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- PDB-7sam: tRNA-like Structure from Brome Mosaic Virus -

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Basic information

Entry
Database: PDB / ID: 7sam
TitletRNA-like Structure from Brome Mosaic Virus
ComponentsViral RNA
KeywordsRNA / Viral RNA / 3' UTR
Function / homologyRNA / RNA (> 10) / RNA (> 100)
Function and homology information
Biological speciesBrome mosaic virus
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.3 Å
AuthorsKieft, J.S. / Bonilla, S.L.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM118070 United States
National Science Foundation (NSF, United States)F32GM139385 United States
Howard Hughes Medical Institute (HHMI)Hanna H. Gray Fellowship United States
CitationJournal: Science / Year: 2021
Title: A viral RNA hijacks host machinery using dynamic conformational changes of a tRNA-like structure.
Authors: Steve L Bonilla / Madeline E Sherlock / Andrea MacFadden / Jeffrey S Kieft /
Abstract: Viruses require multifunctional structured RNAs to hijack their host’s biochemistry, but their mechanisms can be obscured by the difficulty of solving conformationally dynamic RNA structures. Using ...Viruses require multifunctional structured RNAs to hijack their host’s biochemistry, but their mechanisms can be obscured by the difficulty of solving conformationally dynamic RNA structures. Using cryo–electron microscopy (cryo-EM), we visualized the structure of the mysterious viral transfer RNA (tRNA)–like structure (TLS) from the brome mosaic virus, which affects replication, translation, and genome encapsidation. Structures in isolation and those bound to tyrosyl-tRNA synthetase (TyrRS) show that this ~55-kilodalton purported tRNA mimic undergoes large conformational rearrangements to bind TyrRS in a form that differs substantially from that of tRNA. Our study reveals how viral RNAs can use a combination of static and dynamic RNA structures to bind host machinery through highly noncanonical interactions, and we highlight the utility of cryo-EM for visualizing small, conformationally dynamic structured RNAs.
History
DepositionSep 22, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 1, 2021Provider: repository / Type: Initial release
Revision 1.1Jun 5, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

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Assembly

Deposited unit
A: Viral RNA


Theoretical massNumber of molelcules
Total (without water)55,0451
Polymers55,0451
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: native gel electrophoresis, Conformational heterogeneity was assessed using gel electrophoresis under native conditions.
TypeNameSymmetry operationNumber
identity operation1_5551
MethodPISA

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Components

#1: RNA chain Viral RNA


Mass: 55044.559 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: In vitro transcribed / Source: (synth.) Brome mosaic virus

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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: tRNA-like structure at the 3' UTR of the brome mosaic virus (BMV) genome
Type: COMPLEX
Details: RNA was generated by in vitro transcription of sequence-verified linear DNA template
Entity ID: all / Source: MULTIPLE SOURCES
Molecular weightValue: 0.05456 MDa / Experimental value: NO
Source (natural)Organism: Brome mosaic virus
Buffer solutionpH: 7 / Details: Solution contained 10 mM magnesium chloride.
Buffer componentConc.: 50 mM / Name: NaMOPS
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: This sample was monodisperse as determined by native gel electrophoresis.
Specimen supportGrid mesh size: 400 divisions/in. / Grid type: C-flat-1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 278 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: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: -800 nm / Nominal defocus min: -2000 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 30 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

EM softwareName: cryoSPARC / Category: 3D reconstruction
CTF correctionType: NONE
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 128226 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
RefinementStereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 103.85 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00134021
ELECTRON MICROSCOPYf_angle_d0.396263
ELECTRON MICROSCOPYf_chiral_restr0.0216845
ELECTRON MICROSCOPYf_plane_restr0.0029169
ELECTRON MICROSCOPYf_dihedral_angle_d13.28252025

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