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- EMDB-4118: CryoEM structure of the membrane pore complex of Pneumolysin at 4.5A -

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Basic information

Entry
Database: EMDB / ID: 4118
TitleCryoEM structure of the membrane pore complex of Pneumolysin at 4.5A
Map dataPLY pore complex
SamplePneumolysin pore complex
  • Pneumolysin
Function/homologyThiol-activated cytolysins signature. / Thiol-activated cytolysin / Thiol-activated cytolysin C-terminal / Thiol-activated cytolysin superfamily / Thiol-activated cytolysin, alpha-beta domain superfamily / Thiol-activated cytolysin / Thiol-activated cytolysin beta sandwich domain / cholesterol binding / hemolysis in other organism / toxin activity ...Thiol-activated cytolysins signature. / Thiol-activated cytolysin / Thiol-activated cytolysin C-terminal / Thiol-activated cytolysin superfamily / Thiol-activated cytolysin, alpha-beta domain superfamily / Thiol-activated cytolysin / Thiol-activated cytolysin beta sandwich domain / cholesterol binding / hemolysis in other organism / toxin activity / host cell plasma membrane / integral component of membrane / extracellular region / Pneumolysin
Function and homology information
SourceStreptococcus pneumoniae / / bacteria
MethodCryo EM / single particle reconstruction / 4.5 Å resolution
Authorsvan Pee K / Neuhaus A
CitationJournal: Elife / Year: 2017
Title: CryoEM structures of membrane pore and prepore complex reveal cytolytic mechanism of Pneumolysin.
Authors: Katharina van Pee / Alexander Neuhaus / Edoardo D'Imprima / Deryck J Mills / Werner Kühlbrandt / Özkan Yildiz
Abstract: Many pathogenic bacteria produce pore-forming toxins to attack and kill human cells. We have determined the 4.5 Å structure of the ~2.2 MDa pore complex of pneumolysin, the main virulence factor of ...Many pathogenic bacteria produce pore-forming toxins to attack and kill human cells. We have determined the 4.5 Å structure of the ~2.2 MDa pore complex of pneumolysin, the main virulence factor of , by cryoEM. The pneumolysin pore is a 400 Å ring of 42 membrane-inserted monomers. Domain 3 of the soluble toxin refolds into two ~85 Å β-hairpins that traverse the lipid bilayer and assemble into a 168-strand β-barrel. The pore complex is stabilized by salt bridges between β-hairpins of adjacent subunits and an internal α-barrel. The apolar outer barrel surface with large sidechains is immersed in the lipid bilayer, while the inner barrel surface is highly charged. Comparison of the cryoEM pore complex to the prepore structure obtained by electron cryo-tomography and the x-ray structure of the soluble form reveals the detailed mechanisms by which the toxin monomers insert into the lipid bilayer to perforate the target membrane.
Validation ReportPDB-ID: 5ly6

SummaryFull reportAbout validation report
DateDeposition: Sep 24, 2016 / Header (metadata) release: Oct 26, 2016 / Map release: Apr 5, 2017 / Last update: Aug 2, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.02
  • Imaged by UCSF CHIMERA
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  • Surface view colored by cylindrical radius
  • Surface level: 0.02
  • Imaged by UCSF CHIMERA
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  • Surface view with fitted model
  • Atomic models: : PDB-5ly6
  • Surface level: 0.022
  • Imaged by UCSF CHIMERA
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  • Simplified surface model + fitted atomic model
  • Atomic models: PDB-5ly6
  • Imaged by Jmol
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3D viewer
Supplemental images

Downloads & links

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Map

Fileemd_4118.map.gz (map file in CCP4 format, 186625 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
360 pix
1.4 Å/pix.
= 504. Å
360 pix
1.4 Å/pix.
= 504. Å
360 pix
1.4 Å/pix.
= 504. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Voxel sizeX=Y=Z: 1.4 Å
Density
Contour Level:0.0282 (by author), 0.02 (movie #1):
Minimum - Maximum-0.022703713 - 0.079944775
Average (Standard dev.)0.0008561607 (0.0045818426)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions360360360
Origin000
Limit359359359
Spacing360360360
CellA=B=C: 504 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.41.41.4
M x/y/z360360360
origin x/y/z0.0000.0000.000
length x/y/z504.000504.000504.000
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS360360360
D min/max/mean-0.0230.0800.001

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Supplemental data

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Sample components

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Entire Pneumolysin pore complex

EntireName: Pneumolysin pore complex / Number of components: 2
MassTheoretical: 2.2 MDa

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Component #1: cellular-component, Pneumolysin pore complex

Cellular-componentName: Pneumolysin pore complex / Recombinant expression: No
MassTheoretical: 2.2 MDa
SourceSpecies: Streptococcus pneumoniae / / bacteria
Source (engineered)Expression System: Escherichia coli bl21 / / bacteria / image: Escherichia coli
Vector: pET15

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Component #2: protein, Pneumolysin

ProteinName: Pneumolysin / Recombinant expression: No
MassTheoretical: 52.866066 kDa
Source (engineered)Expression System: Streptococcus pneumoniae serotype 2 (strain d39 / nctc 7466) / / bacteria

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: Cryo EM
Sample solutionSpecimen conc.: 1 mg/ml / pH: 7
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company
ImagingMicroscope: FEI POLARA 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 1.02 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C42 (42 fold cyclic) / Number of projections: 6461
3D reconstructionSoftware: RELION / Resolution: 4.5 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution assessment)

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Atomic model buiding

Modeling #1Refinement protocol: flexible
Input PDB model: 5AOD
Chain ID: 5AOD_A
Output model

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