+Open data
-Basic information
Entry | Database: PDB / ID: 6y97 | ||||||
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Title | Structure of full-length CD20 in complex with Obinutuzumab Fab | ||||||
Components |
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Keywords | MEMBRANE PROTEIN / Cancer immunotherapy / Therapeutic antibody | ||||||
Function / homology | Function and homology information store-operated calcium entry / positive regulation of calcium ion import across plasma membrane / calcium ion import into cytosol / epidermal growth factor receptor binding / B cell proliferation / plasma membrane raft / B cell activation / immunoglobulin binding / humoral immune response / B cell differentiation ...store-operated calcium entry / positive regulation of calcium ion import across plasma membrane / calcium ion import into cytosol / epidermal growth factor receptor binding / B cell proliferation / plasma membrane raft / B cell activation / immunoglobulin binding / humoral immune response / B cell differentiation / response to bacterium / B cell receptor signaling pathway / protein tetramerization / MHC class II protein complex binding / cell surface receptor signaling pathway / external side of plasma membrane / cell surface / extracellular space / extracellular exosome / nucleoplasm / identical protein binding / plasma membrane Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.33 Å | ||||||
Authors | Kumar, A. / Reyes, N. | ||||||
Funding support | France, 1items
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Citation | Journal: Science / Year: 2020 Title: Binding mechanisms of therapeutic antibodies to human CD20. Authors: Anand Kumar / Cyril Planchais / Rémi Fronzes / Hugo Mouquet / Nicolas Reyes / Abstract: Monoclonal antibodies (mAbs) targeting human antigen CD20 (cluster of differentiation 20) constitute important immunotherapies for the treatment of B cell malignancies and autoimmune diseases. Type I ...Monoclonal antibodies (mAbs) targeting human antigen CD20 (cluster of differentiation 20) constitute important immunotherapies for the treatment of B cell malignancies and autoimmune diseases. Type I and II therapeutic mAbs differ in B cell binding properties and cytotoxic effects, reflecting differential interaction mechanisms with CD20. Here we present 3.7- to 4.7-angstrom cryo-electron microscopy structures of full-length CD20 in complexes with prototypical type I rituximab and ofatumumab and type II obinutuzumab. The structures and binding thermodynamics demonstrate that upon binding to CD20, type II mAbs form terminal complexes that preclude recruitment of additional mAbs and complement components, whereas type I complexes act as molecular seeds to increase mAb local concentration for efficient complement activation. Among type I mAbs, ofatumumab complexes display optimal geometry for complement recruitment. The uncovered mechanisms should aid rational design of next-generation immunotherapies targeting CD20. | ||||||
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 | 6y97.cif.gz | 108.7 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6y97.ent.gz | 81.8 KB | Display | PDB format |
PDBx/mmJSON format | 6y97.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6y97_validation.pdf.gz | 832.8 KB | Display | wwPDB validaton report |
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Full document | 6y97_full_validation.pdf.gz | 836.7 KB | Display | |
Data in XML | 6y97_validation.xml.gz | 23.1 KB | Display | |
Data in CIF | 6y97_validation.cif.gz | 32.5 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/y9/6y97 ftp://data.pdbj.org/pub/pdb/validation_reports/y9/6y97 | HTTPS FTP |
-Related structure data
Related structure data | 10733MC 6y90C 6y92C 6y9aC 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: 18735.373 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Details: Wt CD20 / Source: (gene. exp.) Homo sapiens (human) / Cell: B-Lymphocyte / Gene: MS4A1, CD20 / Plasmid: pcDNA3.1+ / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: P11836 #2: Antibody | | Mass: 12855.385 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Plasmid: IgG1 Expression vector / Cell line (production host): HEK293F / Production host: Homo sapiens (human) #3: Antibody | | Mass: 11962.462 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Plasmid: pcDNA3.1 + / Cell line (production host): HEK293F / Production host: Homo sapiens (human) Has protein modification | 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
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Molecular weight |
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Source (natural) |
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Source (recombinant) |
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Buffer solution | pH: 7.4 | |||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 1.2 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: Quantifoil R1.2/1.3 | |||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K / Details: Grids were blot for 3.5 Sec. |
-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 / Calibrated defocus min: 800 nm / Calibrated defocus max: 2000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm |
Specimen holder | Cryogen: NITROGEN |
Image recording | Average exposure time: 8 sec. / Electron dose: 42.84 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 10545 |
Image scans | Movie frames/image: 40 / Used frames/image: 1-40 |
-Processing
Software | Name: PHENIX / Version: 1.18.2_3874: / Classification: refinement | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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EM software |
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CTF correction | Details: Gctf / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1316678 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.33 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 65203 / Num. of class averages: 3 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building |
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Atomic model building | Accession code: 3PP4 / Initial refinement model-ID: 1 / PDB-ID: 3PP4 / Source name: PDB / Type: experimental model
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Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 112.8 Å2 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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