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Yorodumi- PDB-7bg4: Multidrug resistance transporter BmrA mutant E504A bound with ATP... -
+Open data
-Basic information
Entry | Database: PDB / ID: 7bg4 | ||||||||||||
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Title | Multidrug resistance transporter BmrA mutant E504A bound with ATP, Mg, and Rhodamine 6G solved by Cryo-EM | ||||||||||||
Components | Multidrug resistance ABC transporter ATP-binding/permease protein BmrA | ||||||||||||
Keywords | TRANSPORT PROTEIN / membrane protein / ABC transporter / multidrug resistance | ||||||||||||
Function / homology | Function and homology information ATPase-coupled lipid transmembrane transporter activity / Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate / ABC-type transporter activity / transmembrane transport / response to antibiotic / ATP hydrolysis activity / ATP binding / plasma membrane Similarity search - Function | ||||||||||||
Biological species | Bacillus subtilis (bacteria) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.2 Å | ||||||||||||
Authors | Wiseman, B. / Chaptal, V. / Zampieri, V. / Magnard, S. / Hogbom, M. / Falson, P. | ||||||||||||
Funding support | France, Sweden, 3items
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Citation | Journal: Sci Adv / Year: 2022 Title: Substrate-bound and substrate-free outward-facing structures of a multidrug ABC exporter. Authors: Vincent Chaptal / Veronica Zampieri / Benjamin Wiseman / Cédric Orelle / Juliette Martin / Kim-Anh Nguyen / Alexia Gobet / Margot Di Cesare / Sandrine Magnard / Waqas Javed / Jad Eid / ...Authors: Vincent Chaptal / Veronica Zampieri / Benjamin Wiseman / Cédric Orelle / Juliette Martin / Kim-Anh Nguyen / Alexia Gobet / Margot Di Cesare / Sandrine Magnard / Waqas Javed / Jad Eid / Arnaud Kilburg / Marine Peuchmaur / Julien Marcoux / Luca Monticelli / Martin Hogbom / Guy Schoehn / Jean-Michel Jault / Ahcène Boumendjel / Pierre Falson / Abstract: Multidrug ABC transporters translocate drugs across membranes by a mechanism for which the molecular features of drug release are so far unknown. Here, we resolved three ATP-Mg-bound outward-facing ...Multidrug ABC transporters translocate drugs across membranes by a mechanism for which the molecular features of drug release are so far unknown. Here, we resolved three ATP-Mg-bound outward-facing conformations of the (homodimeric) BmrA by x-ray crystallography and single-particle cryo-electron microscopy (EM) in detergent solution, one of them with rhodamine 6G (R6G), a substrate exported by BmrA when overexpressed in . Two R6G molecules bind to the drug-binding cavity at the level of the outer leaflet, between transmembrane (TM) helices 1-2 of one monomer and TM5'-6' of the other. They induce a rearrangement of TM1-2, highlighting a local flexibility that we confirmed by hydrogen/deuterium exchange and molecular dynamics simulations. In the absence of R6G, simulations show a fast postrelease occlusion of the cavity driven by hydrophobicity, while when present, R6G can move within the cavity, maintaining it open. | ||||||||||||
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 | 7bg4.cif.gz | 208 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7bg4.ent.gz | 165.1 KB | Display | PDB format |
PDBx/mmJSON format | 7bg4.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7bg4_validation.pdf.gz | 993.1 KB | Display | wwPDB validaton report |
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Full document | 7bg4_full_validation.pdf.gz | 1017.7 KB | Display | |
Data in XML | 7bg4_validation.xml.gz | 42.8 KB | Display | |
Data in CIF | 7bg4_validation.cif.gz | 62.8 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/bg/7bg4 ftp://data.pdbj.org/pub/pdb/validation_reports/bg/7bg4 | HTTPS FTP |
-Related structure data
Related structure data | 12170MC 4749C 6r72C 6r81C 7ow8C C: citing same article (ref.) M: map data used to model this data |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
#1: Protein | Mass: 65747.141 Da / Num. of mol.: 2 / Mutation: E504A Source method: isolated from a genetically manipulated source Source: (gene. exp.) Bacillus subtilis (bacteria) / Gene: bmrA, yvcC, BSU34820 / Plasmid: pET15b / Production host: Escherichia coli (E. coli) / Variant (production host): C43 References: UniProt: O06967, Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate #2: Chemical | #3: Chemical | #4: Chemical | Has ligand of interest | N | |
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-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: Multidrug resistance transporter from Bacillus subtilis bound with ATP, Mg, and Rhodamine 6G. Type: COMPLEX / Details: mutant E504A / Entity ID: #1 / Source: RECOMBINANT |
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Molecular weight | Units: MEGADALTONS / Experimental value: NO |
Source (natural) | Organism: Bacillus subtilis (bacteria) |
Source (recombinant) | Organism: Escherichia coli (E. coli) / Strain: C43, delta-AcrB / Plasmid: pET15b |
Buffer solution | pH: 8 |
Specimen | Conc.: 3.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: C-flat-1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-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 / Cs: 2.7 mm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 5 sec. / Electron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 3477 |
Image scans | Movie frames/image: 40 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 486404 | ||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 128372 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 187 / Protocol: FLEXIBLE FIT / Space: REAL |