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- PDB-8wvg: Transporter bound with inhibitor -

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Basic information

Entry
Database: PDB / ID: 8wvg
TitleTransporter bound with inhibitor
Components
  • FabH
  • FabL
  • Synaptic vesicular amine transporter
KeywordsMEMBRANE PROTEIN / transporter
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
: / Synaptic vesicular amine transporter
Similarity search - Component
Biological speciesHomo sapiens (human)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.18 Å
AuthorsIm, D. / Iwata, S.
Funding support Japan, 1items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS) Japan
CitationJournal: Nat Commun / Year: 2024
Title: Neurotransmitter recognition by human vesicular monoamine transporter 2.
Authors: Dohyun Im / Mika Jormakka / Narinobu Juge / Jun-Ichi Kishikawa / Takayuki Kato / Yukihiko Sugita / Takeshi Noda / Tomoko Uemura / Yuki Shiimura / Takaaki Miyaji / Hidetsugu Asada / So Iwata /
Abstract: Human vesicular monoamine transporter 2 (VMAT2), a member of the SLC18 family, plays a crucial role in regulating neurotransmitters in the brain by facilitating their uptake and storage within ...Human vesicular monoamine transporter 2 (VMAT2), a member of the SLC18 family, plays a crucial role in regulating neurotransmitters in the brain by facilitating their uptake and storage within vesicles, preparing them for exocytotic release. Because of its central role in neurotransmitter signalling and neuroprotection, VMAT2 is a target for neurodegenerative diseases and movement disorders, with its inhibitor being used as therapeutics. Despite the importance of VMAT2 in pharmacophysiology, the molecular basis of VMAT2-mediated neurotransmitter transport and its inhibition remains unclear. Here we show the cryo-electron microscopy structure of VMAT2 in the substrate-free state, in complex with the neurotransmitter dopamine, and in complex with the inhibitor tetrabenazine. In addition to these structural determinations, monoamine uptake assays, mutational studies, and pKa value predictions were performed to characterize the dynamic changes in VMAT2 structure. These results provide a structural basis for understanding VMAT2-mediated vesicular transport of neurotransmitters and a platform for modulation of current inhibitor design.
History
DepositionOct 23, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Sep 25, 2024Provider: repository / Type: Initial release
Revision 1.1Oct 2, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.page_first / _citation.page_last ..._citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation_author.identifier_ORCID / _citation_author.name / _em_admin.last_update
Revision 1.2Nov 20, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
H: FabH
L: FabL
A: Synaptic vesicular amine transporter
hetero molecules


Theoretical massNumber of molelcules
Total (without water)116,9104
Polymers116,5933
Non-polymers3171
Water181
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 FabH


Mass: 36424.773 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#2: Antibody FabL


Mass: 24418.936 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#3: Protein Synaptic vesicular amine transporter


Mass: 55749.352 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC18A2 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q05940
#4: Chemical ChemComp-XEQ / (3~{S},11~{b}~{R})-9,10-dimethoxy-3-(2-methylpropyl)-1,3,4,6,7,11~{b}-hexahydrobenzo[a]quinolizin-2-one


Mass: 317.423 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C19H27NO3 / Feature type: SUBJECT OF INVESTIGATION
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY
Has protein modificationY

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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: VMAT2-Fab0801 bound with inhibitor / Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT
Molecular weightValue: 0.09 MDa / Experimental value: YES
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.5
SpecimenConc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 281.15 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 defocus max: 1800 nm / Nominal defocus min: 800 nm
Specimen holderSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 65 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.21rc1_5109: / Category: model refinement
CTF correctionType: NONE
3D reconstructionResolution: 3.18 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 697706 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0056516
ELECTRON MICROSCOPYf_angle_d0.6398871
ELECTRON MICROSCOPYf_dihedral_angle_d6.74900
ELECTRON MICROSCOPYf_chiral_restr0.0451022
ELECTRON MICROSCOPYf_plane_restr0.0051107

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