+Open data
-Basic information
Entry | Database: PDB / ID: 8wzi | |||||||||
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Title | One RBD up state of Spike glycoprotein, SARS-CoV-2 | |||||||||
Components | Spike glycoprotein | |||||||||
Keywords | VIRAL PROTEIN / Glycoprotein / SARS-CoV-2 | |||||||||
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 / receptor-mediated endocytosis of virus by host cell / membrane fusion / 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 | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å | |||||||||
Authors | Yadav, S. / Vinothkumar, K.R. | |||||||||
Funding support | India, 2items
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Citation | Journal: Acta Crystallogr D Struct Biol / Year: 2024 Title: Factors affecting macromolecule orientations in thin films formed in cryo-EM. Authors: Swati Yadav / Kutti R Vinothkumar / Abstract: The formation of a vitrified thin film embedded with randomly oriented macromolecules is an essential prerequisite for cryogenic sample electron microscopy. Most commonly, this is achieved using the ...The formation of a vitrified thin film embedded with randomly oriented macromolecules is an essential prerequisite for cryogenic sample electron microscopy. Most commonly, this is achieved using the plunge-freeze method first described nearly 40 years ago. Although this is a robust method, the behaviour of different macromolecules shows great variation upon freezing and often needs to be optimized to obtain an isotropic, high-resolution reconstruction. For a macromolecule in such a film, the probability of encountering the air-water interface in the time between blotting and freezing and adopting preferred orientations is very high. 3D reconstruction using preferentially oriented particles often leads to anisotropic and uninterpretable maps. Currently, there are no general solutions to this prevalent issue, but several approaches largely focusing on sample preparation with the use of additives and novel grid modifications have been attempted. In this study, the effect of physical and chemical factors on the orientations of macromolecules was investigated through an analysis of selected well studied macromolecules, and important parameters that determine the behaviour of proteins on cryo-EM grids were revealed. These insights highlight the nature of the interactions that cause preferred orientations and can be utilized to systematically address orientation bias for any given macromolecule and to provide a framework to design small-molecule additives to enhance sample stability and behaviour. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8wzi.cif.gz | 629.1 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8wzi.ent.gz | 507.4 KB | Display | PDB format |
PDBx/mmJSON format | 8wzi.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8wzi_validation.pdf.gz | 2.8 MB | Display | wwPDB validaton report |
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Full document | 8wzi_full_validation.pdf.gz | 2.8 MB | Display | |
Data in XML | 8wzi_validation.xml.gz | 92.3 KB | Display | |
Data in CIF | 8wzi_validation.cif.gz | 141 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/wz/8wzi ftp://data.pdbj.org/pub/pdb/validation_reports/wz/8wzi | HTTPS FTP |
-Related structure data
Related structure data | 37953MC 8wv4C 8wv5C 8wv6C 8wzhC 8wzjC 8wzkC 8wzmC M: map data used to model this data 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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-Components
#1: Protein | Mass: 139706.984 Da / Num. of mol.: 3 / Mutation: (RRAR)682A,K986P, V987P Source method: isolated from a genetically manipulated source Details: linker (1216-SGRLVPRGSP-1225) and a fibritin trimerization motif (1226-GSGYIPEAPRDGQAYVRKDGEWVLLSTFLG-1254) followed by another linker (1256-GTK-1258) followed by the HRV3c post cleavage ...Details: linker (1216-SGRLVPRGSP-1225) and a fibritin trimerization motif (1226-GSGYIPEAPRDGQAYVRKDGEWVLLSTFLG-1254) followed by another linker (1256-GTK-1258) followed by the HRV3c post cleavage fragment (1259-LEVLFQ-1264) Source: (gene. exp.) Severe acute respiratory syndrome coronavirus 2 Gene: S / Production host: Homo sapiens (human) / References: UniProt: P0DTC2 #2: Polysaccharide | 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source #3: Polysaccharide | 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta- ...2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta-D-glucopyranose | Source method: isolated from a genetically manipulated source #4: Polysaccharide | alpha-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...alpha-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source #5: Sugar | ChemComp-NAG / 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: Spike glycoprotein, SARS-CoV-2 / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Value: 0.432 MDa / Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: Severe acute respiratory syndrome coronavirus 2 | ||||||||||||||||||||
Source (recombinant) | Organism: Homo sapiens (human) | ||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 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 R0.6/1 | ||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 293 K / Details: Blot force, 0 |
-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: 75000 X / Calibrated magnification: 130841 X / Nominal defocus max: 3000 nm / Nominal defocus min: 1800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 60 sec. / Electron dose: 25 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Details: Movies were collected at 40 frames per second |
Image scans | Sampling size: 14 µm / Width: 4096 / Height: 4096 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 261703 / Algorithm: BACK PROJECTION / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 84.6 / Protocol: OTHER / Space: RECIPROCAL | ||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 8d56 Accession code: 8d56 / Source name: PDB / Type: experimental model | ||||||||||||||||||||||||||||||||||||||||||||
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