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- PDB-8p00: Cryo-EM structure of Rotavirus B NSP2 -

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Basic information

Entry
Database: PDB / ID: 8p00
TitleCryo-EM structure of Rotavirus B NSP2
ComponentsNon-structural protein 2
KeywordsVIRAL PROTEIN / Octamer / RNA binding / Rotavirus / RNA chaperone
Function / homologyhydrolase activity, acting on acid anhydrides / Rotavirus non-structural protein 2 / viral genome replication / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / host cell cytoplasm / RNA binding / ATP binding / metal ion binding / Non-structural protein 2
Function and homology information
Biological speciesHuman rotavirus B strain CAL-1
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsChamera, S. / Nowotny, M.
Funding support1items
OrganizationGrant numberCountry
Other government
CitationJournal: J Virol / Year: 2024
Title: Cryo-EM structure of rotavirus B NSP2 reveals its unique tertiary architecture.
Authors: Sebastian Chamera / Krzysztof Wycisk / Mariusz Czarnocki-Cieciura / Marcin Nowotny /
Abstract: Rotavirus (RV) NSP2 is a multifunctional RNA chaperone that exhibits numerous activities that are essential for replication and viral genome packaging. We performed an analysis that highlighted a ...Rotavirus (RV) NSP2 is a multifunctional RNA chaperone that exhibits numerous activities that are essential for replication and viral genome packaging. We performed an analysis that highlighted a distant relationship of NSP2 from rotavirus B (RVB) to proteins from other human RVs. We solved a cryo-electron microscopy structure of RVB NSP2 that shows structural differences with corresponding proteins from other human RVs. Based on the structure, we identified amino acid residues that are involved in RNA interactions. Anisotropy titration experiments showed that these residues are important for nucleic acid binding. We also identified structural motifs that are conserved in all RV species. Collectively, our data complete the structural characterization of rotaviral NSP2 protein and demonstrate its structural diversity among RV species.IMPORTANCERotavirus B (RVB), also known as adult diarrhea rotavirus, has caused epidemics of severe diarrhea in China, India, and Bangladesh. Thousands of people are infected in a single RVB epidemic. However, information on this group of rotaviruses remains limited. As NSP2 is an essential protein in the viral life cycle, including its role in the formation of replication factories, it may be a target for future antiviral strategy against viruses with similar mechanisms.
History
DepositionMay 9, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 28, 2024Provider: repository / Type: Initial release
Revision 1.1Mar 13, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2Mar 27, 2024Group: Database references / Category: citation / Item: _citation.journal_volume
Revision 1.3May 8, 2024Group: Source and taxonomy / Category: entity_src_gen
Item: _entity_src_gen.pdbx_gene_src_ncbi_taxonomy_id / _entity_src_gen.pdbx_gene_src_scientific_name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Non-structural protein 2
B: Non-structural protein 2
C: Non-structural protein 2
D: Non-structural protein 2
E: Non-structural protein 2
F: Non-structural protein 2
G: Non-structural protein 2
H: Non-structural protein 2


Theoretical massNumber of molelcules
Total (without water)276,6058
Polymers276,6058
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area9770 Å2
ΔGint-50 kcal/mol
Surface area127240 Å2
MethodPISA

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Components

#1: Protein
Non-structural protein 2 / NSP2 / NCVP3 / Non-structural RNA-binding protein 35 / NS35


Mass: 34575.672 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human rotavirus B strain CAL-1 / Gene: NSP2 / Production host: Escherichia coli (E. coli)
References: UniProt: Q6XD98, Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement

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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: Homooctamer of NSP2 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Rotavirus B / Strain: Mexico
Source (recombinant)Organism: Escherichia coli (E. coli)
Details of virusIsolate: OTHER / Type: VIRION
Buffer solutionpH: 7
SpecimenConc.: 0.36 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: C-flat-2/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 %

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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 defocus max: 2500 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 40.92 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.20.1_4487refinement
PHENIX1.20.1_4487refinement
EM softwareName: PHENIX / Version: 1.20.1_4487: / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 22131 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 118.68 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00217376
ELECTRON MICROSCOPYf_angle_d0.55323672
ELECTRON MICROSCOPYf_dihedral_angle_d3.7822496
ELECTRON MICROSCOPYf_chiral_restr0.0362792
ELECTRON MICROSCOPYf_plane_restr0.0053016

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