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- EMDB-60932: Cryo-EM structure of the CHIKV nsP3 peptide in complex with the N... -

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Entry
Database: EMDB / ID: EMD-60932
TitleCryo-EM structure of the CHIKV nsP3 peptide in complex with the NTF2L domain of G3BP1 (Conformation I)
Map data
Sample
  • Complex: CHIKV nsP3 in complex with human G3BP1
    • Protein or peptide: Ras GTPase-activating protein-binding protein 1
    • Protein or peptide: Polyprotein P1234
KeywordsChikungunya virus / nsP3 / G3BP1 / protein-protein interaction. / VIRAL PROTEIN
Function / homology
Function and homology information


DNA/RNA helicase activity / positive regulation of stress granule assembly / host cell filopodium / mRNA methyltransferase activity / polynucleotide 5'-phosphatase activity / poly(A) RNA polymerase activity / mRNA modification / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / ribosomal small subunit binding / 7-methylguanosine mRNA capping ...DNA/RNA helicase activity / positive regulation of stress granule assembly / host cell filopodium / mRNA methyltransferase activity / polynucleotide 5'-phosphatase activity / poly(A) RNA polymerase activity / mRNA modification / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / ribosomal small subunit binding / 7-methylguanosine mRNA capping / positive regulation of type I interferon production / DNA helicase activity / cysteine-type peptidase activity / stress granule assembly / ribonucleoside triphosphate phosphatase activity / negative regulation of canonical Wnt signaling pathway / molecular condensate scaffold activity / host cell cytoplasmic vesicle membrane / cytoplasmic stress granule / endonuclease activity / DNA helicase / methylation / defense response to virus / perikaryon / forked DNA-dependent helicase activity / single-stranded 3'-5' DNA helicase activity / four-way junction helicase activity / double-stranded DNA helicase activity / Ras protein signal transduction / RNA helicase activity / RNA helicase / chromatin extrusion motor activity / ATP-dependent H2AZ histone chaperone activity / ATP-dependent H3-H4 histone complex chaperone activity / cohesin loader activity / viral RNA genome replication / DNA clamp loader activity / innate immune response / symbiont-mediated suppression of host gene expression / focal adhesion / RNA-directed RNA polymerase activity / DNA-templated transcription / mRNA binding / host cell nucleus / GTP binding / SARS-CoV-2 activates/modulates innate and adaptive immune responses / host cell plasma membrane / ATP hydrolysis activity / proteolysis / DNA binding / RNA binding / ATP binding / nucleus / metal ion binding / cytosol / cytoplasm
Similarity search - Function
G3BP1, RNA recognition motif / Ras GTPase-activating protein-binding protein / Nuclear transport factor 2, eukaryote / Nuclear transport factor 2 domain profile. / Nuclear transport factor 2 (NTF2) domain / Nuclear transport factor 2 domain / Alphavirus nsp2 protease (nsp2pro) domain / Alphavirus nsP2 protease domain superfamily / : / Peptidase family C9 ...G3BP1, RNA recognition motif / Ras GTPase-activating protein-binding protein / Nuclear transport factor 2, eukaryote / Nuclear transport factor 2 domain profile. / Nuclear transport factor 2 (NTF2) domain / Nuclear transport factor 2 domain / Alphavirus nsp2 protease (nsp2pro) domain / Alphavirus nsP2 protease domain superfamily / : / Peptidase family C9 / Tomato mosaic virus helicase, N-terminal domain / Alphavirus nsp2 protease (nsp2pro) domain profile. / : / Non-structural protein 3, zinc-binding domain / Viral methyltransferase / Alphavirus-like methyltransferase (MT) domain / Alphavirus-like methyltransferase (MT) domain profile. / Tymovirus, RNA-dependent RNA polymerase / RNA dependent RNA polymerase / Viral (Superfamily 1) RNA helicase / NTF2-like domain superfamily / RNA recognition motif / RNA recognition motif / Eukaryotic RNA Recognition Motif (RRM) profile. / RNA recognition motif domain / RNA-binding domain superfamily / (+) RNA virus helicase core domain / (+)RNA virus helicase core domain profile. / Non-structural protein 3, X-domain-like / Appr-1"-p processing enzyme / Macro domain / Macro domain / Macro domain profile. / Macro domain-like / Nucleotide-binding alpha-beta plait domain superfamily / RNA-directed RNA polymerase, catalytic domain / RdRp of positive ssRNA viruses catalytic domain profile. / S-adenosyl-L-methionine-dependent methyltransferase superfamily / DNA/RNA polymerase superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Polyprotein P1234 / Ras GTPase-activating protein-binding protein 1
Similarity search - Component
Biological speciesChikungunya virus / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.66 Å
AuthorsWang J / Liu YZ / Lei J / Wang KT
Funding support China, 2 items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, China)2021YFF0702004 China
Ministry of Science and Technology (MoST, China)2022YFC2303701 China
CitationJournal: mBio / Year: 2025
Title: Cryo-EM reveals a double oligomeric ring scaffold of the CHIKV nsP3 peptide in complex with the NTF2L domain of host G3BP1.
Authors: Yuanzhi Liu / Jie Wang / Yinze Han / Xiaoyan Xia / Rui Zeng / Xinyu Fan / Bo Zhang / Kaituo Wang / Jian Lei /
Abstract: Chikungunya virus (CHIKV) poses a severe threat to global public health. The interaction between CHIKV nsP3 and host G3BP1 is critical for viral replication. However, the exact structural mechanism ...Chikungunya virus (CHIKV) poses a severe threat to global public health. The interaction between CHIKV nsP3 and host G3BP1 is critical for viral replication. However, the exact structural mechanism of the nsP3-G3BP1 interaction is scarce. Here, we report a cryo-electron microscopy structure of an octameric-heterotrimer formed by CHIKV nsP3 peptide (designated as CHIKV-43) in complex with the nuclear translocation factor 2-like (NTF2L) domain of G3BP1. The overall structure presents a double-layer ring scaffold. Two FGDF motifs and two alpha helices of CHIKV-43 are essential to bind NTF2L. Particularly, the secondary alpha helix plays key roles in forming the CHIKV-43-NTF2L high-order oligomer. We next analyzed the detailed interactions between CHIKV-43 and the NTF2L domain. The different binding patterns of NTF2L with its various partners were described as well. Subsequently, we demonstrated that the CHIKV-43 peptide is a crucial factor for nsP3 co-localization with G3BP1, reducing stress granule formation and interfering with interferon production. Overall, our findings present the structural and functional mechanisms on CHIKV nsP3 modulating host G3BP1 and provide a potential antiviral target based on the protein-protein interaction interface.
IMPORTANCE: Chikungunya virus (CHIKV) is an arbovirus responsible for causing fever, headache, and severe joint pain in humans, with widespread outbreaks affecting millions worldwide. The CHIKV nsP3 ...IMPORTANCE: Chikungunya virus (CHIKV) is an arbovirus responsible for causing fever, headache, and severe joint pain in humans, with widespread outbreaks affecting millions worldwide. The CHIKV nsP3 is a key protein that interacts with multiple host proteins. In this study, we present the cryo-electron microscopy structure of a high-order oligomer formed by the CHIKV nsP3 peptide and the nuclear translocation factor 2-like (NTF2L) domain of host protein G3BP1, revealing a completely novel interaction model. The detailed interactions within this oligomer were illustrated. We also analyzed the binding patterns of the NTF2L domain of G3BP1 with its various partners, providing essential insights for the development of peptide-mimetic inhibitors targeting nsP3 and/or G3BP1. Furthermore, our data indicate that the nsP3-G3BP1 interaction reduces stress granule formation and antagonizes interferon production. Overall, this study provides new knowledge on CHIKV biology and suggests a potential target for developing antiviral therapeutics.
History
DepositionJul 24, 2024-
Header (metadata) releaseApr 9, 2025-
Map releaseApr 9, 2025-
UpdateApr 30, 2025-
Current statusApr 30, 2025Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_60932.png

