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Yorodumi- EMDB-19979: Inhibitor-free outward-open structure of Drosophila dopamine tran... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-19979 | ||||||||||||||||||||||||||||||||||||
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Title | Inhibitor-free outward-open structure of Drosophila dopamine transporter | ||||||||||||||||||||||||||||||||||||
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Sample |
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Keywords | SLC6A3 / Dopamine transporter / neurotransmitter sodium symporters / MEMBRANE PROTEIN | ||||||||||||||||||||||||||||||||||||
Function / homology | Function and homology information Dopamine clearance from the synaptic cleft / Na+/Cl- dependent neurotransmitter transporters / circadian sleep/wake cycle / cocaine binding / dopamine:sodium symporter activity / norepinephrine:sodium symporter activity / norepinephrine transport / response to odorant / regulation of presynaptic cytosolic calcium ion concentration / dopamine transport ...Dopamine clearance from the synaptic cleft / Na+/Cl- dependent neurotransmitter transporters / circadian sleep/wake cycle / cocaine binding / dopamine:sodium symporter activity / norepinephrine:sodium symporter activity / norepinephrine transport / response to odorant / regulation of presynaptic cytosolic calcium ion concentration / dopamine transport / sleep / dopamine uptake involved in synaptic transmission / amino acid transport / neuronal cell body membrane / sodium ion transmembrane transport / adult locomotory behavior / presynaptic membrane / axon / metal ion binding / plasma membrane Similarity search - Function | ||||||||||||||||||||||||||||||||||||
Biological species | Drosophila melanogaster (fruit fly) / Mus musculus (house mouse) | ||||||||||||||||||||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.0 Å | ||||||||||||||||||||||||||||||||||||
Authors | Pedersen CN / Yang F / Ita S / Xu Y / Akunuri R / Trampari S / Neumann CMT / Desdorf LM / Schioett B / Salvino JM ...Pedersen CN / Yang F / Ita S / Xu Y / Akunuri R / Trampari S / Neumann CMT / Desdorf LM / Schioett B / Salvino JM / Mortensen OV / Nissen P / Shahsavar A | ||||||||||||||||||||||||||||||||||||
Funding support | Denmark, United States, 11 items
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Citation | Journal: J Neurochem / Year: 2024 Title: Cryo-EM structure of the dopamine transporter with a novel atypical non-competitive inhibitor bound to the orthosteric site. Authors: Clara Nautrup Pedersen / Fuyu Yang / Samantha Ita / Yibin Xu / Ravikumar Akunuri / Sofia Trampari / Caroline Marie Teresa Neumann / Lasse Messell Desdorf / Birgit Schiøtt / Joseph M Salvino ...Authors: Clara Nautrup Pedersen / Fuyu Yang / Samantha Ita / Yibin Xu / Ravikumar Akunuri / Sofia Trampari / Caroline Marie Teresa Neumann / Lasse Messell Desdorf / Birgit Schiøtt / Joseph M Salvino / Ole Valente Mortensen / Poul Nissen / Azadeh Shahsavar / Abstract: The regulation of dopamine (DA) removal from the synaptic cleft is a crucial process in neurotransmission and is facilitated by the sodium- and chloride-coupled dopamine transporter DAT. ...The regulation of dopamine (DA) removal from the synaptic cleft is a crucial process in neurotransmission and is facilitated by the sodium- and chloride-coupled dopamine transporter DAT. Psychostimulant drugs, cocaine, and amphetamine, both block the uptake of DA, while amphetamine also triggers the release of DA. As a result, they prolong or even amplify neurotransmitter signaling. Atypical inhibitors of DAT lack cocaine-like rewarding effects and offer a promising strategy for the treatment of drug use disorders. Here, we present the 3.2 Å resolution