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Yorodumi- PDB-6x01: Crystal structure of the GltPh V216C-A391C mutant cross-linked in... -
+Open data
-Basic information
Entry | Database: PDB / ID: 6x01 | ||||||
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Title | Crystal structure of the GltPh V216C-A391C mutant cross-linked in outward-facing state | ||||||
Components | Glutamate transporter homolog | ||||||
Keywords | TRANSPORT PROTEIN / Glutamate transporter homolog | ||||||
Function / homology | Function and homology information amino acid:sodium symporter activity / L-aspartate transmembrane transport / L-aspartate transmembrane transporter activity / L-aspartate import across plasma membrane / chloride transmembrane transporter activity / protein homotrimerization / chloride transmembrane transport / identical protein binding / metal ion binding / plasma membrane Similarity search - Function | ||||||
Biological species | Pyrococcus horikoshii (archaea) | ||||||
Method | X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.65 Å | ||||||
Authors | Chen, I. / Font, J. / Ryan, R. | ||||||
Funding support | Australia, 1items
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Citation | Journal: Nature / Year: 2021 Title: Glutamate transporters have a chloride channel with two hydrophobic gates. Authors: Ichia Chen / Shashank Pant / Qianyi Wu / Rosemary J Cater / Meghna Sobti / Robert J Vandenberg / Alastair G Stewart / Emad Tajkhorshid / Josep Font / Renae M Ryan / Abstract: Glutamate is the most abundant excitatory neurotransmitter in the central nervous system, and its precise control is vital to maintain normal brain function and to prevent excitotoxicity. The removal ...Glutamate is the most abundant excitatory neurotransmitter in the central nervous system, and its precise control is vital to maintain normal brain function and to prevent excitotoxicity. The removal of extracellular glutamate is achieved by plasma-membrane-bound transporters, which couple glutamate transport to sodium, potassium and pH gradients using an elevator mechanism. Glutamate transporters also conduct chloride ions by means of a channel-like process that is thermodynamically uncoupled from transport. However, the molecular mechanisms that enable these dual-function transporters to carry out two seemingly contradictory roles are unknown. Here we report the cryo-electron microscopy structure of a glutamate transporter homologue in an open-channel state, which reveals an aqueous cavity that is formed during the glutamate transport cycle. The functional properties of this cavity, combined with molecular dynamics simulations, reveal it to be an aqueous-accessible chloride permeation pathway that is gated by two hydrophobic regions and is conserved across mammalian and archaeal glutamate transporters. Our findings provide insight into the mechanism by which glutamate transporters support their dual function, and add information that will assist in mapping the complete transport cycle shared by the solute carrier 1A transporter family. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 6x01.cif.gz | 228.9 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6x01.ent.gz | 184 KB | Display | PDB format |
PDBx/mmJSON format | 6x01.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6x01_validation.pdf.gz | 471.8 KB | Display | wwPDB validaton report |
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Full document | 6x01_full_validation.pdf.gz | 493.6 KB | Display | |
Data in XML | 6x01_validation.xml.gz | 42.1 KB | Display | |
Data in CIF | 6x01_validation.cif.gz | 56.3 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/x0/6x01 ftp://data.pdbj.org/pub/pdb/validation_reports/x0/6x01 | HTTPS FTP |
-Related structure data
Related structure data | 6wyjC 6wykC 6wylC 6wzbC 2nwxS S: Starting model for refinement C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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Unit cell |
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Noncrystallographic symmetry (NCS) | NCS domain:
NCS domain segments: Component-ID: 1 / Ens-ID: 1 / Beg auth comp-ID: VAL / Beg label comp-ID: VAL / End auth comp-ID: GLU / End label comp-ID: GLU / Auth seq-ID: 12 - 416 / Label seq-ID: 12 - 416
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-Components
#1: Protein | Mass: 44585.035 Da / Num. of mol.: 3 / Mutation: V216C, A391C, C321S Source method: isolated from a genetically manipulated source Source: (gene. exp.) Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3) (archaea) Strain: ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3 Gene: PH1295 / Production host: Escherichia coli (E. coli) / References: UniProt: O59010 #2: Chemical | #3: Chemical | ChemComp-NA / Has ligand of interest | N | |
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-Experimental details
-Experiment
Experiment | Method: X-RAY DIFFRACTION / Number of used crystals: 1 |
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-Sample preparation
Crystal | Density Matthews: 4.51 Å3/Da / Density % sol: 72.73 % |
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Crystal grow | Temperature: 277.15 K / Method: vapor diffusion, hanging drop Details: 15/23% PEG 1000, 100mM Lithium sulfate, 50mM citric acid, 50mM Disodium phosphate |
-Data collection
Diffraction | Mean temperature: 100 K / Serial crystal experiment: N |
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Diffraction source | Source: SYNCHROTRON / Site: Australian Synchrotron / Beamline: MX2 / Wavelength: 0.9537 Å |
Detector | Type: DECTRIS EIGER X 16M / Detector: PIXEL / Date: May 6, 2017 |
Radiation | Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray |
Radiation wavelength | Wavelength: 0.9537 Å / Relative weight: 1 |
Reflection | Resolution: 3.65→48.79 Å / Num. obs: 26118 / % possible obs: 100 % / Redundancy: 21.6 % / CC1/2: 0.999 / Rmerge(I) obs: 0.128 / Net I/σ(I): 14 |
Reflection shell | Resolution: 3.65→3.9 Å / Rmerge(I) obs: 3.559 / Mean I/σ(I) obs: 1.2 / Num. unique obs: 4731 / CC1/2: 0.851 |
-Processing
Software |
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Refinement | Method to determine structure: MOLECULAR REPLACEMENT Starting model: 2NWX Resolution: 3.65→44.84 Å / SU ML: 0.59 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 45.85 / Stereochemistry target values: ML
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Solvent computation | Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso max: 280.65 Å2 / Biso mean: 176.7668 Å2 / Biso min: 76.74 Å2 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement step | Cycle: final / Resolution: 3.65→44.84 Å
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Refine LS restraints NCS |
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LS refinement shell | Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 9
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