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- EMDB-50891: Cryo-EM structure of native SV2A in complex with TeNT-Hc, Pro-Mac... -

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Entry
Database: EMDB / ID: EMD-50891
TitleCryo-EM structure of native SV2A in complex with TeNT-Hc, Pro-Macrobody 5 and Levetiracetam
Map dataCryo-EM structure of native SV2A in complex with TeNT-Hc, Pro-macrobody 5 and Levetiracetam
Sample
  • Complex: SV2A bound to TeNT-Hc, Pro-Macrobody 5 and Levetiracetam
    • Complex: Synaptic vesicle glycoprotein 2A
      • Protein or peptide: Synaptic vesicle glycoprotein 2A
    • Complex: TeNT-Hc
      • Protein or peptide: Tetanus toxin heavy chain
    • Complex: Pro-Macrobody 5
      • Protein or peptide: Pro-Macrobody 5,Maltose/maltodextrin-binding periplasmic protein,Maltose/maltodextrin-binding periplasmic protein
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: omega-undecylenyl-beta-D-maltopyranoside
  • Ligand: (2S)-2-(2-oxidanylidenepyrrolidin-1-yl)butanamide
Keywordssynaptic vesicles / MFS transporter / MEMBRANE PROTEIN / epilepsy / Levetiracetam / clostridial neurotoxin
Function / homology
Function and homology information


tentoxilysin / symbiont-mediated perturbation of host neurotransmitter secretion / Toxicity of tetanus toxin (tetX) / synaptic vesicle priming / detection of maltose stimulus / maltose transport complex / carbohydrate transport / presynaptic active zone / transmembrane transporter activity / protein transmembrane transporter activity ...tentoxilysin / symbiont-mediated perturbation of host neurotransmitter secretion / Toxicity of tetanus toxin (tetX) / synaptic vesicle priming / detection of maltose stimulus / maltose transport complex / carbohydrate transport / presynaptic active zone / transmembrane transporter activity / protein transmembrane transporter activity / carbohydrate transmembrane transporter activity / maltose binding / maltose transport / maltodextrin transmembrane transport / ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing / ATP-binding cassette (ABC) transporter complex / GABA-ergic synapse / cell chemotaxis / clathrin-coated endocytic vesicle membrane / neuromuscular junction / metalloendopeptidase activity / intracellular calcium ion homeostasis / synaptic vesicle membrane / endocytic vesicle membrane / cell-cell junction / outer membrane-bounded periplasmic space / toxin activity / periplasmic space / neuron projection / DNA damage response / protein kinase binding / glutamatergic synapse / endoplasmic reticulum / proteolysis / extracellular region / zinc ion binding / membrane / plasma membrane / cytosol
Similarity search - Function
: / SV2A/B/C luminal domain / Synaptic vesicle protein SV2 / Sugar transport proteins signature 2. / Sugar transporter, conserved site / Major facilitator, sugar transporter-like / Sugar (and other) transporter / Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain / Clostridium neurotoxin, translocation domain ...: / SV2A/B/C luminal domain / Synaptic vesicle protein SV2 / Sugar transport proteins signature 2. / Sugar transporter, conserved site / Major facilitator, sugar transporter-like / Sugar (and other) transporter / Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain / Clostridium neurotoxin, translocation domain / Clostridial neurotoxin zinc protease / Botulinum/Tetanus toxin, catalytic chain / Clostridium neurotoxin, receptor-binding C-terminal / Clostridium neurotoxin, receptor binding N-terminal / Clostridium neurotoxin, C-terminal receptor binding / Clostridium neurotoxin, N-terminal receptor binding / Kunitz inhibitor STI-like superfamily / Major facilitator superfamily / Major Facilitator Superfamily / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / Maltose/Cyclodextrin ABC transporter, substrate-binding protein / Solute-binding family 1, conserved site / Bacterial extracellular solute-binding proteins, family 1 signature. / MFS transporter superfamily / Bacterial extracellular solute-binding protein / Bacterial extracellular solute-binding protein / Neutral zinc metallopeptidases, zinc-binding region signature. / Concanavalin A-like lectin/glucanase domain superfamily
Similarity search - Domain/homology
Synaptic vesicle glycoprotein 2A / Tetanus toxin / Maltose/maltodextrin-binding periplasmic protein
Similarity search - Component
Biological speciesOvis aries (sheep) / Clostridium tetani E88 (bacteria) / Lama glama (llama) / Escherichia coli K-12 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsSchenck S / Brunner JD
Funding support Belgium, 1 items
OrganizationGrant numberCountry
Vrije Universiteit Brussel Belgium
CitationJournal: Nat Commun / Year: 2025
Title: Structures of native SV2A reveal the binding mode for tetanus neurotoxin and anti-epileptic racetams.
Authors: Stephan Schenck / Toon Laeremans / Jan Steyaert / Janine D Brunner /
Abstract: The synaptic vesicle glycoprotein 2A (SV2A) is a synaptic vesicle (SV) resident with homology to the major facilitator superfamily (MFS) and essential in vertebrate neurotransmission. Despite its ...The synaptic vesicle glycoprotein 2A (SV2A) is a synaptic vesicle (SV) resident with homology to the major facilitator superfamily (MFS) and essential in vertebrate neurotransmission. Despite its unclear physiological role, SV2A is of high medical relevance as it is the target of the anti-epileptic drug Levetiracetam (LEV) and a receptor for clostridial neurotoxins (CNTs), among them presumably tetanus neurotoxin (TeNT). To obtain detailed insights about these molecular interactions we subjected native SV2A, purified from brain tissue, to cryo-EM. We discover that TeNT binds SV2A strikingly different from botulinum neurotoxin A and unveil the precise geometry of TeNT binding to dipartite SV2-ganglioside receptors. The structures deliver compelling support for SV2A as the protein receptor for TeNT in central neurons and reinforce the concepts of the dual receptor hypothesis for CNT entry into neurons. Further, our LEV-bound structure of SV2A reveals the drug-interacting residues, delineates a putative substrate pocket in SV2A and provides insights into the SV2-isoform-specificity of LEV. Our work has implications for CNT engineering from a hitherto unrecognized SV2 binding interface and for improved designs of anti-convulsant drugs in epilepsy treatment.
History
DepositionJul 3, 2024-
Header (metadata) releaseMay 21, 2025-
Map releaseMay 21, 2025-
UpdateMay 21, 2025-
Current statusMay 21, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_50891.map.gz / Format: CCP4 / Size: 343 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM structure of native SV2A in complex with TeNT-Hc, Pro-macrobody 5 and Levetiracetam
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.71 Å/pix.
x 448 pix.
= 318.08 Å
0.71 Å/pix.
x 448 pix.
= 318.08 Å
0.71 Å/pix.
x 448 pix.
= 318.08 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.71 Å
Density
Contour LevelBy AUTHOR: 0.0547
Minimum - Maximum-0.13135806 - 0.2808858
Average (Standard dev.)0.00043160154 (±0.0068635317)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions448448448
Spacing448448448
CellA=B=C: 318.08 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_50891_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Phenix-density-modified map of 'Focused refienment on Levetiracetam'