cryo-electron microscopy structure of the Drosophila melanogaster dopamine transporter (dDAT) in complex with the atypical non-competitive inhibitor AC-4-248. The inhibitor partially binds at the central binding site, extending into the extracellular vestibule, and locks the transporter in an outward open conformation. Our findings propose mechanisms for the non-competitive inhibition of DAT and attenuation of cocaine potency by AC-4-248 and provide a basis for the rational design of more efficacious atypical inhibitors. | ||||||||||||||||||||||||||||||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_19979.map.gz | 59.6 MB | EMDB map data format | |
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Header (meta data) | emd-19979-v30.xml emd-19979.xml | 24.2 KB 24.2 KB | Display Display | EMDB header |
Images | emd_19979.png | 49.7 KB | ||
Filedesc metadata | emd-19979.cif.gz | 7.3 KB | ||
Others | emd_19979_half_map_1.map.gz emd_19979_half_map_2.map.gz | 59.5 MB 59.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-19979 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-19979 | HTTPS FTP |
-Validation report
Summary document | emd_19979_validation.pdf.gz | 795.3 KB | Display | EMDB validaton report |
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Full document | emd_19979_full_validation.pdf.gz | 794.9 KB | Display | |
Data in XML | emd_19979_validation.xml.gz | 12.5 KB | Display | |
Data in CIF | emd_19979_validation.cif.gz | 14.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-19979 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-19979 | HTTPS FTP |
-Related structure data
Related structure data | 9eupMC 9euoC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_19979.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.294 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: Half map A
File | emd_19979_half_map_1.map | ||||||||||||
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Annotation | Half map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map B
File | emd_19979_half_map_2.map | ||||||||||||
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Annotation | Half map B | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Inhibitor-free Drosophila melanogaster dopamine transporter in co...
Entire | Name: Inhibitor-free Drosophila melanogaster dopamine transporter in complex with Fab 9D5 |
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Components |
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-Supramolecule #1: Inhibitor-free Drosophila melanogaster dopamine transporter in co...
Supramolecule | Name: Inhibitor-free Drosophila melanogaster dopamine transporter in complex with Fab 9D5 type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
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Source (natural) | Organism: Drosophila melanogaster (fruit fly) |
Molecular weight | Theoretical: 107.602 KDa |
-Macromolecule #1: Sodium-dependent dopamine transporter
Macromolecule | Name: Sodium-dependent dopamine transporter / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Drosophila melanogaster (fruit fly) |
Molecular weight | Theoretical: 60.93952 KDa |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: MNSISDERET WSGKVDFLLS VIGFAVDLAN VWRFPYLCYK NGGGAFLVPY GIMLAVGGIP LFYMELALGQ HNRKGAITCW GRLVPLFKG IGYAVVLIAF YVDFYYNVII AWSLRFFFAS FTNSLPWTSC NNIWNTPNCR PFESQGFQSA ASEYFNRYIL E LNRSEGIH ...String: MNSISDERET WSGKVDFLLS VIGFAVDLAN VWRFPYLCYK NGGGAFLVPY GIMLAVGGIP LFYMELALGQ HNRKGAITCW GRLVPLFKG IGYAVVLIAF YVDFYYNVII AWSLRFFFAS FTNSLPWTSC NNIWNTPNCR PFESQGFQSA ASEYFNRYIL E LNRSEGIH DLGAIKWDMA LCLLIVYLIC YFSLWKGIST SGKVVWFTAL FPYAALLILL IRGLTLPGSF LGIQYYLTPN FS AIYKAEV WADAATQVFF SLGPGFGVLL AYASYNKYHN NVYKDALLTS FINSATSFIA GFVIFSVLGY MAHTLGVRIE DVA TEGPGL VFVVYPAAIA TMPASTFWAL IFFMMLATLG LDSSFGGSEA IITALSDEFP KIKRNRELFV AGLFSLYFVV GLAS CTQGG FYFFHLLDRY AAGYSILVAV FFEAIAVSWI YGTNRFSEDI RDMIGFPPGR YWQVCWRFVA PIFLLFITVY LLIGY EPLT YADYVYPSWA NALGWCIAGS SVVMIPAVAI FKLLSTPGSL RQRFTILTTP WRDQQLVPR UniProtKB: Sodium-dependent dopamine transporter |
