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Yorodumi- PDB-7nf8: Ovine (b0,+AT-rBAT)2 hetero-tetramer, asymmetric unit, rigid-body... -
+Open data
-Basic information
Entry | Database: PDB / ID: 7nf8 | |||||||||
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Title | Ovine (b0,+AT-rBAT)2 hetero-tetramer, asymmetric unit, rigid-body fitted | |||||||||
Components |
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Keywords | TRANSLOCASE / Transporter / Cystinuria / Complex | |||||||||
Function / homology | Function and homology information broad specificity neutral L-amino acid:basic L-amino acid antiporter activity / L-cystine transmembrane transporter activity / brush border membrane / carbohydrate metabolic process / protein heterodimerization activity / apical plasma membrane Similarity search - Function | |||||||||
Biological species | Ovis aries (sheep) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.83 Å | |||||||||
Authors | Lee, Y. / Kuehlbrandt, W. | |||||||||
Funding support | Germany, 2items
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Citation | Journal: Nat Commun / Year: 2022 Title: Ca-mediated higher-order assembly of heterodimers in amino acid transport system b biogenesis and cystinuria. Authors: Yongchan Lee / Pattama Wiriyasermkul / Pornparn Kongpracha / Satomi Moriyama / Deryck J Mills / Werner Kühlbrandt / Shushi Nagamori / Abstract: Cystinuria is a genetic disorder characterized by overexcretion of dibasic amino acids and cystine, causing recurrent kidney stones and kidney failure. Mutations of the regulatory glycoprotein rBAT ...Cystinuria is a genetic disorder characterized by overexcretion of dibasic amino acids and cystine, causing recurrent kidney stones and kidney failure. Mutations of the regulatory glycoprotein rBAT and the amino acid transporter bAT, which constitute system b, are linked to type I and non-type I cystinuria respectively and they exhibit distinct phenotypes due to protein trafficking defects or catalytic inactivation. Here, using electron cryo-microscopy and biochemistry, we discover that Ca mediates higher-order assembly of system b. Ca stabilizes the interface between two rBAT molecules, leading to super-dimerization of bAT-rBAT, which in turn facilitates N-glycan maturation and protein trafficking. A cystinuria mutant T216M and mutations of the Ca site of rBAT cause the loss of higher-order assemblies, resulting in protein trapping at the ER and the loss of function. These results provide the molecular basis of system b biogenesis and type I cystinuria and serve as a guide to develop new therapeutic strategies against it. More broadly, our findings reveal an unprecedented link between transporter oligomeric assembly and protein-trafficking diseases. #1: Journal: Biorxiv / Year: 2021 Title: Ca 2+ -mediated higher-order assembly of b 0,+ AT-rBAT is a key step for system b 0,+ biogenesis and cystinuria Authors: Lee, Y. / Wiriyasermkul, P. / Moriyama, S. / Mills, D.J. / Kuhlbrandt, W. / Nagamori, S. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7nf8.cif.gz | 212.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7nf8.ent.gz | 164 KB | Display | PDB format |
PDBx/mmJSON format | 7nf8.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/nf/7nf8 ftp://data.pdbj.org/pub/pdb/validation_reports/nf/7nf8 | HTTPS FTP |
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-Related structure data
Related structure data | 12298MC 7nf6C 7nf7C M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Protein , 2 types, 2 molecules BA
#1: Protein | Mass: 56318.047 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Ovis aries (sheep) / Gene: SLC7A9 / Production host: Homo sapiens (human) / References: UniProt: A0A6P3EL78 |
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#2: Protein | Mass: 78754.281 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Ovis aries (sheep) / Gene: SLC3A1 / Production host: Homo sapiens (human) / References: UniProt: A0A6P7DVK7 |
-Sugars , 2 types, 6 molecules
#3: Polysaccharide | Source method: isolated from a genetically manipulated source #7: Sugar | ChemComp-NAG / |
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-Non-polymers , 3 types, 4 molecules
#4: Chemical | #5: Chemical | ChemComp-LBN / | #6: Chemical | ChemComp-CA / | |
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-Details
Has ligand of interest | Y |
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Has protein modification | Y |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: (b0,+AT-rBAT) hetero-tetramer / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT |
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Molecular weight | Value: 270 kDa/nm / Experimental value: NO |
Source (natural) | Organism: Ovis aries (sheep) |
Source (recombinant) | Organism: Homo sapiens (human) |
Buffer solution | pH: 8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
-Processing
Software | Name: PHENIX / Version: dev_3689: / Classification: refinement | ||||||||||||||||||||||||
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Symmetry | Point symmetry: C2 (2 fold cyclic) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 2.83 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 581697 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT | ||||||||||||||||||||||||
Refine LS restraints |
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