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- EMDB-41269: Structure of the human vesicular monoamine transporter 2 (VMAT2) ... -

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Basic information

Entry
Database: EMDB / ID: EMD-41269
TitleStructure of the human vesicular monoamine transporter 2 (VMAT2) bound to Tetrabenazine in an occluded conformation
Map dataDeepEMhancer sharpened with local mask
Sample
  • Complex: Human VMAT2 complexed with Tetrabenazine
    • Protein or peptide: fluorescent protein mVenus,Synaptic vesicular amine transporter,GFP nano body,Synaptic vesicular amine transporter,Synaptic vesicular amine transporterFluorescence
  • Ligand: (3S,5R,11bS)-9,10-dimethoxy-3-(2-methylpropyl)-1,3,4,6,7,11b-hexahydro-2H-pyrido[2,1-a]isoquinolin-2-one
Keywordsmonoamine / neurotransmitter / synaptic vesicles / major facilitator superfamily / SLC18 / TRANSPORT PROTEIN
Function / homology
Function and homology information


serotonin secretion by mast cell / sequestering of neurotransmitter / somato-dendritic dopamine secretion / histamine uptake / neurotransmitter loading into synaptic vesicle / monoamine:proton antiporter activity / aminergic neurotransmitter loading into synaptic vesicle / clathrin-sculpted monoamine transport vesicle membrane / Serotonin Neurotransmitter Release Cycle / serotonin:sodium:chloride symporter activity ...serotonin secretion by mast cell / sequestering of neurotransmitter / somato-dendritic dopamine secretion / histamine uptake / neurotransmitter loading into synaptic vesicle / monoamine:proton antiporter activity / aminergic neurotransmitter loading into synaptic vesicle / clathrin-sculpted monoamine transport vesicle membrane / Serotonin Neurotransmitter Release Cycle / serotonin:sodium:chloride symporter activity / Dopamine Neurotransmitter Release Cycle / Norepinephrine Neurotransmitter Release Cycle / serotonin uptake / dopaminergic synapse / dopamine transport / Na+/Cl- dependent neurotransmitter transporters / monoamine transmembrane transporter activity / monoamine transport / histamine secretion by mast cell / neurotransmitter transport / negative regulation of reactive oxygen species biosynthetic process / response to amphetamine / post-embryonic development / secretory granule membrane / locomotory behavior / terminal bouton / response to toxic substance / synaptic vesicle membrane / synaptic vesicle / chemical synaptic transmission / axon / intracellular membrane-bounded organelle / centrosome / dendrite / membrane / plasma membrane
Similarity search - Function
Major facilitator superfamily / Major Facilitator Superfamily / Major facilitator superfamily domain / Major facilitator superfamily (MFS) profile. / MFS transporter superfamily
Similarity search - Domain/homology
Synaptic vesicular amine transporter
Similarity search - Component
Biological speciesHomo sapiens (human) / Vicugna pacos (alpaca)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.12 Å
AuthorsDalton MP / Coleman JA
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Elife / Year: 2024
Title: Structural mechanisms for VMAT2 inhibition by tetrabenazine.
Authors: Michael P Dalton / Mary Hongying Cheng / Ivet Bahar / Jonathan A Coleman /
Abstract: The vesicular monoamine transporter 2 (VMAT2) is a proton-dependent antiporter responsible for loading monoamine neurotransmitters into synaptic vesicles. Dysregulation of VMAT2 can lead to several ...The vesicular monoamine transporter 2 (VMAT2) is a proton-dependent antiporter responsible for loading monoamine neurotransmitters into synaptic vesicles. Dysregulation of VMAT2 can lead to several neuropsychiatric disorders including Parkinson's disease and schizophrenia. Furthermore, drugs such as amphetamine and MDMA are known to act on VMAT2, exemplifying its role in the mechanisms of actions for drugs of abuse. Despite VMAT2's importance, there remains a critical lack of mechanistic understanding, largely driven by a lack of structural information. Here, we report a 3.1 Å resolution cryo-electron microscopy (cryo-EM) structure of VMAT2 complexed with tetrabenazine (TBZ), a non-competitive inhibitor used in the treatment of Huntington's chorea. We find TBZ interacts with residues in a central binding site, locking VMAT2 in an occluded conformation and providing a mechanistic basis for non-competitive inhibition. We further identify residues critical for cytosolic and lumenal gating, including a cluster of hydrophobic residues which are involved in a lumenal gating strategy. Our structure also highlights three distinct polar networks that may determine VMAT2 conformational dynamics and play a role in proton transduction. The structure elucidates mechanisms of VMAT2 inhibition and transport, providing insights into VMAT2 architecture, function, and the design of small-molecule therapeutics.
History
DepositionJul 17, 2023-
Header (metadata) releaseOct 25, 2023-
Map releaseOct 25, 2023-
UpdateMay 8, 2024-
Current statusMay 8, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41269.png

Downloads & links

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Map

FileDownload / File: emd_41269.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationDeepEMhancer sharpened with local mask
Voxel sizeX=Y=Z: 0.647 Å
Density
Contour LevelBy AUTHOR: 0.338
Minimum - Maximum-0.0010661092 - 2.9606125
Average (Standard dev.)0.0010804834 (±0.02661487)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 248.448 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_41269_msk_1.map
Projections & Slices
AxesZYX

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Additional map: unsharpened map

Fileemd_41269_additional_1.map
Annotationunsharpened map
Projections & Slices
AxesZYX

