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Yorodumi- PDB-8jlh: Cryo-EM structure of SV2A dimer in complex with BoNT/A2 Hc and le... -
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-Basic information
Entry | Database: PDB / ID: 8jlh | ||||||
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Title | Cryo-EM structure of SV2A dimer in complex with BoNT/A2 Hc and levetiracetam | ||||||
Components |
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Keywords | TRANSPORT PROTEIN / Synaptic vesicle / epilepsy | ||||||
Function / homology | Function and homology information regulation of gamma-aminobutyric acid secretion / Toxicity of botulinum toxin type F (botF) / Toxicity of botulinum toxin type D (botD) / Toxicity of botulinum toxin type E (botE) / Toxicity of botulinum toxin type A (botA) / synaptic vesicle priming / presynaptic active zone / transmembrane transporter activity / protein transmembrane transporter activity / GABA-ergic synapse ...regulation of gamma-aminobutyric acid secretion / Toxicity of botulinum toxin type F (botF) / Toxicity of botulinum toxin type D (botD) / Toxicity of botulinum toxin type E (botE) / Toxicity of botulinum toxin type A (botA) / synaptic vesicle priming / presynaptic active zone / transmembrane transporter activity / protein transmembrane transporter activity / GABA-ergic synapse / neuromuscular junction / metalloendopeptidase activity / synaptic vesicle membrane / intracellular calcium ion homeostasis / cell-cell junction / synaptic vesicle / toxin activity / neuron projection / neuronal cell body / glutamatergic synapse / dendrite / protein kinase binding / endoplasmic reticulum / proteolysis / zinc ion binding / extracellular region / plasma membrane Similarity search - Function | ||||||
Biological species | Homo sapiens (human) Clostridium botulinum (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å | ||||||
Authors | Yamagata, A. | ||||||
Funding support | Japan, 1items
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Citation | Journal: Nat Commun / Year: 2024 Title: Structural basis for antiepileptic drugs and botulinum neurotoxin recognition of SV2A. Authors: Atsushi Yamagata / Kaori Ito / Takehiro Suzuki / Naoshi Dohmae / Tohru Terada / Mikako Shirouzu / Abstract: More than one percent of people have epilepsy worldwide. Levetiracetam (LEV) is a successful new-generation antiepileptic drug (AED), and its derivative, brivaracetam (BRV), shows improved efficacy. ...More than one percent of people have epilepsy worldwide. Levetiracetam (LEV) is a successful new-generation antiepileptic drug (AED), and its derivative, brivaracetam (BRV), shows improved efficacy. Synaptic vesicle glycoprotein 2a (SV2A), a putative membrane transporter in the synaptic vesicles (SVs), has been identified as a target of LEV and BRV. SV2A also serves as a receptor for botulinum neurotoxin (BoNT), which is the most toxic protein and has paradoxically emerged as a potent reagent for therapeutic and cosmetic applications. Nevertheless, no structural analysis on AEDs and BoNT recognition by full-length SV2A has been available. Here we describe the cryo-electron microscopy structures of the full-length SV2A in complex with the BoNT receptor-binding domain, BoNT/A2 H and either LEV or BRV. The large fourth luminal domain of SV2A binds to BoNT/A2 H through protein-protein and protein-glycan interactions. LEV and BRV occupy the putative substrate-binding site in an outward-open conformation. A propyl group in BRV creates additional contacts with SV2A, explaining its higher binding affinity than that of LEV, which was further supported by label-free spectral shift assay. Numerous LEV derivatives have been developed as AEDs and positron emission tomography (PET) tracers for neuroimaging. Our work provides a structural framework for AEDs and BoNT recognition of SV2A and a blueprint for the rational design of additional AEDs and PET tracers. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8jlh.cif.gz | 344 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8jlh.ent.gz | 265.2 KB | Display | PDB format |
PDBx/mmJSON format | 8jlh.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8jlh_validation.pdf.gz | 1.5 MB | Display | wwPDB validaton report |
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Full document | 8jlh_full_validation.pdf.gz | 1.6 MB | Display | |
Data in XML | 8jlh_validation.xml.gz | 61 KB | Display | |
Data in CIF | 8jlh_validation.cif.gz | 89.1 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/jl/8jlh ftp://data.pdbj.org/pub/pdb/validation_reports/jl/8jlh | HTTPS FTP |
-Related structure data
Related structure data | 36397MC 8jlcC 8jleC 8jlfC 8jlgC 8jliC 8js8C 8js9C 8k77C M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 83774.008 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: SV2A / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q7L0J3 #2: Protein | Mass: 49456.191 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Clostridium botulinum (bacteria) / Production host: Escherichia coli (E. coli) / References: UniProt: D2KCK3 #3: Polysaccharide | Source method: isolated from a genetically manipulated source #4: Chemical | ChemComp-UKX / ( | Mass: 170.209 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C8H14N2O2 / Feature type: SUBJECT OF INVESTIGATION #5: Sugar | ChemComp-NAG / Has ligand of interest | Y | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: SV2A dimer in complex with BoNT/A2 Hc domain and anti-epileptic drug, levetiracetam Type: COMPLEX / Entity ID: #2 / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Clostridium botulinum (bacteria) |
Buffer solution | pH: 7.5 Details: 20 mM Hepes (pH7.5), 150 mM NaCl, 0.001% LMNG, 0.0002% cholesterol hemisuccinate |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 297 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm |
Image recording | Electron dose: 50.8 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 6518 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 6409122 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 425512 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement | Resolution: 2.9→159.36 Å / Cor.coef. Fo:Fc: 0.856 / WRfactor Rwork: 0.374 / SU B: 10.939 / SU ML: 0.199 / Average fsc free: 0 / Average fsc overall: 0.8139 / Average fsc work: 0.8139 / Cross valid method: NONE / ESU R: 0.306 Details: Hydrogens have been added in their riding positions
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Solvent computation | Solvent model: NONE | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 158.273 Å2 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement step | Cycle: 1 / Total: 12610 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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LS refinement shell | Refine-ID: ELECTRON MICROSCOPY / Num. reflection Rfree: _ / Total num. of bins used: 20 / % reflection obs: 100 %
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