Fileemd_50891_additional_1.map
AnnotationPhenix-density-modified map of 'Focused refienment on Levetiracetam'
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Composite map

Fileemd_50891_additional_2.map
AnnotationComposite map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Focused refinement on TeNT-Gangliosides

Fileemd_50891_additional_3.map
AnnotationFocused refinement on TeNT-Gangliosides
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: half map A

Fileemd_50891_half_map_1.map
Annotationhalf map A
Projections & Slices
AxesZYX

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Slices (1/2)
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Half map: half map B

Fileemd_50891_half_map_2.map
Annotationhalf map B
Projections & Slices
AxesZYX

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Sample components

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Entire : SV2A bound to TeNT-Hc, Pro-Macrobody 5 and Levetiracetam

EntireName: SV2A bound to TeNT-Hc, Pro-Macrobody 5 and Levetiracetam
Components
  • Complex: SV2A bound to TeNT-Hc, Pro-Macrobody 5 and Levetiracetam
    • Complex: Synaptic vesicle glycoprotein 2A
      • Protein or peptide: Synaptic vesicle glycoprotein 2A
    • Complex: TeNT-Hc
      • Protein or peptide: Tetanus toxin heavy chain
    • Complex: Pro-Macrobody 5
      • Protein or peptide: Pro-Macrobody 5,Maltose/maltodextrin-binding periplasmic protein,Maltose/maltodextrin-binding periplasmic protein
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: omega-undecylenyl-beta-D-maltopyranoside
  • Ligand: (2S)-2-(2-oxidanylidenepyrrolidin-1-yl)butanamide

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Supramolecule #1: SV2A bound to TeNT-Hc, Pro-Macrobody 5 and Levetiracetam

