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Yorodumi- PDB-8elp: Crystal structure of SARS-CoV-2 spike protein receptor-binding do... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8elp | ||||||
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Title | Crystal structure of SARS-CoV-2 spike protein receptor-binding domain in complex with antibody CC12.1 Fab and nanobody Nb-C4-240 | ||||||
Components |
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Keywords | VIRAL PROTEIN/IMMUNE SYSTEM / synthetic / nanobody / SARS-CoV-2 / coronavirus / neutralization / IMMUNE SYSTEM / VIRAL PROTEIN-IMMUNE SYSTEM complex | ||||||
Function / homology | Function and homology information Maturation of spike protein / viral translation / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / suppression by virus of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / entry receptor-mediated virion attachment to host cell ...Maturation of spike protein / viral translation / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / suppression by virus of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / entry receptor-mediated virion attachment to host cell / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / membrane fusion / receptor-mediated endocytosis of virus by host cell / Attachment and Entry / positive regulation of viral entry into host cell / receptor-mediated virion attachment to host cell / receptor ligand activity / symbiont-mediated suppression of host innate immune response / host cell surface receptor binding / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / SARS-CoV-2 activates/modulates innate and adaptive immune responses / host cell plasma membrane / virion membrane / identical protein binding / membrane / plasma membrane Similarity search - Function | ||||||
Biological species | Severe acute respiratory syndrome coronavirus 2 synthetic construct (others) Homo sapiens (human) | ||||||
Method | X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.83 Å | ||||||
Authors | Liu, H. / Wilson, I.A. | ||||||
Funding support | United States, 1items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2023 Title: Fully synthetic platform to rapidly generate tetravalent bispecific nanobody-based immunoglobulins. Authors: Laetitia Misson Mindrebo / Hejun Liu / Gabriel Ozorowski / Quoc Tran / Jordan Woehl / Irene Khalek / Jessica M Smith / Shawn Barman / Fangzhu Zhao / Celina Keating / Oliver Limbo / Megan ...Authors: Laetitia Misson Mindrebo / Hejun Liu / Gabriel Ozorowski / Quoc Tran / Jordan Woehl / Irene Khalek / Jessica M Smith / Shawn Barman / Fangzhu Zhao / Celina Keating / Oliver Limbo / Megan Verma / Jingjia Liu / Robyn L Stanfield / Xueyong Zhu / Hannah L Turner / Devin Sok / Po-Ssu Huang / Dennis R Burton / Andrew B Ward / Ian A Wilson / Joseph G Jardine / Abstract: Nanobodies bind a target antigen with a kinetic profile similar to a conventional antibody, but exist as a single heavy chain domain that can be readily multimerized to engage antigen via multiple ...Nanobodies bind a target antigen with a kinetic profile similar to a conventional antibody, but exist as a single heavy chain domain that can be readily multimerized to engage antigen via multiple interactions. Presently, most nanobodies are produced by immunizing camelids; however, platforms for animal-free production are growing in popularity. Here, we describe the development of a fully synthetic nanobody library based on an engineered human V3-23 variable gene and a multispecific antibody-like format designed for biparatopic target engagement. To validate our library, we selected nanobodies against the SARS-CoV-2 receptor-binding domain and employed an on-yeast epitope binning strategy to rapidly map the specificities of the selected nanobodies. We then generated antibody-like molecules by replacing the V and V domains of a conventional antibody with two different nanobodies, designed as a molecular clamp to engage the receptor-binding domain biparatopically. The resulting bispecific tetra-nanobody immunoglobulins neutralized diverse SARS-CoV-2 variants with potencies similar to antibodies isolated from convalescent donors. Subsequent biochemical analyses confirmed the accuracy of the on-yeast epitope binning and structures of both individual nanobodies, and a tetra-nanobody immunoglobulin revealed that the intended mode of interaction had been achieved. This overall workflow is applicable to nearly any protein target and provides a blueprint for a modular workflow for the development of multispecific molecules. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8elp.cif.gz | 297.8 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8elp.ent.gz | 237.4 KB | Display | PDB format |
PDBx/mmJSON format | 8elp.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8elp_validation.pdf.gz | 473 KB | Display | wwPDB validaton report |
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Full document | 8elp_full_validation.pdf.gz | 484.3 KB | Display | |
Data in XML | 8elp_validation.xml.gz | 26.5 KB | Display | |
Data in CIF | 8elp_validation.cif.gz | 35.8 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/el/8elp ftp://data.pdbj.org/pub/pdb/validation_reports/el/8elp | HTTPS FTP |
-Related structure data
Related structure data | 8dt8C 8eloC 8elqC 6xc3S 7kn5S S: Starting model for refinement C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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Unit cell |
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-Components
#1: Protein | Mass: 23104.867 Da / Num. of mol.: 1 / Fragment: Receptor binding domain, UNP residues 333-530 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Severe acute respiratory syndrome coronavirus 2 Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P0DTC2 |
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#2: Antibody | Mass: 15645.075 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) synthetic construct (others) / Production host: Homo sapiens (human) |
#3: Antibody | Mass: 23178.928 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Production host: Mus musculus (house mouse) |
#4: Antibody | Mass: 23641.365 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Production host: Mus musculus (house mouse) |
#5: Sugar | ChemComp-NAG / |
Has ligand of interest | N |
-Experimental details
-Experiment
Experiment | Method: X-RAY DIFFRACTION / Number of used crystals: 1 |
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-Sample preparation
Crystal | Density Matthews: 2.83 Å3/Da / Density % sol: 56.46 % |
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Crystal grow | Temperature: 293 K / Method: vapor diffusion, sitting drop / pH: 4.6 Details: 0.16 M ammonium sulfate, 0.08 M sodium acetate pH 4.6, 20% (w/v) polyethylene glycol 4000, 20% (v/v) glycerol |
-Data collection
Diffraction | Mean temperature: 100 K / Serial crystal experiment: N | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Diffraction source | Source: SYNCHROTRON / Site: SSRL / Beamline: BL12-2 / Wavelength: 0.97946 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Detector | Type: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Nov 5, 2021 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Radiation | Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Radiation wavelength | Wavelength: 0.97946 Å / Relative weight: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Reflection | Resolution: 2.83→50 Å / Num. obs: 42588 / % possible obs: 96.8 % / Redundancy: 2.1 % / Rmerge(I) obs: 0.955 / Χ2: 0.141 / Net I/σ(I): 4.7 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Reflection shell |
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-Processing
Software |
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Refinement | Method to determine structure: MOLECULAR REPLACEMENT Starting model: 7KN5, 6XC3 Resolution: 2.83→43.15 Å / SU ML: 0.41 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 27.75 / Stereochemistry target values: ML
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Solvent computation | Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement step | Cycle: LAST / Resolution: 2.83→43.15 Å
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Refine LS restraints |
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LS refinement shell |
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Refinement TLS params. | Method: refined / Origin x: 7.5947 Å / Origin y: 38.1285 Å / Origin z: 0.8468 Å
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Refinement TLS group | Selection details: all |