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- PDB-9f25: Cryo-EM structure of Botulinum neurotoxin A LC-HN domain -

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Basic information

Entry
Database: PDB / ID: 9f25
TitleCryo-EM structure of Botulinum neurotoxin A LC-HN domain
ComponentsBotulinum neurotoxin type A
KeywordsTOXIN / Neurotoxin / synaptic vesicle glycoprotein / Clostridium botulinum
Function / homology
Function and homology information


symbiont-mediated suppression of host exocytosis / ganglioside GT1b binding / Toxicity of botulinum toxin type A (botA) / bontoxilysin / host cell presynaptic membrane / host cell cytoplasmic vesicle / host cell cytosol / protein transmembrane transporter activity / metalloendopeptidase activity / toxin activity ...symbiont-mediated suppression of host exocytosis / ganglioside GT1b binding / Toxicity of botulinum toxin type A (botA) / bontoxilysin / host cell presynaptic membrane / host cell cytoplasmic vesicle / host cell cytosol / protein transmembrane transporter activity / metalloendopeptidase activity / toxin activity / host cell plasma membrane / proteolysis / extracellular region / zinc ion binding / membrane
Similarity search - Function
Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain / Clostridium neurotoxin, translocation domain / Clostridium neurotoxin, receptor-binding C-terminal / Clostridium neurotoxin, C-terminal receptor binding / Clostridial neurotoxin zinc protease / Botulinum/Tetanus toxin, catalytic chain / Clostridium neurotoxin, receptor binding N-terminal / Clostridium neurotoxin, N-terminal receptor binding / Kunitz inhibitor STI-like superfamily ...Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain / Clostridium neurotoxin, translocation domain / Clostridium neurotoxin, receptor-binding C-terminal / Clostridium neurotoxin, C-terminal receptor binding / Clostridial neurotoxin zinc protease / Botulinum/Tetanus toxin, catalytic chain / Clostridium neurotoxin, receptor binding N-terminal / Clostridium neurotoxin, N-terminal receptor binding / Kunitz inhibitor STI-like superfamily / Neutral zinc metallopeptidases, zinc-binding region signature. / Concanavalin A-like lectin/glucanase domain superfamily
Similarity search - Domain/homology
Botulinum neurotoxin type A
Similarity search - Component
Biological speciesClostridium botulinum (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsKhanppnavar, B. / Leka, O. / Korkhov, V. / Kammerer, R.
Funding support Switzerland, 2items
OrganizationGrant numberCountry
Swiss National Science Foundation310030_212253 Switzerland
Swiss National Science Foundation198545 Switzerland
CitationJournal: Nat Commun / Year: 2025
Title: Cryo-EM structure of the botulinum neurotoxin A/SV2B complex and its implications for translocation.
Authors: Basavraj Khanppnavar / Oneda Leka / Sushant K Pal / Volodymyr M Korkhov / Richard A Kammerer /
Abstract: Botulinum neurotoxin A1 (BoNT/A1) belongs to the most potent toxins and is used as a major therapeutic agent. Neurotoxin conformation is crucial for its translocation to the neuronal cytosol, a key ...Botulinum neurotoxin A1 (BoNT/A1) belongs to the most potent toxins and is used as a major therapeutic agent. Neurotoxin conformation is crucial for its translocation to the neuronal cytosol, a key process for intoxication that is only poorly understood. To gain molecular insights into the steps preceding toxin translocation, we determine cryo-EM structures of BoNT/A1 alone and in complex with its receptor synaptic vesicle glycoprotein 2B (SV2B). In solution, BoNT/A1 adopts a unique, semi-closed conformation. The toxin changes its structure into an open state upon receptor binding with the translocation domain (H) and the catalytic domain (LC) remote from the membrane, suggesting translocation incompatibility. Under acidic pH conditions, where translocation is initiated, receptor-bound BoNT/A1 switches back into a semi-closed conformation. This conformation brings the LC and H close to the membrane, suggesting that a translocation-competent state of the toxin is required for successful LC transport into the neuronal cytosol.
History
DepositionApr 22, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 12, 2025Provider: repository / Type: Initial release
Revision 1.0Mar 12, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Mar 12, 2025Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
Revision 1.0Mar 12, 2025Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Mar 12, 2025Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Mar 12, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Mar 12, 2025Data content type: Mask / Part number: 1 / Data content type: Mask / Provider: repository / Type: Initial release
Revision 1.0Mar 12, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release

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Structure visualization

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Assembly

Deposited unit
A: Botulinum neurotoxin type A


Theoretical massNumber of molelcules
Total (without water)153,0841
Polymers153,0841
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein Botulinum neurotoxin type A / BoNT/A / Bontoxilysin-A / BOTOX / Botulinum neurotoxin type A1


Mass: 153083.734 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Clostridium botulinum (bacteria) / Gene: botA, atx, bonT / Production host: Escherichia coli (E. coli) / References: UniProt: P0DPI0
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Botulinum neurotoxin A / Type: COMPLEX / Details: Botulinum neurotoxin A / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.15 MDa / Experimental value: NO
Source (natural)Organism: Clostridium botulinum (bacteria)
Source (recombinant)Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2400 nm / Nominal defocus min: 600 nm
Image recordingElectron dose: 80 e/Å2 / Film or detector model: GATAN K3 BIOCONTINUUM (6k x 4k)

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Processing

EM softwareName: PHENIX / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 121176 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT
Atomic model buildingPDB-ID: 6UL6

6ul6


Accession code: 6UL6 / Source name: PDB / Type: experimental model
RefinementHighest resolution: 3.7 Å / Cross valid method: NONE
Stereochemistry target values: REAL-SPACE (WEIGHTED MAP SUM AT ATOM CENTERS)
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0026862
ELECTRON MICROSCOPYf_angle_d0.5289294
ELECTRON MICROSCOPYf_dihedral_angle_d16.7692524
ELECTRON MICROSCOPYf_chiral_restr0.041036
ELECTRON MICROSCOPYf_plane_restr0.0041194

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