SupramoleculeName: SV2A bound to TeNT-Hc, Pro-Macrobody 5 and Levetiracetam
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Ovis aries (sheep)
Molecular weightTheoretical: 53.6 KDa

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Supramolecule #2: Synaptic vesicle glycoprotein 2A

SupramoleculeName: Synaptic vesicle glycoprotein 2A / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Ovis aries (sheep)

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Supramolecule #3: TeNT-Hc

SupramoleculeName: TeNT-Hc / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Clostridium tetani E88 (bacteria)

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Supramolecule #4: Pro-Macrobody 5

SupramoleculeName: Pro-Macrobody 5 / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #3
Source (natural)Organism: Lama glama (llama)

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Macromolecule #1: Synaptic vesicle glycoprotein 2A

MacromoleculeName: Synaptic vesicle glycoprotein 2A / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Ovis aries (sheep)
Molecular weightTheoretical: 82.640938 KDa
SequenceString: MEEGFRDRAA FIRGAKDIAK EVKKHAAKKV VKGLDRVQDE YSRRSYSRFE EEDDDDDFPA PADGYYRGEG AQDEEEGGAS SDATEGHDE DDEIYEGEYQ GIPRAESGGK GERMADGAPL AGVRGGLGDG EGPPGGRGEA QRRKEREELA QQYEAILREC G HGRFQWTL ...String:
MEEGFRDRAA FIRGAKDIAK EVKKHAAKKV VKGLDRVQDE YSRRSYSRFE EEDDDDDFPA PADGYYRGEG AQDEEEGGAS SDATEGHDE DDEIYEGEYQ GIPRAESGGK GERMADGAPL AGVRGGLGDG EGPPGGRGEA QRRKEREELA QQYEAILREC G HGRFQWTL YFVLGLALMA DGVEVFVVGF VLPSAEKDMC LSDSNKGMLG LIVYLGMMVG AFLWGGLADR LGRRQCLLIS LS VNSVFAF FSSFVQGYGT FLFCRLLSGV GIGGSIPIVF SYFSEFLAQE KRGEHLSWLC MFWMIGGVYA AAMAWAIIPH YGW SFQMGS AYQFHSWRVF VLVCAFPSVF AIGALTTQPE SPRFFLENGK HDEAWMVLKQ VHDTNMRAKG HPERVFSVTH IKTI HQEDE LIEIQSDTGA WYQRWGVRAL SLGGQVWGNF LSCFGPEYRR ITLMMMGVWF TMSFSYYGLT VWFPDMIRHL QAVDY AART KVFPGERVEH VTFNFTLENQ IHRGGQYFND KFIGLRLKSV SFEDSLFEEC YFEDVTSSNT FFRNCTFINT VFYNTD LFE YKFVNSRLVN STFLHNKEGC PLDVTGTGEG AYMVYFVSFL GTLAVLPGNI VSALLMDKIG RLRMLAGSSV MSCVSCF FL SFGNSESAMI ALLCLFGGVS IASWNALDVL TVELYPSDKR TTAFGFLNAL CKLAAVLGIS IFTSFVGITK AAPILFAS A ALALGSSLAL KLPETRGQVL Q

UniProtKB: Synaptic vesicle glycoprotein 2A

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Macromolecule #2: Tetanus toxin heavy chain

MacromoleculeName: Tetanus toxin heavy chain / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Clostridium tetani E88 (bacteria)
Molecular weightTheoretical: 53.718535 KDa
Recombinant expressionOrganism: Escherichia coli K-12 (bacteria)
SequenceString: MGKHHHHHHH HHHGSASLEV LFQGPSHMED IDVILKKSTI LNLDINNDII SDISGFNSSV ITYPDAQLVP GINGKAIHLV NNESSEVIV HKAMDIEYND MFNNFTVSFW LRVPKVSASH LEQYGTNEYS IISSMKKHSL SIGSGWSVSL KGNNLIWTLK D SAGEVRQI ...String:
MGKHHHHHHH HHHGSASLEV LFQGPSHMED IDVILKKSTI LNLDINNDII SDISGFNSSV ITYPDAQLVP GINGKAIHLV NNESSEVIV HKAMDIEYND MFNNFTVSFW LRVPKVSASH LEQYGTNEYS IISSMKKHSL SIGSGWSVSL KGNNLIWTLK D SAGEVRQI TFRDLPDKFN AYLANKWVFI TITNDRLSSA NLYINGVLMG SAEITGLGAI REDNNITLKL DRCNNNNQYV SI DKFRIFC KALNPKEIEK LYTSYLSITF LRDFWGNPLR YDTEYYLIPV ASSSKDVQLK NITDYMYLTN APSYTNGKLN IYY RRLYNG LKFIIKRYTP NNEIDSFVKS GDFIKLYVSY NNNEHIVGYP KDGNAFNNLD RILRVGYNAP GIPLYKKMEA VKLR DLKTY SVQLKLYDDK NASLGLVGTH NGQIGNDPNR DILIASNWYF NHLKDKILGC DWYFVPTDEG WTND

UniProtKB: Tetanus toxin

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Macromolecule #3: Pro-Macrobody 5,Maltose/maltodextrin-binding periplasmic protein,...

MacromoleculeName: Pro-Macrobody 5,Maltose/maltodextrin-binding periplasmic protein,Maltose/maltodextrin-binding periplasmic protein
type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli K-12 (bacteria)
Molecular weightTheoretical: 53.674238 KDa
Recombinant expressionOrganism: Escherichia coli K-12 (bacteria)
SequenceString: GPSQVQLVES GGGLVQAGGS LRLSCAASGR TFSRFIMGWF RQAPGKEREF VAAVGKSGDT TYYADSMSGR FAISRDNAKN TVYLQMISL KPEDTAVYYC AADSSYFYHT HESEYDYWGQ GTQVTVPPLV IWINGDKGYN GLAEVGKKFE KDTGIKVTVE H PDKLEEKF ...String:
GPSQVQLVES GGGLVQAGGS LRLSCAASGR TFSRFIMGWF RQAPGKEREF VAAVGKSGDT TYYADSMSGR FAISRDNAKN TVYLQMISL KPEDTAVYYC AADSSYFYHT HESEYDYWGQ GTQVTVPPLV IWINGDKGYN GLAEVGKKFE KDTGIKVTVE H PDKLEEKF PQVAATGDGP DIIFWAHDRF GGYAQSGLLA EITPDKAFQD KLYPFTWDAV RYNGKLIAYP IAVEALSLIY NK DLLPNPP KTWEEIPALD KELKAKGKSA LMFNLQEPYF TWPLIAADGG YAFKYENGKY DIKDVGVDNA GAKAGLTFLV DLI KNKHMN ADTDYSIAEA AFNKGETAMT INGPWAWSNI DTSKVNYGVT VLPTFKGQPS KPFVGVLSAG INAASPNKEL AKEF LENYL LTDEGLEAVN KDKPLGAVAL KSYEEELAKD PRIAATMENA QKGEIMPNIP QMSAFWYAVR TAVINAASGR QTVDE ALKD AQTPGA

UniProtKB: Maltose/maltodextrin-binding periplasmic protein

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Macromolecule #6: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 6 / Number of copies: 1 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #7: omega-undecylenyl-beta-D-maltopyranoside

MacromoleculeName: omega-undecylenyl-beta-D-maltopyranoside / type: ligand / ID: 7 / Number of copies: 1 / Formula: 6UZ
Molecular weightTheoretical: 494.573 Da
Chemical component information

ChemComp-6UZ:
omega-undecylenyl-beta-D-maltopyranoside

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Macromolecule #8: (2S)-2-(2-oxidanylidenepyrrolidin-1-yl)butanamide

MacromoleculeName: (2S)-2-(2-oxidanylidenepyrrolidin-1-yl)butanamide / type: ligand / ID: 8 / Number of copies: 1 / Formula: UKX
Molecular weightTheoretical: 170.209 Da

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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

Concentration7 mg/mL
BufferpH: 7.5
Details: 10 mM Hepes-NaOH 7.5, 150 mM NaCl, 3 mM maltose, 0.03 % n-dodecyl-beta-D-maltopyranoside, 0.006% cholesterol hemisuccinate, 250 uM Levetiracetam
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 293 K / Instrument: GATAN CRYOPLUNGE 3

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number real images: 17809 / Average exposure time: 2.796 sec. / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 60000
Sample stageSpecimen holder model: JEOL CRYOSPECPORTER / Cooling holder cryogen: NITROGEN

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Image processing

CTF correctionSoftware - Name: cryoSPARC (ver. cryoSPARC-v.4) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. cryoSPARC-v.4) / Number images used: 158703
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-9fyr:
Cryo-EM structure of native SV2A in complex with TeNT-Hc, Pro-Macrobody 5 and Levetiracetam

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