|Entry||Database: PDB / ID: 5ly6|
|Title||CryoEM structure of the membrane pore complex of Pneumolysin at 4.5A|
|Keywords||TOXIN / Bacterial toxins / pore complex / cryoEM structure / cholesterol-dependent cytolysin / Pneumolysin / membrane pore|
|Function/homology||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 ...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
|Specimen source||Streptococcus pneumoniae serotype 2 / / bacteria|
|Method||Electron microscopy (4.5 Å resolution / Particle / Single particle) / Transmission electron microscopy|
|Authors||van Pee, K. / Neuhaus, A. / D'Imprima, E. / Mills, D.J. / Kuehlbrandt, W. / Yildiz, O.|
|Citation||Journal: 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.
SummaryFull reportAbout validation report
|Date||Deposition: Sep 24, 2016 / Release: Apr 5, 2017|
Downloads & links
|#1: Protein/peptide|| |
Mass: 52866.066 Da / Num. of mol.: 1
Source: (gene. exp.) Streptococcus pneumoniae serotype 2 (strain d39 / nctc 7466) / / bacteria
Gene: ply, SPD_1726 / Production host: Escherichia coli BL21(DE3) / References: UniProt:Q04IN8
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / Reconstruction method: SINGLE PARTICLE|
|Component||Name: Pneumolysin pore complex / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: 1 / Source: RECOMBINANT|
|Molecular weight||Value: 2.2 deg. / Units: MEGADALTONS|
|Source (natural)||Organism: Streptococcus pneumoniae|
|Source (recombinant)||Organism: Escherichia coli BL21 / Plasmid: pET15|
|Buffer solution||pH: 7|
|Specimen||Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Vitrification||Cryogen name: ETHANE|
-Electron microscopy imaging
Model: Tecnai Polara / Image courtesy: FEI Company
|Microscopy||Microscope model: FEI POLARA 300|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELD|
|Image recording||Average exposure time: 6 sec. / Electron dose: 1.02 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)|
|CTF correction||Type: PHASE FLIPPING AND AMPLITUDE CORRECTION|
|Symmetry||Point symmetry: C42|
|3D reconstruction||Resolution: 4.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 6461 / Symmetry type: POINT|
|Atomic model building||Ref protocol: AB INITIO MODEL|
|Atomic model building||PDB-ID: 5AOD|
Pdb chain ID: A / Pdb chain residue range: 1-471
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