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- EMDB-36934: Local map of SV2A LD4-BoNT/A2 Hc from SV2A-BoNT/A2 Hc-brivaraceta... -

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Basic information

Entry
Database: EMDB / ID: EMD-36934
TitleLocal map of SV2A LD4-BoNT/A2 Hc from SV2A-BoNT/A2 Hc-brivaracetam complex
Map data
Sample
  • Complex: Synaptic vesicle glycroprotein 2A in complex with brivaracetam and botulinum neurotoxin A2 Hc domain
    • Complex: Synaptic vesicle glycoprotein
      • Protein or peptide: Synaptic vesicle glycoprotein
    • Complex: Botulinum neurotoxin A2
      • Protein or peptide: Botulinum neurotoxin A2
KeywordsSynapse / epilepsy / antiepileptic drug / botulinum neurotoxin / MEMBRANE PROTEIN
Biological speciesHomo sapiens (human) / Clostridium botulinum (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.07 Å
AuthorsYamagata A
Funding support Japan, 1 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)JP22H02564 Japan
CitationJournal: 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
DepositionJul 26, 2023-
Header (metadata) releaseMay 15, 2024-
Map releaseMay 15, 2024-
UpdateMay 15, 2024-
Current statusMay 15, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_36934.png

Downloads & links

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Map

FileDownload / File: emd_36934.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.83 Å
Density
Contour LevelBy AUTHOR: 0.237
Minimum - Maximum-4.137098 - 6.4636617
Average (Standard dev.)-0.0004501057 (±0.029882763)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 249.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_36934_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
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Histogram

Histogram (log scale)

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Half map: #1

Fileemd_36934_half_map_1.map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Half map: #2

Fileemd_36934_half_map_2.map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Sample components

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Entire : Synaptic vesicle glycroprotein 2A in complex with brivaracetam an...

EntireName: Synaptic vesicle glycroprotein 2A in complex with brivaracetam and botulinum neurotoxin A2 Hc domain
Components
  • Complex: Synaptic vesicle glycroprotein 2A in complex with brivaracetam and botulinum neurotoxin A2 Hc domain
    • Complex: Synaptic vesicle glycoprotein
      • Protein or peptide: Synaptic vesicle glycoprotein
    • Complex: Botulinum neurotoxin A2
      • Protein or peptide: Botulinum neurotoxin A2

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Supramolecule #1: Synaptic vesicle glycroprotein 2A in complex with brivaracetam an...

SupramoleculeName: Synaptic vesicle glycroprotein 2A in complex with brivaracetam and botulinum neurotoxin A2 Hc domain
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #2: Synaptic vesicle glycoprotein

SupramoleculeName: Synaptic vesicle glycoprotein / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Clostridium botulinum (bacteria)

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Supramolecule #3: Botulinum neurotoxin A2

SupramoleculeName: Botulinum neurotoxin A2 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2

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Macromolecule #1: Synaptic vesicle glycoprotein

MacromoleculeName: Synaptic vesicle glycoprotein / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MDYKDDDDKE EGFRDRAAFI RGAKDIAKEV KKHAAKKVVK GLDRVQDEYS RRSYSRFEEE DDDDDFPAP SDGYYRGEGT QDEEEGGASS DATEGHDEDD EIYEGEYQGI PRAESGGKGE R MADGAPLA GVRGGLSDGE GPPGGRGEAQ RRKEREELAQ QYEAILRECG ...String:
MDYKDDDDKE EGFRDRAAFI RGAKDIAKEV KKHAAKKVVK GLDRVQDEYS RRSYSRFEEE DDDDDFPAP SDGYYRGEGT QDEEEGGASS DATEGHDEDD EIYEGEYQGI PRAESGGKGE R MADGAPLA GVRGGLSDGE GPPGGRGEAQ RRKEREELAQ QYEAILRECG HGRFQWTLYF VL GLALMAD GVEVFVVGFV LPSAEKDMCL SDSNKGMLGL IVYLGMMVGA FLWGGLADRL GRR QCLLIS LSVNSVFAFF SSFVQGYGTF LFCRLLSGVG IGGSIPIVFS YFSEFLAQEK RGEH LSWLC MFWMIGGVYA AAMAWAIIPH YGWSFQMGSA YQFHSWRVFV LVCAFPSVFA IGALT TQPE SPRFFLENGK HDEAWMVLKQ VHDTNMRAKG HPERVFSVTH IKTIHQEDEL IEIQSD TGT WYQRWGVRAL SLGGQVWGNF LSCFGPEYRR ITLMMMGVWF TMSFSYYGLT VWFPDMI RH LQAVDYASRT KVFPGERVEH VTFNFTLENQ IHRGGQYFND KFIGLRLKSV SFEDSLFE E CYFEDVTSSN TFFRNCTFIN TVFYNTDLFE YKFVNSRLIN STFLHNKEGC PLDVTGTGE GAYMVYFVSF LGTLAVLPGN IVSALLMDKI GRLRMLAGSS VMSCVSCFFL SFGNSESAMI ALLCLFGGV SIASWNALDV LTVELYPSDK RTTAFGFLNA LCKLAAVLGI SIFTSFVGIT K AAPILFAS AALALGSSLA LKLPETRGQV LQ

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Macromolecule #2: Botulinum neurotoxin A2

MacromoleculeName: Botulinum neurotoxin A2 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Clostridium botulinum (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: KNIVNTSILS IVYKKDDLID LSRYGAKINI GDRVYYDSID KNQIKLINLE SSTIEVILKN AIVYNSMYE NFSTSFWIKI PKYFSKINLN NEYTIINCIE NNSGWKVSLN YGEIIWTLQD N KQNIQRVV FKYSQMVNIS DYINRWIFVT ITNNRLTKSK IYINGRLIDQ ...String:
KNIVNTSILS IVYKKDDLID LSRYGAKINI GDRVYYDSID KNQIKLINLE SSTIEVILKN AIVYNSMYE NFSTSFWIKI PKYFSKINLN NEYTIINCIE NNSGWKVSLN YGEIIWTLQD N KQNIQRVV FKYSQMVNIS DYINRWIFVT ITNNRLTKSK IYINGRLIDQ KPISNLGNIH AS NKIMFKL DGCRDPRRYI MIKYFNLFDK ELNEKEIKDL YDSQSNSGIL KDFWGNYLQY DKP YYMLNL FDPNKYVDVN NIGIRGYMYL KGPRGSVVTT NIYLNSTLYE GTKFIIKKYA SGNE DNIVR NNDRVYINVV VKNKEYRLAT NASQAGVEKI LSALEIPDVG NLSQVVVMKS KDDQG IRNK CKMNLQDNNG NDIGFIGFHL YDNIAKLVAS NWYNRQVGKA SRTFGCSWEF IPVDDG WGE SSL

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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

BufferpH: 7.5
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 53.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.6 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 4734449
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

8js8

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.07 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 479130
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
FSC plot (resolution estimation)

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