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Additional map: DeepEMhancer sharpened with a solvent mask

Fileemd_41269_additional_2.map
AnnotationDeepEMhancer sharpened with a solvent mask
Projections & Slices
AxesZYX

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Histogram

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Half map: half map A

Fileemd_41269_half_map_1.map
Annotationhalf map A
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Half map: half map B

Fileemd_41269_half_map_2.map
Annotationhalf map B
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Sample components

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Entire : Human VMAT2 complexed with Tetrabenazine

EntireName: Human VMAT2 complexed with Tetrabenazine
Components
  • Complex: Human VMAT2 complexed with Tetrabenazine
    • Protein or peptide: fluorescent protein mVenus,Synaptic vesicular amine transporter,GFP nano body,Synaptic vesicular amine transporter,Synaptic vesicular amine transporterFluorescence
  • Ligand: (3S,5R,11bS)-9,10-dimethoxy-3-(2-methylpropyl)-1,3,4,6,7,11b-hexahydro-2H-pyrido[2,1-a]isoquinolin-2-one

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Supramolecule #1: Human VMAT2 complexed with Tetrabenazine

SupramoleculeName: Human VMAT2 complexed with Tetrabenazine / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 93.7 KDa

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Macromolecule #1: fluorescent protein mVenus,Synaptic vesicular amine transporter,G...

MacromoleculeName: fluorescent protein mVenus,Synaptic vesicular amine transporter,GFP nano body,Synaptic vesicular amine transporter,Synaptic vesicular amine transporter
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Vicugna pacos (alpaca)
Molecular weightTheoretical: 93.85093 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MWSHPQFEKG GGSGGGSGGS AWSHPQFEKH HHHHHHHHHG SGVSKGEELF TGVVPILVEL DGDVNGHKFS VSGEGEGDAT YGKLTLKLI CTTGKLPVPW PTLVTTLGYG LQCFARYPDH MKQHDFFKSA MPEGYVQERT IFFKDDGNYK TRAEVKFEGD T LVNRIELK ...String:
MWSHPQFEKG GGSGGGSGGS AWSHPQFEKH HHHHHHHHHG SGVSKGEELF TGVVPILVEL DGDVNGHKFS VSGEGEGDAT YGKLTLKLI CTTGKLPVPW PTLVTTLGYG LQCFARYPDH MKQHDFFKSA MPEGYVQERT IFFKDDGNYK TRAEVKFEGD T LVNRIELK GIDFKEDGNI LGHKLEYNYN SHNVYITADK QKNGIKANFK IRHNIEDGGV QLADHYQQNT PIGDGPVLLP DN HYLSYQS KLSKDPNEKR DHMVLLEFVT AAGITLGMDE LYKSRKLILF IVFLALLLDN MLLTVVVPII PSYLYSIKHE KNA TEIQTA RPVHTASISD SFQSIFSYYD NSTMVTGNAT RDLTLHQTAT QHMVTNASAV PSDCPSEDKD LLNENVQVGL LFAS KATVQ LITNPFIGLL TNRIGYPIPI FAGFCIMFVS TIMFAFSSSY AFLLIARSLQ GIGSSCSSVA GMGMLASVYT DDEER GNVM GIALGGLAMG VLVGPPFGSV LYEFVGKTAP FLVLAALVLL DGAIQLFVLQ PSRVQPESQK GTPLTTLLKD PYILIA AGS ICFANMGIAM LEPALPIWMM ETMCSRKWQL GVAFLPASIS YLIGTNIFGI LAHKMGRWLC ALLGMIIVGV SILCIPF AK NIYGLIAPNF GVGFAIGMVD SSMMPIMGYL VDLRHVSVYG SVYAIADVAF CMGYAIGPSA GGAIAKAIGF PWLMTIIG I IDILFAPLCF FLRSPPAKEE KMAIVQLVES GGALVQPGGS LRLSCAASGF PVNRYSMRWY RQAPGKEREW VAGMSSAGD RSSYEDSVKG RFTISRDDAR NTVYLQMNSL KPEDTAVYYC NVNVGFEYWG QGTQVTVS

UniProtKB: Synaptic vesicular amine transporter

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Macromolecule #2: (3S,5R,11bS)-9,10-dimethoxy-3-(2-methylpropyl)-1,3,4,6,7,11b-hexa...

MacromoleculeName: (3S,5R,11bS)-9,10-dimethoxy-3-(2-methylpropyl)-1,3,4,6,7,11b-hexahydro-2H-pyrido[2,1-a]isoquinolin-2-one
type: ligand / ID: 2 / Number of copies: 1 / Formula: EBZ
Molecular weightTheoretical: 317.423 Da
Chemical component information

ChemComp-EBZ:
(3S,5R,11bS)-9,10-dimethoxy-3-(2-methylpropyl)-1,3,4,6,7,11b-hexahydro-2H-pyrido[2,1-a]isoquinolin-2-one

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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

Concentration6 mg/mL
BufferpH: 8
Details: 1 mM DDM, .125 mM CHS, 1 mM DTT, 100 uM TBZ, 25 mM Tris HCl pH 8, 150 mM NaCl
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR / Details: 25 mA
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 77279
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 60.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER / Details: ab-initio model created in cryosparc
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.12 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 65516
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

0-f1


Chain - Source name: AlphaFold / Chain - Initial model type: in silico model
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8thr:
Structure of the human vesicular monoamine transporter 2 (VMAT2) bound to tetrabenazine in an occluded conformation

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