED MDVTRDGDGW VIRGNNDGGC D GYRCGDKT ...String:
AEPVYPDQLR LFSLGQGVCG DKYRPVNREE AQSVKSNIVG MMGQWQISGL ANGWVIMGPG YNGEIKPGTA SNTWCYPTNP VTGEIPTLS ALDIPDGDEV DVQWRLVHDS ANFIKPTSYL AHYLGYAWVG GNHSQYVGED MDVTRDGDGW VIRGNNDGGC D GYRCGDKT AIKVSNFAYN LDPDSFKHGD VTQSDRQLVK TVVGWAVNDS DTPQSGYDVT LRYDTATNWS KTNTYGLSEK VT TKNKFCW PLVGETELSI CIAANQSWAS QNGGSTTTSL SQSVRPTVPA RSKIPVKIEL YKADISYPYE FKADVSYDLT LSG FLRWGG NAWYTHPDNR PNWNHTFVIG PYKDKASSIR YQWDKRYIPG EVKWWDWNWT IQQNGLSTMQ NNLARVLRPV RAGI TGDFS AESQFAGNIE IGAPVPLAA

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.4 mg/mL
BufferpH: 7.4
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 70 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV
Details: 30 seconds glow discharged grids, 4ul sample, 5 seconds blot.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Average exposure time: 1.0 sec. / Average electron dose: 22.42 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 260958
CTF correctionSoftware - Name: RELION (ver. 1.3)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.46 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.3) / Number images used: 70766
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 1.3)
Final angle assignmentType: ANGULAR RECONSTITUTION
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

3c0n


Chain - Chain ID: B
RefinementProtocol: FLEXIBLE FIT
Output model

PDB-5jzw:
Cryo-EM structures of aerolysin post-prepore and quasipore

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