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- PDB-8thr: Structure of the human vesicular monoamine transporter 2 (VMAT2) ... -

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Basic information

Entry
Database: PDB / ID: 8thr
TitleStructure of the human vesicular monoamine transporter 2 (VMAT2) bound to tetrabenazine in an occluded conformation
Componentsfluorescent protein mVenus,Synaptic vesicular amine transporter,GFP nano body,Synaptic vesicular amine transporter,Synaptic vesicular amine transporter
KeywordsTRANSPORT PROTEIN / monoamine / neurotransmitter / synaptic vesicles / major facilitator superfamily / SLC18
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 / dopamine transport / dopaminergic synapse / monoamine transmembrane transporter activity / histamine secretion by mast cell / monoamine transport / Na+/Cl- dependent neurotransmitter transporters / 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
Chem-EBZ / Synaptic vesicular amine transporter
Similarity search - Component
Biological speciesAequorea victoria (jellyfish)
Homo sapiens (human)
Vicugna pacos (alpaca)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.12 Å
AuthorsDalton, M.P. / Coleman, J.A.
Funding support1items
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, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 25, 2023Provider: repository / Type: Initial release
Revision 1.1May 8, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: fluorescent protein mVenus,Synaptic vesicular amine transporter,GFP nano body,Synaptic vesicular amine transporter,Synaptic vesicular amine transporter
hetero molecules


Theoretical massNumber of molelcules
Total (without water)94,1682
Polymers93,8511
Non-polymers3171
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Antibody fluorescent protein mVenus,Synaptic vesicular amine transporter,GFP nano body,Synaptic vesicular amine transporter,Synaptic vesicular amine transporter


Mass: 93850.930 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Aequorea victoria (jellyfish), (gene. exp.) Homo sapiens (human), (gene. exp.) Vicugna pacos (alpaca)
Gene: SLC18A2 / Cell line (production host): HEK293 GnTi- / Production host: Homo sapiens (human) / References: UniProt: Q05940
#2: Chemical 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


Mass: 317.423 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C19H27NO3 / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Human VMAT2 complexed with Tetrabenazine / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: .0937 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
Details: 1 mM DDM, .125 mM CHS, 1 mM DTT, 100 uM TBZ, 25 mM Tris HCl pH 8, 150 mM NaCl
SpecimenConc.: 6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 25 mA / Grid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 77279 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV

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Processing

EM software
IDNameVersionCategory
2SerialEMimage acquisition
7Coot0.98model fitting
8ISOLDEmodel fitting
10PHENIX1.2model refinement
14cryoSPARC4.2.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.12 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 65516 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingAccession code: AF-Q05940-F1 / Source name: AlphaFold / Type: in silico model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0043007
ELECTRON MICROSCOPYf_angle_d0.6834090
ELECTRON MICROSCOPYf_dihedral_angle_d9.925414
ELECTRON MICROSCOPYf_chiral_restr0.041487
ELECTRON MICROSCOPYf_plane_restr0.006501

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