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- EMDB-18521: Conserved Structures and Dynamics in 5-Proximal Regions of Betaco... -

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Entry
Database: EMDB / ID: EMD-18521
TitleConserved Structures and Dynamics in 5-Proximal Regions of Betacoronavirus RNA Genomes
Map data
Sample
  • Virus: Rousettus bat coronavirus GCCDC1
    • RNA: RoBat-CoV-SL5
KeywordsRNA structure / 5-proximal region / Coronavirus / Cryo-EM / VIRUS
Biological speciesRousettus bat coronavirus GCCDC1
Methodsingle particle reconstruction / cryo EM / Resolution: 6.6 Å
AuthorsMoura TR / Purta E / Bernat A / Baulin E / Mukherjee S / Bujnicki JM
Funding support Poland, 1 items
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, 2023-
Header (metadata) releaseMar 6, 2024-
Map releaseMar 6, 2024-
UpdateApr 24, 2024-
Current statusApr 24, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18521.png

Downloads & links

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Map

FileDownload / File: emd_18521.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.86 Å
Density
Contour LevelBy AUTHOR: 0.05
Minimum - Maximum-0.104578964 - 0.49119845
Average (Standard dev.)0.0000036263405 (±0.005397401)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions512512512
Spacing512512512
CellA=B=C: 440.32 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_18521_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_18521_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Rousettus bat coronavirus GCCDC1

EntireName: Rousettus bat coronavirus GCCDC1
Components
  • Virus: Rousettus bat coronavirus GCCDC1
    • RNA: RoBat-CoV-SL5

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Supramolecule #1: Rousettus bat coronavirus GCCDC1

SupramoleculeName: Rousettus bat coronavirus GCCDC1 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all / Details: in vitro transcription / NCBI-ID: 2501962 / Sci species name: Rousettus bat coronavirus GCCDC1 / Virus type: VIROID / Virus isolate: OTHER / Virus enveloped: Yes / Virus empty: Yes
Host (natural)Organism: synthetic construct (others)

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Macromolecule #1: RoBat-CoV-SL5

MacromoleculeName: RoBat-CoV-SL5 / type: rna / ID: 1 / Number of copies: 1
Source (natural)Organism: Rousettus bat coronavirus GCCDC1
Molecular weightTheoretical: 37.739094 KDa
SequenceString:
AAGCUUUUGU GUCUUCAGCU UCCCUCUUCU CCUCGUGGGG UUCCGUUCCC GCGGUAGUGA GCCUGGUGUG GUUCCGCCCG CGCCAGUGA UGGAGGGUGC UUUAGGCACU AACAAGCUU

GENBANK: GENBANK: NC_030886.1

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.9 mg/mL
BufferpH: 7.5
Component:
ConcentrationNameFormula
20.0 mMHepes
50.0 mMsodium chlorideNaClSodium chloride
50.0 mMPotassium chlorideKCl

Details: 20 mM HEPES pH 7.5, 50 mM KCl, 50 mM NaCl
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK II

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.1 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 40.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL / In silico model: Generated by SimRNA
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 4)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 4)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 6.6 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 29234
FSC plot (resolution estimation)

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