-Macromolecule #2: 9D5 ANTIBODY, HEAVY CHAIN
Macromolecule | Name: 9D5 ANTIBODY, HEAVY CHAIN / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Mus musculus (house mouse) |
Molecular weight | Theoretical: 25.921338 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MNFGLRLVFL VLILKGVQCE VQLVESGGGL VKPGGSLKLS CAASGFTFSS YAMSWVRQSP EKRLEWVAEI SSGGRYIYYS DTVTGRFTI SRDNARNILH LEMSSLRSED TAMYYCARGE VRQRGFDYWG QGTTLTVSSA KTTAPSVYPL APVCGDTTGS S VTLGCLVK ...String: MNFGLRLVFL VLILKGVQCE VQLVESGGGL VKPGGSLKLS CAASGFTFSS YAMSWVRQSP EKRLEWVAEI SSGGRYIYYS DTVTGRFTI SRDNARNILH LEMSSLRSED TAMYYCARGE VRQRGFDYWG QGTTLTVSSA KTTAPSVYPL APVCGDTTGS S VTLGCLVK GYFPEPVTLT WNSGSLSSGV HTFPAVLQSD LYTLSSSVTV TSSTWPSQSI TCNVAHPASS TKVDKKIEPR GP |
-Macromolecule #3: 9D5 ANTIBODY, LIGHT CHAIN
Macromolecule | Name: 9D5 ANTIBODY, LIGHT CHAIN / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO |
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Source (natural) | Organism: Mus musculus (house mouse) |
Molecular weight | Theoretical: 25.840607 KDa |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MDFQVQIFSF LLISASVAMS RGENVLTQSP AIMSTSPGEK VTMTCRASSS VGSSYLHWYQ QKSGASPKLW IYSTSNLASG VPARFSGSG SGTSYSLTIS SVEAEDAATY YCQQFSGYPL TFGSGTKLEM KRADAAPTVS IFPPSSEQLT SGGASVVCFL N NFYPKDIN ...String: MDFQVQIFSF LLISASVAMS RGENVLTQSP AIMSTSPGEK VTMTCRASSS VGSSYLHWYQ QKSGASPKLW IYSTSNLASG VPARFSGSG SGTSYSLTIS SVEAEDAATY YCQQFSGYPL TFGSGTKLEM KRADAAPTVS IFPPSSEQLT SGGASVVCFL N NFYPKDIN VKWKIDGSER QNGVLNSWTD QDSKDSTYSM SSTLTLTKDE YERHNSYTCE ATHKTSTSPI VKSFNRNEC |
-Macromolecule #4: CHOLESTEROL HEMISUCCINATE
Macromolecule | Name: CHOLESTEROL HEMISUCCINATE / type: ligand / ID: 4 / Number of copies: 1 / Formula: Y01 |
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Molecular weight | Theoretical: 486.726 Da |
Chemical component information | ChemComp-Y01: |
-Macromolecule #5: TRIS-HYDROXYMETHYL-METHYL-AMMONIUM
Macromolecule | Name: TRIS-HYDROXYMETHYL-METHYL-AMMONIUM / type: ligand / ID: 5 / Number of copies: 1 / Formula: 144 |
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Molecular weight | Theoretical: 122.143 Da |
Chemical component information | ChemComp-144: |
-Macromolecule #6: CHOLESTEROL
Macromolecule | Name: CHOLESTEROL / type: ligand / ID: 6 / Number of copies: 1 / Formula: CLR |
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Molecular weight | Theoretical: 386.654 Da |
Chemical component information | ChemComp-CLR: |
-Macromolecule #7: SODIUM ION
Macromolecule | Name: SODIUM ION / type: ligand / ID: 7 / Number of copies: 2 |
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Molecular weight | Theoretical: 22.99 Da |
-Macromolecule #8: CHLORIDE ION
Macromolecule | Name: CHLORIDE ION / type: ligand / ID: 8 / Number of copies: 1 / Formula: CL |
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Molecular weight | Theoretical: 35.453 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 2.3 mg/mL | |||||||||||||||
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Buffer | pH: 8 Component:
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Grid | Model: C-flat-1.2/1.3 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: OTHER | |||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 4294 / Average electron dose: 59.2 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: PDB ENTRY PDB model - PDB ID: |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 137600 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |