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- PDB-8y2e: Cryo-EM structure of human dopamine transporter in complex with b... -

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Basic information

Entry
Database: PDB / ID: 8y2e
TitleCryo-EM structure of human dopamine transporter in complex with benztropine
ComponentsSodium-dependent dopamine transporter
KeywordsMEMBRANE PROTEIN / human dopamine transporter in complex with benztropine
Function / homology
Function and homology information


Defective SLC6A3 causes Parkinsonism-dystonia infantile (PKDYS) / Defective SLC6A3 causes Parkinsonism-dystonia infantile (PKDYS) / Dopamine clearance from the synaptic cleft / amine binding / adenohypophysis development / dopamine uptake / hyaloid vascular plexus regression / dopamine binding / norepinephrine:sodium symporter activity / dopamine:sodium symporter activity ...Defective SLC6A3 causes Parkinsonism-dystonia infantile (PKDYS) / Defective SLC6A3 causes Parkinsonism-dystonia infantile (PKDYS) / Dopamine clearance from the synaptic cleft / amine binding / adenohypophysis development / dopamine uptake / hyaloid vascular plexus regression / dopamine binding / norepinephrine:sodium symporter activity / dopamine:sodium symporter activity / norepinephrine transport / regulation of dopamine metabolic process / neurotransmitter transmembrane transporter activity / dopaminergic synapse / dopamine transport / flotillin complex / dopamine catabolic process / monoamine transmembrane transporter activity / monoamine transport / Na+/Cl- dependent neurotransmitter transporters / positive regulation of multicellular organism growth / response to iron ion / neurotransmitter transport / dopamine biosynthetic process / heterocyclic compound binding / amino acid transport / dopamine uptake involved in synaptic transmission / neuronal cell body membrane / sodium ion transmembrane transport / prepulse inhibition / axon terminus / response to cAMP / lactation / protein phosphatase 2A binding / response to cocaine / cognition / locomotory behavior / response to nicotine / sensory perception of smell / presynaptic membrane / protease binding / postsynaptic membrane / response to ethanol / neuron projection / response to xenobiotic stimulus / membrane raft / axon / signaling receptor binding / neuronal cell body / protein-containing complex binding / cell surface / membrane / metal ion binding / plasma membrane / cytoplasm
Similarity search - Function
Sodium:neurotransmitter symporter, dopamine / Sodium:neurotransmitter symporter family signature 2. / Sodium:neurotransmitter symporter family signature 1. / Sodium:neurotransmitter symporter / Sodium:neurotransmitter symporter superfamily / Sodium:neurotransmitter symporter family / Sodium:neurotransmitter symporter family profile.
Similarity search - Domain/homology
benztropine / Sodium-dependent dopamine transporter
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.03 Å
AuthorsZhao, Y. / Li, Y.
Funding support China, 1items
OrganizationGrant numberCountry
Chinese Academy of Sciences China
CitationJournal: Nature / Year: 2024
Title: Dopamine reuptake and inhibitory mechanisms in human dopamine transporter.
Authors: Yue Li / Xianping Wang / Yufei Meng / Tuo Hu / Jun Zhao / Renjie Li / Qinru Bai / Pu Yuan / Jun Han / Kun Hao / Yiqing Wei / Yunlong Qiu / Na Li / Yan Zhao /
Abstract: The dopamine transporter has a crucial role in regulation of dopaminergic neurotransmission by uptake of dopamine into neurons and contributes to the abuse potential of psychomotor stimulants. ...The dopamine transporter has a crucial role in regulation of dopaminergic neurotransmission by uptake of dopamine into neurons and contributes to the abuse potential of psychomotor stimulants. Despite decades of study, the structure, substrate binding, conformational transitions and drug-binding poses of human dopamine transporter remain unknown. Here we report structures of the human dopamine transporter in its apo state, and in complex with the substrate dopamine, the attention deficit hyperactivity disorder drug methylphenidate, and the dopamine-uptake inhibitors GBR12909 and benztropine. The dopamine-bound structure in the occluded state precisely illustrates the binding position of dopamine and associated ions. The structures bound to drugs are captured in outward-facing or inward-facing states, illuminating distinct binding modes and conformational transitions during substrate transport. Unlike the outward-facing state, which is stabilized by cocaine, GBR12909 and benztropine stabilize the dopamine transporter in the inward-facing state, revealing previously unseen drug-binding poses and providing insights into how they counteract the effects of cocaine. This study establishes a framework for understanding the functioning of the human dopamine transporter and developing therapeutic interventions for dopamine transporter-related disorders and cocaine addiction.
History
DepositionJan 25, 2024Deposition site: PDBJ / Processing site: PDBC
Revision 1.0Aug 14, 2024Provider: repository / Type: Initial release
Revision 1.1Aug 21, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.pdbx_database_id_PubMed / _citation.title / _em_admin.last_update
Revision 1.2Aug 28, 2024Group: Data collection / Database references / Category: citation / em_admin
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Sodium-dependent dopamine transporter
hetero molecules


Theoretical massNumber of molelcules
Total (without water)62,3404
Polymers61,5911
Non-polymers7503
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: Protein Sodium-dependent dopamine transporter / DA transporter / DAT / Solute carrier family 6 member 3


Mass: 61590.508 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SLC6A3, DAT1 / Production host: Homo sapiens (human) / References: UniProt: Q01959
#2: Sugar ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
#3: Chemical ChemComp-CXQ / benztropine


Mass: 307.429 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C21H25NO / Feature type: SUBJECT OF INVESTIGATION / Comment: medication, antipsychotic*YM
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 dopamine transporter in complex with benztropine
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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: 2200 nm / Nominal defocus min: 1200 nm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING ONLY
3D reconstructionResolution: 3.03 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 63350 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0034451
ELECTRON MICROSCOPYf_angle_d0.5386077
ELECTRON MICROSCOPYf_dihedral_angle_d4.118617
ELECTRON MICROSCOPYf_chiral_restr0.038692
ELECTRON MICROSCOPYf_plane_restr0.005744

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