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- PDB-7m33: The structure of Bacillus subtilis BmrCD in the inward-facing con... -
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Open data
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Basic information
Entry | Database: PDB / ID: 7m33 | |||||||||
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Title | The structure of Bacillus subtilis BmrCD in the inward-facing conformation bound to Hoechst-33342 and ATP | |||||||||
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![]() | TRANSPORT PROTEIN / ABC transporter / multi-drug efflux transporter / ABC exporter | |||||||||
Function / homology | ![]() ATPase-coupled lipid transmembrane transporter activity / Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate / ATPase-coupled transmembrane transporter activity / ABC-type transporter activity / transmembrane transport / response to antibiotic / ATP hydrolysis activity / ATP binding / plasma membrane Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.55 Å | |||||||||
![]() | Thaker, T.M. / Tomasiak, T.M. | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Asymmetric drug binding in an ATP-loaded inward-facing state of an ABC transporter. Authors: Tarjani M Thaker / Smriti Mishra / Wenchang Zhou / Michael Mohan / Qingyu Tang / José D Faraldo-Goméz / Hassane S Mchaourab / Thomas M Tomasiak / ![]() Abstract: Substrate efflux by ATP-binding cassette (ABC) transporters, which play a major role in multidrug resistance, entails the ATP-powered interconversion between transporter intermediates. Despite recent ...Substrate efflux by ATP-binding cassette (ABC) transporters, which play a major role in multidrug resistance, entails the ATP-powered interconversion between transporter intermediates. Despite recent progress in structure elucidation, a number of intermediates have yet to be visualized and mechanistically interpreted. Here, we combine cryogenic-electron microscopy (cryo-EM), double electron-electron resonance spectroscopy and molecular dynamics simulations to profile a previously unobserved intermediate of BmrCD, a heterodimeric multidrug ABC exporter from Bacillus subtilis. In our cryo-EM structure, ATP-bound BmrCD adopts an inward-facing architecture featuring two molecules of the substrate Hoechst-33342 in a striking asymmetric head-to-tail arrangement. Deletion of the extracellular domain capping the substrate-binding chamber or mutation of Hoechst-coordinating residues abrogates cooperative stimulation of ATP hydrolysis. Together, our findings support a mechanistic role for symmetry mismatch between the nucleotide binding and the transmembrane domains in the conformational cycle of ABC transporters and is of notable importance for rational design of molecules for targeted ABC transporter inhibition. | |||||||||
History |
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Structure visualization
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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PDBx/mmCIF format | ![]() | 423 KB | Display | ![]() |
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PDB format | ![]() | 340.8 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Summary document | ![]() | 1.4 MB | Display | ![]() |
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Full document | ![]() | 1.4 MB | Display | |
Data in XML | ![]() | 48 KB | Display | |
Data in CIF | ![]() | 71.8 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 23641MC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 67602.961 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: yheI, BSU09710 / Production host: ![]() ![]() References: UniProt: O07550, Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate | ||||
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#2: Protein | Mass: 77369.898 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() Gene: yheH, BSU09720 / Production host: ![]() ![]() References: UniProt: O07549, Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate | ||||
#3: Chemical | #4: Chemical | Has ligand of interest | Y | |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: BmrCD / Type: COMPLEX Details: heterodimeric multi-drug ABC exporter from Bacillus subtilis Entity ID: #1-#2 / Source: RECOMBINANT |
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Molecular weight | Value: 0.14139865 MDa / Experimental value: NO |
Source (natural) | Organism: ![]() ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 7.4 |
Specimen | Conc.: 2.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: Purified by size exclusion chromatography using a Superose 6 column. |
Specimen support | Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 85 % / Chamber temperature: 283 K / Details: blot for 4 seconds before plunging |
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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: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2100 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 37.2 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 6919 |
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Processing
Software | Name: PHENIX / Version: 1.17.1_3660: / Classification: refinement | ||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.55 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 157021 / Symmetry type: POINT | ||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||
Displacement parameters | Biso mean: 59.49 Å2 | ||||||||||||||||||||||||
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