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- EMDB-43720: Cryo-EM Map of KSHV ORF74-BRIL-BAK5-Nb complex at 3.7A -

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Basic information

Entry
Database: EMDB / ID: EMD-43720
TitleCryo-EM Map of KSHV ORF74-BRIL-BAK5-Nb complex at 3.7A
Map dataCryo-EM Map of KSHV ORF74-BRIL-BAG2-Nb complex at 3.7A
Sample
  • Complex: Apo Inactive Conformation of KSHV ORF74 (Viral G Protein-Coupled Receptor)-BRIL in Complex with BAG2-Nb
    • Protein or peptide: Cryo-EM Map of KSHV ORF74-BRIL-BAG2-Nb complex
KeywordsKSHV vGPCR / Viral GPCR / KSHV ORF74 / ORF74-BRIL-BAG2-Nb / ORF74 Apo / ORF74 Inactive / ORF74 Apo Monomer / VIRAL PROTEIN
Biological speciesHuman gammaherpesvirus 8
Methodsingle particle reconstruction / cryo EM / Resolution: 3.73 Å
AuthorsSahoo B / Seo HD / Dai X / Jung J
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)R01CA251275 United States
CitationJournal: Nat Commun / Year: 2025
Title: Structural basis for ligand promiscuity and high signaling activity of Kaposi's Sarcoma-associated Herpesvirus-encoded GPCR.
Authors: Jun Bae Park / Bibekananda Sahoo / Amita Rani Sahoo / Dokyun Kim / Hogyu David Seo / James Bowman / Mi-Jeong Kwak / Sophia Suh / Matthias Buck / Xinghong Dai / Jae U Jung /
Abstract: Kaposi's Sarcoma-associated Herpesvirus encodes ORF74, a viral G protein-coupled receptor homologous to CXCR2, which plays a crucial role in Kaposi's Sarcoma development through its high basal ...Kaposi's Sarcoma-associated Herpesvirus encodes ORF74, a viral G protein-coupled receptor homologous to CXCR2, which plays a crucial role in Kaposi's Sarcoma development through its high basal signaling activity. Our cryoEM analysis of ORF74 in ligand-free, BRIL-fused ligand-free, and CXCL1/Gi-bound forms elucidates its ligand-independent signaling activity. A widely open, static extracellular cavity facilitates ligand promiscuity by enabling dynamic access and diverse binding modes. Structural alterations in CWxP, E/DRY, and NPxxY micro-switches stabilize the active conformation, leading to constitutive signaling. Metadynamics simulations reveal a dynamic ensemble between local switch structures corresponding to the inactive and active states, supporting spontaneous activation. CXCR2-ORF74 chimeras highlight intracellular loops 2 and 3 as key modulators of basal and agonist-induced activity. This study defines the structural basis of ORF74's ligand promiscuity, spontaneous activation, and high basal signaling, providing insights into its role in viral oncogenesis.
History
DepositionFeb 15, 2024-
Header (metadata) releaseAug 27, 2025-
Map releaseAug 27, 2025-
UpdateOct 8, 2025-
Current statusOct 8, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43720.png

Downloads & links

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Map

FileDownload / File: emd_43720.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM Map of KSHV ORF74-BRIL-BAG2-Nb complex at 3.7A
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.66 Å/pix.
x 360 pix.
= 237.6 Å		0.66 Å/pix.
x 360 pix.
= 237.6 Å		0.66 Å/pix.
x 360 pix.
= 237.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.66 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.17
Minimum - Maximum-0.81163126 - 1.3447232
Average (Standard dev.)0.00022749757 (±0.022775248)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 237.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: KSHV ORF74-BRIL-BAG2-Nb complex Cryo-EM Half Map A

Fileemd_43720_half_map_1.map
AnnotationKSHV ORF74-BRIL-BAG2-Nb complex Cryo-EM Half Map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: KSHV ORF74-BRIL-BAG2-Nb complex Cryo-EM Half Map B

Fileemd_43720_half_map_2.map
AnnotationKSHV ORF74-BRIL-BAG2-Nb complex Cryo-EM Half Map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Apo Inactive Conformation of KSHV ORF74 (Viral G Protein-Coupled ...

EntireName: Apo Inactive Conformation of KSHV ORF74 (Viral G Protein-Coupled Receptor)-BRIL in Complex with BAG2-Nb
Components
  • Complex: Apo Inactive Conformation of KSHV ORF74 (Viral G Protein-Coupled Receptor)-BRIL in Complex with BAG2-Nb
    • Protein or peptide: Cryo-EM Map of KSHV ORF74-BRIL-BAG2-Nb complex

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Supramolecule #1: Apo Inactive Conformation of KSHV ORF74 (Viral G Protein-Coupled ...

SupramoleculeName: Apo Inactive Conformation of KSHV ORF74 (Viral G Protein-Coupled Receptor)-BRIL in Complex with BAG2-Nb
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Human gammaherpesvirus 8
Molecular weightTheoretical: 53.9 KDa

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Macromolecule #1: Cryo-EM Map of KSHV ORF74-BRIL-BAG2-Nb complex

MacromoleculeName: Cryo-EM Map of KSHV ORF74-BRIL-BAG2-Nb complex / type: protein_or_peptide / ID: 1
Details: The KSHV-BRIL (sequence provided above) construct bound to BAG2-Nb (purified separately) makes up the ORF74-BRIL-BAG2-Nb complex.
Enantiomer: LEVO
Source (natural)Organism: Human gammaherpesvirus 8
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MAAEDFLTIF LDDDESWNET LNMSGYDYSC NFSLEVSVCE MTTVVPYTWN VGILSLIFLI NVLGNGLVTY IFCKHRSRAG AIDILLLGIC LNSLCLSISL LAEVLMFLFP NIISTGLCRL EIFFYYLYVY LDIFSVVCVS LVRYLLVAYS TRSWPKKQSL GWVLTSAAWL ...String:
MAAEDFLTIF LDDDESWNET LNMSGYDYSC NFSLEVSVCE MTTVVPYTWN VGILSLIFLI NVLGNGLVTY IFCKHRSRAG AIDILLLGIC LNSLCLSISL LAEVLMFLFP NIISTGLCRL EIFFYYLYVY LDIFSVVCVS LVRYLLVAYS TRSWPKKQSL GWVLTSAAWL IALVLSGDAC RHRSRVVDPV SKQAMCYENA GNMTADWRLH VRTVSVTAGF LLPLALLILF YALTWCVVRR TKSAADLEDN WETLNDNLKV IEKADNAAQV KDALTKMRAA ALDAQKATPP KLEDKSPDSP EMKDFRHGFD ILVGQIDDAL KLANEGKVKE AQAAAEQLKT TRNAYIQKYL SALQARRKVR GVIVAVVVLF FVFCFPYHVL NLLDTLLRRR WIRDSCYTRG LINVGLAVTS LLQALYSAVV PLIYSCLGSK FRQRMYGLFQ SLRQSFMSGA DYKDDDDKle vlfqgpHHHH HHHHHH

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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

Concentration10 mg/mL
BufferpH: 7.5 / Component - Concentration: 10.0 mM / Component - Formula: C8H18N2O4S / Component - Name: HPEPS
Details: 10 mM HEPES pH 7.5, 150 mM NaCl, 0.005% (w/v) LMNG, 0.0005% (w/v) CHS
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 278 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER / Details: AB-INITIO RECONSTRUCTION
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.73 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 111016
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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