Downloads & links

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Map

FileDownload / File: emd_60932.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

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AxesZ (Sec.)Y (Row.)X (Col.)
0.73 Å/pix.
x 320 pix.
= 232. Å		0.73 Å/pix.
x 320 pix.
= 232. Å		0.73 Å/pix.
x 320 pix.
= 232. Å

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.725 Å
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Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0485
Minimum - Maximum-0.120356835 - 0.23276116
Average (Standard dev.)0.000045792898 (±0.009653668)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 232.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_60932_half_map_1.map
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Half map: #1

Fileemd_60932_half_map_2.map
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Sample components

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Entire : CHIKV nsP3 in complex with human G3BP1

EntireName: CHIKV nsP3 in complex with human G3BP1
Components
  • Complex: CHIKV nsP3 in complex with human G3BP1
    • Protein or peptide: Ras GTPase-activating protein-binding protein 1
    • Protein or peptide: Polyprotein P1234

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Supramolecule #1: CHIKV nsP3 in complex with human G3BP1

SupramoleculeName: CHIKV nsP3 in complex with human G3BP1 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Chikungunya virus

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Macromolecule #1: Ras GTPase-activating protein-binding protein 1

MacromoleculeName: Ras GTPase-activating protein-binding protein 1 / type: protein_or_peptide / ID: 1 / Number of copies: 16 / Enantiomer: LEVO / EC number: DNA helicase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 16.057231 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
GASMVMEKPS PLLVGREFVR QYYTLLNQAP DMLHRFYGKN SSYVHGGLDS NGKPADAVYG QKEIHRKVMS QNFTNCHTKI RHVDAHATL NDGVVVQVMG LLSNNNQALR RFMQTFVLAP EGSVANKFYV HNDIFRYQDE VF

UniProtKB: Ras GTPase-activating protein-binding protein 1

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Macromolecule #2: Polyprotein P1234

MacromoleculeName: Polyprotein P1234 / type: protein_or_peptide / ID: 2 / Number of copies: 8 / Enantiomer: LEVO
Source (natural)Organism: Chikungunya virus
Molecular weightTheoretical: 6.000295 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
GPLGSETFPI TFGDFNDGEI ESLSSELLTF GDFLPGEVDD LTDSDWSTHH HHHH

UniProtKB: Polyprotein P1234

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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

BufferpH: 7.5
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: OTHER
Details: The value given for _em_vitrification.instrument is CRYOSOL VITROJET. This is not in a list of allowed values {'HOMEMADE PLUNGER', 'REICHERT-JUNG PLUNGER', 'LEICA KF80', 'GATAN CRYOPLUNGE ...Details: The value given for _em_vitrification.instrument is CRYOSOL VITROJET. This is not in a list of allowed values {'HOMEMADE PLUNGER', 'REICHERT-JUNG PLUNGER', 'LEICA KF80', 'GATAN CRYOPLUNGE 3', 'FEI VITROBOT MARK IV', 'LEICA PLUNGER', 'FEI VITROBOT MARK I', 'EMS-002 RAPID IMMERSION FREEZER', 'LEICA EM CPC', 'OTHER', 'SPOTITON', 'FEI VITROBOT MARK II', 'FEI VITROBOT MARK III', 'LEICA EM GP'} so OTHER is written into the XML file.

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.66 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 442651
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC

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