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- EMDB-50562: Aerolysin heptamer in membrane inserted form reconstituted in amp... -

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Basic information

Entry
Database: EMDB / ID: EMD-50562
TitleAerolysin heptamer in membrane inserted form reconstituted in amphipoles.
Map datapostprocessed map of aerolysin heptamer in A8-35.
Sample
  • Complex: aerolysin heptamer in A8-35
    • Protein or peptide: Aerolysin
Keywordspore forming toxin / aerolysin / amphipole reconstitution / cryo-EM / TOXIN
Function / homology
Function and homology information


symbiont-mediated cytolysis of host cell / 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: 2.2 Å
AuthorsIacovache I / Zuber B
Funding support Switzerland, 3 items
OrganizationGrant numberCountry
Swiss National Science Foundation31003A_179520 Switzerland
Swiss National Science Foundation32NE30_185536 Switzerland
Swiss National Science Foundation32NE30_185536 Switzerland
CitationJournal: J Am Chem Soc / Year: 2025
Title: Aerolysin Nanopore Structures Revealed at High Resolution in a Lipid Environment.
Authors: Jana S Anton / Ioan Iacovache / Juan F Bada Juarez / Luciano A Abriata / Louis W Perrin / Chan Cao / Maria J Marcaida / Benoît Zuber / Matteo Dal Peraro /
Abstract: Aerolysin is a β-pore-forming toxin produced by most Aeromonas bacteria, which has attracted large attention in the field of nanopore sensing due to its narrow and charged pore lumen. Structurally ...Aerolysin is a β-pore-forming toxin produced by most Aeromonas bacteria, which has attracted large attention in the field of nanopore sensing due to its narrow and charged pore lumen. Structurally similar proteins, belonging to the aerolysin-like family, are present throughout all kingdoms of life, but very few of them have been structurally characterized in a lipid environment. Here, we present the first high-resolution atomic cryo-EM structures of aerolysin prepore and pore in a membrane-like environment. These structures allow the identification of key interactions, which are relevant for understanding the pore formation mechanism and for correctly positioning the pore β-barrel and its anchoring β-turn motif in the membrane. Moreover, we elucidate at high resolution the architecture of key pore mutations and precisely identify four constriction rings in the pore lumen that are highly relevant for nanopore sensing experiments.
History
DepositionJun 6, 2024-
Header (metadata) releaseFeb 12, 2025-
Map releaseFeb 12, 2025-
UpdateFeb 19, 2025-
Current statusFeb 19, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_50562.png

Downloads & links

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Map

FileDownload / File: emd_50562.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationpostprocessed map of aerolysin heptamer in A8-35.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.94 Å/pix.
x 240 pix.
= 225.6 Å		0.94 Å/pix.
x 240 pix.
= 225.6 Å		0.94 Å/pix.
x 240 pix.
= 225.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.94 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.015
Minimum - Maximum-0.10551148 - 0.18214285
Average (Standard dev.)0.00022193223 (±0.005799765)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 225.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_50562_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: refinement map

Fileemd_50562_additional_1.map
Annotationrefinement map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 2

Fileemd_50562_half_map_1.map
Annotationhalf map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 1

Fileemd_50562_half_map_2.map
Annotationhalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : aerolysin heptamer in A8-35

EntireName: aerolysin heptamer in A8-35
Components
  • Complex: aerolysin heptamer in A8-35
    • Protein or peptide: Aerolysin

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Supramolecule #1: aerolysin heptamer in A8-35

SupramoleculeName: aerolysin heptamer in A8-35 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Heptamerization was induced by trypsinization of proaerolysin.
Source (natural)Organism: Aeromonas hydrophila (bacteria)
Molecular weightTheoretical: 350 KDa

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Macromolecule #1: Aerolysin

MacromoleculeName: Aerolysin / type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO
Source (natural)Organism: Aeromonas hydrophila (bacteria)
Molecular weightTheoretical: 51.980453 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
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 TKNKFKW PLVGETELSI EIAANQSWAS QNGGSTTTSL SQSVRPTVPA RSKIPVKIEL YKADISYPYE FKADVSYDLT LSG FLRWGG NAWYTHPDNR PNWNHTFVIG PYKDKASSIR YQWDKRYIPG EVKWWDWNWT IQQNGLSTMQ NNLARVLRPV RAGI TGDFS AESQFAGNIE IGAPVPLAAD SKVRRARSVD GAGQGLRLEI PLDAQELSGL GFNNVSLSVT PAANQ

UniProtKB: Aerolysin

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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

Concentration1 mg/mL
BufferpH: 7.4 / Details: 20mM Hepes 7.4 100mM NaCl 0.05% A8-35
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 5 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 2e-05 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.2 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: EMDB MAP
EMDB ID:

8187

Final reconstructionApplied symmetry - Point group: C7 (7 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 5) / Number images used: 1070455
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 5)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: Other / Chain - Initial model type: other / Details: modelangelo
DetailsModelAngelo was used with the final map to generate the first model.
RefinementProtocol: OTHER
Output model

PDB-9fml:
Aerolysin heptamer in membrane inserted form reconstituted in amphipoles.

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