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- PDB-8qo5: Conserved Structures and Dynamics in 5-Proximal Regions of Betaco... -

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Basic information

Entry
Database: PDB / ID: 8qo5
TitleConserved Structures and Dynamics in 5-Proximal Regions of Betacoronavirus RNA Genomes
ComponentsSARS-CoV-2-SL5
KeywordsVIRUS / RNA structure / 5-proximal region / Coronavirus / Cryo-EM
Function / homology: / RNA / RNA (> 10) / RNA (> 100)
Function and homology information
Biological speciesSevere acute respiratory syndrome coronavirus 2
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 6.5 Å
AuthorsMoura, T.R. / Purta, E. / Bernat, A. / Baulin, E. / Mukherjee, S. / Bujnicki, J.M.
Funding support Poland, 1items
OrganizationGrant numberCountry
Polish National Science Centre Poland
CitationJournal: Nucleic Acids Res / Year: 2024
Title: Conserved structures and dynamics in 5'-proximal regions of Betacoronavirus RNA genomes.
Authors: Tales Rocha de Moura / Elżbieta Purta / Agata Bernat / Eva M Martín-Cuevas / Małgorzata Kurkowska / Eugene F Baulin / Sunandan Mukherjee / Jakub Nowak / Artur P Biela / Michał Rawski / ...Authors: Tales Rocha de Moura / Elżbieta Purta / Agata Bernat / Eva M Martín-Cuevas / Małgorzata Kurkowska / Eugene F Baulin / Sunandan Mukherjee / Jakub Nowak / Artur P Biela / Michał Rawski / Sebastian Glatt / Fernando Moreno-Herrero / Janusz M Bujnicki /
Abstract: Betacoronaviruses are a genus within the Coronaviridae family of RNA viruses. They are capable of infecting vertebrates and causing epidemics as well as global pandemics in humans. Mitigating the ...Betacoronaviruses are a genus within the Coronaviridae family of RNA viruses. They are capable of infecting vertebrates and causing epidemics as well as global pandemics in humans. Mitigating the threat posed by Betacoronaviruses requires an understanding of their molecular diversity. The development of novel antivirals hinges on understanding the key regulatory elements within the viral RNA genomes, in particular the 5'-proximal region, which is pivotal for viral protein synthesis. Using a combination of cryo-electron microscopy, atomic force microscopy, chemical probing, and computational modeling, we determined the structures of 5'-proximal regions in RNA genomes of Betacoronaviruses from four subgenera: OC43-CoV, SARS-CoV-2, MERS-CoV, and Rousettus bat-CoV. We obtained cryo-electron microscopy maps and determined atomic-resolution models for the stem-loop-5 (SL5) region at the translation start site and found that despite low sequence similarity and variable length of the helical elements it exhibits a remarkable structural conservation. Atomic force microscopy imaging revealed a common domain organization and a dynamic arrangement of structural elements connected with flexible linkers across all four Betacoronavirus subgenera. Together, these results reveal common features of a critical regulatory region shared between different Betacoronavirus RNA genomes, which may allow targeting of these RNAs by broad-spectrum antiviral therapeutics.
History
DepositionSep 28, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 6, 2024Provider: repository / Type: Initial release
Revision 1.1Mar 13, 2024Group: Database references / Category: citation / citation_author / Item: _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Apr 24, 2024Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: SARS-CoV-2-SL5


Theoretical massNumber of molelcules
Total (without water)46,5431
Polymers46,5431
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: RNA chain SARS-CoV-2-SL5


Mass: 46543.441 Da / Num. of mol.: 1 / Source method: obtained synthetically
Source: (synth.) Severe acute respiratory syndrome coronavirus 2
References: GenBank: MN908947.3

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Severe acute respiratory syndrome coronavirus 2SARS-CoV-2
Type: VIRUS / Details: in vitro transcription / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2
Source (recombinant)Organism: synthetic construct (others)
Details of virusEmpty: YES / Enveloped: YES / Isolate: OTHER / Type: VIROID
Natural hostOrganism: synthetic construct
Buffer solutionpH: 7.5 / Details: 20 mM HEPES pH 7.5, 50 mM KCl, 50 mM NaCl
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHepes1
250 mMsodium chlorideNaClSodium chloride1
350 mMPotassium chlorideKCl1
SpecimenConc.: 1.18 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK II / Cryogen name: ETHANE / Humidity: 90 % / Chamber temperature: 298 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 105000 X / Nominal defocus max: 2100 nm / Nominal defocus min: 900 nm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 40 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM software
IDNameVersionCategory
1cryoSPARC4particle selection
4cryoSPARC4CTF correction
10cryoSPARC4initial Euler assignment
11cryoSPARC4final Euler assignment
CTF correctionType: NONE
3D reconstructionResolution: 6.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 29234 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0023437
ELECTRON MICROSCOPYf_angle_d0.4955354
ELECTRON MICROSCOPYf_dihedral_angle_d13.7081737
ELECTRON MICROSCOPYf_chiral_restr0.023725
ELECTRON MICROSCOPYf_plane_restr0.003145

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