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Yorodumi- EMDB-12524: Cryo-EM structure of Human excitatory amino acid transporters-1 (... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-12524 | |||||||||
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Title | Cryo-EM structure of Human excitatory amino acid transporters-1 (EAAT1) in potassium buffer | |||||||||
Map data | Cryo-EM map of Human excitatory amino acid transporters (EAATs) | |||||||||
Sample |
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Keywords | Human Membrane Protein / Transporter / Glutamate transporter / MEMBRANE PROTEIN | |||||||||
Function / homology | Function and homology information Defective SLC1A3 causes episodic ataxia 6 (EA6) / Astrocytic Glutamate-Glutamine Uptake And Metabolism / membrane protein complex / auditory behavior / neurotransmitter uptake / cranial nerve development / cell morphogenesis involved in neuron differentiation / gamma-aminobutyric acid biosynthetic process / high-affinity L-glutamate transmembrane transporter activity / glutamate:sodium symporter activity ...Defective SLC1A3 causes episodic ataxia 6 (EA6) / Astrocytic Glutamate-Glutamine Uptake And Metabolism / membrane protein complex / auditory behavior / neurotransmitter uptake / cranial nerve development / cell morphogenesis involved in neuron differentiation / gamma-aminobutyric acid biosynthetic process / high-affinity L-glutamate transmembrane transporter activity / glutamate:sodium symporter activity / L-glutamate import / Transport of inorganic cations/anions and amino acids/oligopeptides / L-glutamate transmembrane transport / L-glutamate transmembrane transporter activity / D-aspartate import across plasma membrane / L-aspartate import across plasma membrane / Glutamate Neurotransmitter Release Cycle / neutral L-amino acid transmembrane transporter activity / L-glutamate import across plasma membrane / transepithelial transport / intracellular sodium ion homeostasis / cellular response to cocaine / neurotransmitter transport / glutamate binding / transport across blood-brain barrier / neuromuscular process controlling balance / response to light stimulus / positive regulation of synaptic transmission / monoatomic ion transport / chloride transmembrane transport / potassium ion transmembrane transport / basal plasma membrane / sensory perception of sound / response to wounding / cytoplasmic vesicle / chemical synaptic transmission / neuron projection / response to xenobiotic stimulus / response to antibiotic / neuronal cell body / synapse / perinuclear region of cytoplasm / cell surface / membrane / metal ion binding / plasma membrane Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.99 Å | |||||||||
Authors | Kumar A / Reyes N | |||||||||
Funding support | France, 1 items
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Citation | Journal: EMBO J / Year: 2022 Title: The ion-coupling mechanism of human excitatory amino acid transporters. Authors: Juan C Canul-Tec / Anand Kumar / Jonathan Dhenin / Reda Assal / Pierre Legrand / Martial Rey / Julia Chamot-Rooke / Nicolas Reyes / Abstract: Excitatory amino acid transporters (EAATs) maintain glutamate gradients in the brain essential for neurotransmission and to prevent neuronal death. They use ionic gradients as energy source and co- ...Excitatory amino acid transporters (EAATs) maintain glutamate gradients in the brain essential for neurotransmission and to prevent neuronal death. They use ionic gradients as energy source and co-transport transmitter into the cytoplasm with Na and H , while counter-transporting K to re-initiate the transport cycle. However, the molecular mechanisms underlying ion-coupled transport remain incompletely understood. Here, we present 3D X-ray crystallographic and cryo-EM structures, as well as thermodynamic analysis of human EAAT1 in different ion bound conformations, including elusive counter-transport ion bound states. Binding energies of Na and H , and unexpectedly Ca , are coupled to neurotransmitter binding. Ca competes for a conserved Na site, suggesting a regulatory role for Ca in glutamate transport at the synapse, while H binds to a conserved glutamate residue stabilizing substrate occlusion. The counter-transported ion binding site overlaps with that of glutamate, revealing the K -based mechanism to exclude the transmitter during the transport cycle and to prevent its neurotoxic release on the extracellular side. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_12524.map.gz | 230 MB | EMDB map data format | |
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Header (meta data) | emd-12524-v30.xml emd-12524.xml | 14.3 KB 14.3 KB | Display Display | EMDB header |
Images | emd_12524.png | 133 KB | ||
Filedesc metadata | emd-12524.cif.gz | 6.1 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-12524 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-12524 | HTTPS FTP |
-Validation report
Summary document | emd_12524_validation.pdf.gz | 483.2 KB | Display | EMDB validaton report |
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Full document | emd_12524_full_validation.pdf.gz | 482.8 KB | Display | |
Data in XML | emd_12524_validation.xml.gz | 7.2 KB | Display | |
Data in CIF | emd_12524_validation.cif.gz | 8.2 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-12524 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-12524 | HTTPS FTP |
-Related structure data
Related structure data | 7npwMC 7awlC 7awmC 7awnC 7awpC 7awqC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_12524.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Cryo-EM map of Human excitatory amino acid transporters (EAATs) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.814 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Human excitatory amino acid transporter 1
Entire | Name: Human excitatory amino acid transporter 1 |
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Components |
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-Supramolecule #1: Human excitatory amino acid transporter 1
Supramolecule | Name: Human excitatory amino acid transporter 1 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Homo sapiens (human) |
-Macromolecule #1: Excitatory amino acid transporter 1
Macromolecule | Name: Excitatory amino acid transporter 1 / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 51.078273 KDa |
Recombinant expression | Organism: Homo sapiens (human) |
Sequence | String: AKKKVQNITK EDVKSYLFRN AFVLLTVTAV IVGTILGFTL RPYRMSYREV KYFSFPGELL MRMLQMLVLP LIISSLVTGM AALDSKASG KMGMRAVVYY MTTTIIAVVI GIIIVIIIHP GKGTKENMHR EGKIVRVTAA DAFLDLIRNM FPPNLVEACF K QFKTNYEK ...String: AKKKVQNITK EDVKSYLFRN AFVLLTVTAV IVGTILGFTL RPYRMSYREV KYFSFPGELL MRMLQMLVLP LIISSLVTGM AALDSKASG KMGMRAVVYY MTTTIIAVVI GIIIVIIIHP GKGTKENMHR EGKIVRVTAA DAFLDLIRNM FPPNLVEACF K QFKTNYEK RSFKVPIQAN ETLVGAVINN VSEAMETLTR ITEELVPVPG SVNGVNALGL VVFSMCFGFV IGNMKEQGQA LR EFFDSLN EAIMRLVAVI MWYAPVGILF LIAGKIVEME DMGVIGGQLA MYTVTVIVGL LIHAVIVLPL LYFLVTRKNP WVF IGGLLQ ALITALGTSS SSATLPITFK CLEENNGVDK RVTRFVLPVG ATINMDGTAL YEALAAIFIA QVNNFELNFG QIIT ISITA TAASIGAAGI PQAGLVTMVI VLTSVGLPTD DITLIIAVDW FLDRLRTTTN VLGDSLGAGI VEHL UniProtKB: Excitatory amino acid transporter 1 |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 4 mg/mL | ||||||||||||||||||
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Buffer | pH: 7.4 Component:
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Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR | ||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV / Details: Grids were blotted for 4 Sec. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-40 / Number grids imaged: 1 / Number real images: 13607 / Average electron dose: 42.3 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / 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 | Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Details | For model building, the scaD and tranD of Rb+/Ba2+ bound X-ray structure were fitted into the EM map as separate rigid-bodies using Chimera. |
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Refinement | Space: REAL / Protocol: RIGID BODY FIT |
Output model | PDB-7npw: |