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- EMDB-42431: Structure of synaptic vesicle protein 2A in complex with a nanobody -

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Basic information

Entry
Database: EMDB / ID: EMD-42431
TitleStructure of synaptic vesicle protein 2A in complex with a nanobody
Map dataDeepEMhancer sharpened map
Sample
  • Complex: Protein with a nanobody
    • Protein or peptide: Nanobody
  • Protein or peptide: Synaptic vesicle glycoprotein 2A
  • Ligand: (4S)-1-{[(4S)-2-(methoxymethyl)-6-(trifluoromethyl)imidazo[2,1-b][1,3,4]thiadiazol-5-yl]methyl}-4-(4,4,4-trifluorobutyl)pyrrolidin-2-one
  • Ligand: CHOLESTEROL HEMISUCCINATE
  • Ligand: 1,2-DIDECANOYL-SN-GLYCERO-3-[PHOSPHO-L-SERINE]
KeywordsSynaptic vesicle / SLC22 / Inhibitor / Nanobody / AEDs / TRANSPORT PROTEIN
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 / GABA-ergic synapse / neuromuscular junction ...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 / GABA-ergic synapse / neuromuscular junction / synaptic vesicle membrane / intracellular calcium ion homeostasis / cell-cell junction / synaptic vesicle / neuron projection / neuronal cell body / glutamatergic synapse / dendrite / protein kinase binding / endoplasmic reticulum / plasma membrane
Similarity search - Function
Synaptic vesicle protein SV2 / Major facilitator, sugar transporter-like / Sugar (and other) transporter / Sugar transporter, conserved site / Major facilitator superfamily / Major Facilitator Superfamily / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / MFS transporter superfamily
Similarity search - Domain/homology
Synaptic vesicle glycoprotein 2A
Similarity search - Component
Biological speciesHomo sapiens (human) / Lama glama (llama)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsMittal A / Martin MF / Levin E / Adams C / Yang M / Ledecq M / Horanyi PS / Coleman JA
Funding support United States, 1 items
OrganizationGrant numberCountry
Brain & Behavior Research Foundation30153 United States
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Structures of synaptic vesicle protein 2A and 2B bound to anticonvulsants.
Authors: Anshumali Mittal / Matthew F Martin / Elena J Levin / Christopher Adams / Meng Yang / Laurent Provins / Adrian Hall / Martin Procter / Marie Ledecq / Alexander Hillisch / Christian Wolff / ...Authors: Anshumali Mittal / Matthew F Martin / Elena J Levin / Christopher Adams / Meng Yang / Laurent Provins / Adrian Hall / Martin Procter / Marie Ledecq / Alexander Hillisch / Christian Wolff / Michel Gillard / Peter S Horanyi / Jonathan A Coleman /
Abstract: Epilepsy is a common neurological disorder characterized by abnormal activity of neuronal networks, leading to seizures. The racetam class of anti-seizure medications bind specifically to a membrane ...Epilepsy is a common neurological disorder characterized by abnormal activity of neuronal networks, leading to seizures. The racetam class of anti-seizure medications bind specifically to a membrane protein found in the synaptic vesicles of neurons called synaptic vesicle protein 2 (SV2) A (SV2A). SV2A belongs to an orphan subfamily of the solute carrier 22 organic ion transporter family that also includes SV2B and SV2C. The molecular basis for how anti-seizure medications act on SV2s remains unknown. Here we report cryo-electron microscopy structures of SV2A and SV2B captured in a luminal-occluded conformation complexed with anticonvulsant ligands. The conformation bound by anticonvulsants resembles an inhibited transporter with closed luminal and intracellular gates. Anticonvulsants bind to a highly conserved central site in SV2s. These structures provide blueprints for future drug design and will facilitate future investigations into the biological function of SV2s.
History
DepositionOct 19, 2023-
Header (metadata) releaseMay 22, 2024-
Map releaseMay 22, 2024-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_42431.png

Downloads & links

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Map

FileDownload / File: emd_42431.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationDeepEMhancer sharpened map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.65 Å/pix.
x 384 pix.
= 248.832 Å		0.65 Å/pix.
x 384 pix.
= 248.832 Å		0.65 Å/pix.
x 384 pix.
= 248.832 Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.648 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-0.021140268 - 1.9828718
Average (Standard dev.)0.0007726134 (±0.01688258)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 248.832 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_42431_msk_1.map
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Mask #2

Fileemd_42431_msk_2.map
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Additional map: Unsharpened map

Fileemd_42431_additional_1.map
AnnotationUnsharpened map
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Half map: half map A

Fileemd_42431_half_map_1.map
Annotationhalf map A
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Half map: half map B

Fileemd_42431_half_map_2.map
Annotationhalf map B
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Sample components

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Entire : Protein with a nanobody

EntireName: Protein with a nanobody
Components
  • Complex: Protein with a nanobody
    • Protein or peptide: Nanobody
  • Protein or peptide: Synaptic vesicle glycoprotein 2A
  • Ligand: (4S)-1-{[(4S)-2-(methoxymethyl)-6-(trifluoromethyl)imidazo[2,1-b][1,3,4]thiadiazol-5-yl]methyl}-4-(4,4,4-trifluorobutyl)pyrrolidin-2-one
  • Ligand: CHOLESTEROL HEMISUCCINATE
  • Ligand: 1,2-DIDECANOYL-SN-GLYCERO-3-[PHOSPHO-L-SERINE]

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Supramolecule #1: Protein with a nanobody

SupramoleculeName: Protein with a nanobody / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #2
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 98.1 kDa/nm

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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: Homo sapiens (human)
Molecular weightTheoretical: 75.589188 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGYYRGEGTQ DEEEGGASSD ATEGHDEDDE IYEGEYQGIP RAESGGKGER MADGAPLAGV RGGLSDGEGP PGGRGEAQRR KEREELAQQ YEAILRECGH GRFQWTLYFV LGLALMADGV EVFVVGFVLP SAEKDMCLSD SNKGMLGLIV YLGMMVGAFL W GGLADRLG ...String:
MGYYRGEGTQ DEEEGGASSD ATEGHDEDDE IYEGEYQGIP RAESGGKGER MADGAPLAGV RGGLSDGEGP PGGRGEAQRR KEREELAQQ YEAILRECGH GRFQWTLYFV LGLALMADGV EVFVVGFVLP SAEKDMCLSD SNKGMLGLIV YLGMMVGAFL W GGLADRLG RRQCLLISLS VNSVFAFFSS FVQGYGTFLF CRLLSGVGIG GSIPIVFSYF SEFLAQEKRG EHLSWLCMFW MI GGVYAAA MAWAIIPHYG WSFQMGSAYQ FHSWRVFVLV CAFPSVFAIG ALTTQPESPR FFLENGKHDE AWMVLKQVHD TNM RAKGHP ERVFSVTHIK TIHQEDELIE IQSDTGTWYQ RWGVRALSLG GQVWGNFLSC FGPEYRRITL MMMGVWFTMS FSYY GLTVW FPDMIRHLQA VDYASRTKVF PGERVEHVTF NFTLENQIHR GGQYFNDKFI GLRLKSVSFE DSLFEECYFE DVTSS NTFF RNCTFINTVF YNTDLFEYKF VNSRLINSTF LHNKEGCPLD VTGTGEGAYM VYFVSFLGTL AVLPGNIVSA LLMDKI GRL RMLAGSSVMS CVSCFFLSFG NSESAMIALL CLFGGVSIAS WNALDVLTVE LYPSDKRTTA FGFLNALCKL AAVLGIS IF TSFVGITKAA PILFASAALA LGSSLALKLP ETRGQVLQ

UniProtKB: Synaptic vesicle glycoprotein 2A

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Macromolecule #2: Nanobody

MacromoleculeName: Nanobody / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Lama glama (llama)
Molecular weightTheoretical: 15.469103 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
QVQLVESGGG LVQPGGSLRL SCAASGSIFN MRVMGWYRQA PGEQRESVAS MASGDKTTYA DSVKGRFTIS RDNAKNTVAL QMNSLKPED TAVYYCHAVD LTRNGPRVYW GQGTQVTVSS AAAENLYFQG GSGHHHHHHH HHH

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Macromolecule #5: (4S)-1-{[(4S)-2-(methoxymethyl)-6-(trifluoromethyl)imidazo[2,1-b]...

MacromoleculeName: (4S)-1-{[(4S)-2-(methoxymethyl)-6-(trifluoromethyl)imidazo[2,1-b][1,3,4]thiadiazol-5-yl]methyl}-4-(4,4,4-trifluorobutyl)pyrrolidin-2-one
type: ligand / ID: 5 / Number of copies: 1 / Formula: X49
Molecular weightTheoretical: 444.395 Da

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Macromolecule #6: CHOLESTEROL HEMISUCCINATE

MacromoleculeName: CHOLESTEROL HEMISUCCINATE / type: ligand / ID: 6 / Number of copies: 1 / Formula: Y01
Molecular weightTheoretical: 486.726 Da
Chemical component information

ChemComp-Y01:
CHOLESTEROL HEMISUCCINATE

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Macromolecule #7: 1,2-DIDECANOYL-SN-GLYCERO-3-[PHOSPHO-L-SERINE]

MacromoleculeName: 1,2-DIDECANOYL-SN-GLYCERO-3-[PHOSPHO-L-SERINE] / type: ligand / ID: 7 / Number of copies: 1 / Formula: PS1
Molecular weightTheoretical: 566.642 Da
Chemical component information

ChemComp-PS1:
1,2-DIDECANOYL-SN-GLYCERO-3-[PHOSPHO-L-SERINE]

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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

Concentration4.38 mg/mL
BufferpH: 8
GridModel: Quantifoil R2/1 / Material: GOLD / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / 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: 2.8000000000000003 µm / Nominal defocus min: 0.75 µm / Nominal magnification: 77160
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing #1

Image processing ID1
Startup modelType of model: OTHER / Details: Ab-initio reconstruction
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 250856
Initial angle assignmentType: RANDOM ASSIGNMENT
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Image processing #2

Image processing ID2
Startup modelType of model: OTHER / Details: Ab-initio reconstruction
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 250856
Initial angle assignmentType: RANDOM ASSIGNMENT
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

3-f1


Chain - Source name: AlphaFold / Chain - Initial model type: in silico model
Output model

PDB-8uo9:
Structure of synaptic vesicle protein 2A in complex with a nanobody

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