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Yorodumi- PDB-8bwp: Cryo-EM structure of nanodisc-reconstituted wildtype human MRP4 (... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8bwp | ||||||
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Title | Cryo-EM structure of nanodisc-reconstituted wildtype human MRP4 (in complex with methotrexate) | ||||||
Components | ATP-binding cassette sub-family C member 4 | ||||||
Keywords | TRANSLOCASE / ABC transporter / ABCC4 / MRP4 | ||||||
Function / homology | METHOTREXATE / : Function and homology information | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å | ||||||
Authors | Raj, I. / Bloch, M. / Pape, T.H. / Taylor, N.M.I. | ||||||
Funding support | Denmark, 1items
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Citation | Journal: Structure / Year: 2023 Title: Structural and mechanistic basis of substrate transport by the multidrug transporter MRP4. Authors: Magnus Bloch / Isha Raj / Tillmann Pape / Nicholas M I Taylor / Abstract: Multidrug resistance-associated protein 4 (MRP4) is an ATP-binding cassette (ABC) transporter expressed at multiple tissue barriers where it actively extrudes a wide variety of drug compounds. ...Multidrug resistance-associated protein 4 (MRP4) is an ATP-binding cassette (ABC) transporter expressed at multiple tissue barriers where it actively extrudes a wide variety of drug compounds. Overexpression of MRP4 provides resistance to clinically used antineoplastic agents, making it a highly attractive therapeutic target for countering multidrug resistance. Here, we report cryo-EM structures of multiple physiologically relevant states of lipid bilayer-embedded human MRP4, including complexes between MRP4 and two widely used chemotherapeutic agents and a complex between MRP4 and its native substrate. The structures display clear similarities and distinct differences in the coordination of these chemically diverse substrates and, in combination with functional and mutational analysis, reveal molecular details of the transport mechanism. Our study provides key insights into the unusually broad substrate specificity of MRP4 and constitutes an important contribution toward a general understanding of multidrug transporters. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8bwp.cif.gz | 223.3 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8bwp.ent.gz | 175.5 KB | Display | PDB format |
PDBx/mmJSON format | 8bwp.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8bwp_validation.pdf.gz | 1.1 MB | Display | wwPDB validaton report |
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Full document | 8bwp_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 8bwp_validation.xml.gz | 50.6 KB | Display | |
Data in CIF | 8bwp_validation.cif.gz | 75.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/bw/8bwp ftp://data.pdbj.org/pub/pdb/validation_reports/bw/8bwp | HTTPS FTP |
-Related structure data
Related structure data | 16293MC 8bjfC 8bwoC 8bwqC 8bwrC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 149693.922 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: ABCC4, MOATB, MRP4 / Cell line (production host): HEK / Production host: Homo sapiens (human) References: UniProt: O15439, Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate, ABC-type xenobiotic transporter, ABC-type ...References: UniProt: O15439, Translocases; Catalysing the translocation of other compounds; Linked to the hydrolysis of a nucleoside triphosphate, ABC-type xenobiotic transporter, ABC-type glutathione-S-conjugate transporter |
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#2: Chemical | ChemComp-MTX / |
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 |
-Sample preparation
Component | Name: Cryo-EM structure of nanodisc-reconstituted wildtype human MRP4 (in complex with methotrexate) Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1 / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Homo sapiens (human) / Cell: HEK / Plasmid: BacMam |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: This sample was monodisperse |
Specimen support | Grid type: Quantifoil R2/1 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / 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 / Nominal magnification: 96000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 500 nm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of real images: 5032 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1317394 | ||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 382621 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: AB INITIO MODEL | ||||||||||||||||||||||||||||||||||||||||||||
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