+Open data
-Basic information
Entry | Database: PDB / ID: 7oji | ||||||
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Title | ABCG2 topotecan turnover-2 state | ||||||
Components | Broad substrate specificity ATP-binding cassette transporter ABCG2 | ||||||
Keywords | TRANSPORT PROTEIN / ABC transporter plasma membrane ATP multidrug resistance | ||||||
Function / homology | Function and homology information biotin transmembrane transporter activity / biotin transport / riboflavin transport / riboflavin transmembrane transporter activity / renal urate salt excretion / urate metabolic process / urate transmembrane transporter activity / Abacavir transmembrane transport / organic anion transport / external side of apical plasma membrane ...biotin transmembrane transporter activity / biotin transport / riboflavin transport / riboflavin transmembrane transporter activity / renal urate salt excretion / urate metabolic process / urate transmembrane transporter activity / Abacavir transmembrane transport / organic anion transport / external side of apical plasma membrane / organic anion transmembrane transporter activity / xenobiotic transport across blood-brain barrier / export across plasma membrane / ABC-type xenobiotic transporter / Paracetamol ADME / Ciprofloxacin ADME / transepithelial transport / cellular detoxification / ABC-type xenobiotic transporter activity / NFE2L2 regulating MDR associated enzymes / Heme biosynthesis / Heme degradation / lipid transport / xenobiotic transmembrane transporter activity / efflux transmembrane transporter activity / transport across blood-brain barrier / ATPase-coupled transmembrane transporter activity / mitochondrial membrane / brush border membrane / Iron uptake and transport / transmembrane transport / membrane raft / apical plasma membrane / ATP hydrolysis activity / protein homodimerization activity / nucleoplasm / ATP binding / identical protein binding / plasma membrane Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||
Authors | Yu, Q. / Ni, D. / Kowal, J. / Manolaridis, I. / Jackson, S.M. / Stahlberg, H. / Locher, K.P. | ||||||
Funding support | Switzerland, 1items
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Citation | Journal: Nat Commun / Year: 2021 Title: Structures of ABCG2 under turnover conditions reveal a key step in the drug transport mechanism. Authors: Qin Yu / Dongchun Ni / Julia Kowal / Ioannis Manolaridis / Scott M Jackson / Henning Stahlberg / Kaspar P Locher / Abstract: ABCG2 is a multidrug transporter that affects drug pharmacokinetics and contributes to multidrug resistance of cancer cells. In previously reported structures, the reaction cycle was halted by the ...ABCG2 is a multidrug transporter that affects drug pharmacokinetics and contributes to multidrug resistance of cancer cells. In previously reported structures, the reaction cycle was halted by the absence of substrates or ATP, mutation of catalytic residues, or the presence of small-molecule inhibitors or inhibitory antibodies. Here we present cryo-EM structures of ABCG2 under turnover conditions containing either the endogenous substrate estrone-3-sulfate or the exogenous substrate topotecan. We find two distinct conformational states in which both the transport substrates and ATP are bound. Whereas the state turnover-1 features more widely separated NBDs and an accessible substrate cavity between the TMDs, turnover-2 features semi-closed NBDs and an almost fully occluded substrate cavity. Substrate size appears to control which turnover state is mainly populated. The conformational changes between turnover-1 and turnover-2 states reveal how ATP binding is linked to the closing of the cytoplasmic side of the TMDs. The transition from turnover-1 to turnover-2 is the likely bottleneck or rate-limiting step of the reaction cycle, where the discrimination of substrates and inhibitors occurs. | ||||||
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 | 7oji.cif.gz | 210.7 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7oji.ent.gz | 172.6 KB | Display | PDB format |
PDBx/mmJSON format | 7oji.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/oj/7oji ftp://data.pdbj.org/pub/pdb/validation_reports/oj/7oji | HTTPS FTP |
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-Related structure data
Related structure data | 12952MC 7oj8C 7ojhC 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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1 |
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-Components
#1: Protein | Mass: 73526.938 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: ABCG2, ABCP, BCRP, BCRP1, MXR / Cell (production host): HEK293-EBNA / Production host: Homo sapiens (human) References: UniProt: Q9UNQ0, ABC-type xenobiotic transporter #2: Chemical | ChemComp-TTC / ( | #3: Chemical | ChemComp-CLR / #4: Chemical | #5: Chemical | Has ligand of interest | Y | |
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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: ABCG2 in complex with topotecan under turnover condition Type: COMPLEX Details: ABCG2 was incubated with 5mM ATP, 5mM MgCl2, 0.5mM ADP, 100 uM topotecan at room temperature for 10 min Entity ID: #1 / Source: RECOMBINANT | |||||||||||||||
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Molecular weight | Value: 0.144 MDa / Experimental value: YES | |||||||||||||||
Source (natural) | Organism: Homo sapiens (human) | |||||||||||||||
Source (recombinant) | Organism: Homo sapiens (human) / Cell: HEK293EBNA | |||||||||||||||
Buffer solution | pH: 7.5 | |||||||||||||||
Buffer component |
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Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: The sample was mono-disperse. | |||||||||||||||
Specimen support | Details: 25mA / Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 | |||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277 K / Details: Grids were blotted for 2.5s with blot force 1 |
-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: DARK FIELD / Nominal magnification: 130000 X / Cs: 2.7 mm / C2 aperture diameter: 100 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 58 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
EM imaging optics | Energyfilter name: GIF Bioquantum / Details: no phase plate / Energyfilter slit width: 20 eV |
-Processing
Software |
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 2623169 | ||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 78323 / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL / Target criteria: Correlation coefficient | ||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 6HCO | ||||||||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 64.59 Å2 | ||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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