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- PDB-7ram: Cryo-EM Structure of the HCMV gHgLgO Trimer Derived from AD169 an... -

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Basic information

Entry
Database: PDB / ID: 7ram
TitleCryo-EM Structure of the HCMV gHgLgO Trimer Derived from AD169 and TR strains in complex with PDGFRalpha
Components
  • Envelope glycoprotein H
  • Envelope glycoprotein L
  • Envelope glycoprotein O
  • Platelet-derived growth factor receptor alpha
KeywordsVIRAL PROTEIN / viral entry glycoprotein in complex with receptor
Function / homology
Function and homology information


Imatinib-resistant PDGFR mutants / Sunitinib-resistant PDGFR mutants / Regorafenib-resistant PDGFR mutants / Sorafenib-resistant PDGFR mutants / PDGFR mutants bind TKIs / platelet-derived growth factor receptor-alpha signaling pathway / platelet-derived growth factor receptor-ligand complex / platelet-derived growth factor alpha-receptor activity / metanephric glomerular capillary formation / regulation of mesenchymal stem cell differentiation ...Imatinib-resistant PDGFR mutants / Sunitinib-resistant PDGFR mutants / Regorafenib-resistant PDGFR mutants / Sorafenib-resistant PDGFR mutants / PDGFR mutants bind TKIs / platelet-derived growth factor receptor-alpha signaling pathway / platelet-derived growth factor receptor-ligand complex / platelet-derived growth factor alpha-receptor activity / metanephric glomerular capillary formation / regulation of mesenchymal stem cell differentiation / luteinization / positive regulation of cell proliferation by VEGF-activated platelet derived growth factor receptor signaling pathway / platelet-derived growth factor binding / vascular endothelial growth factor binding / embryonic skeletal system morphogenesis / retina vasculature development in camera-type eye / vascular endothelial growth factor receptor activity / embryonic digestive tract morphogenesis / cardiac myofibril assembly / Leydig cell differentiation / cell activation / Signaling by PDGF / positive regulation of chemotaxis / male genitalia development / embryonic cranial skeleton morphogenesis / phospholipase C activator activity / platelet-derived growth factor receptor binding / estrogen metabolic process / face morphogenesis / signal transduction involved in regulation of gene expression / adrenal gland development / microvillus / platelet-derived growth factor receptor signaling pathway / odontogenesis of dentin-containing tooth / roof of mouth development / negative regulation of platelet activation / white fat cell differentiation / hematopoietic progenitor cell differentiation / Signaling by PDGFRA transmembrane, juxtamembrane and kinase domain mutants / Signaling by PDGFRA extracellular domain mutants / extracellular matrix organization / transmembrane receptor protein tyrosine kinase activity / Downstream signal transduction / positive regulation of calcium-mediated signaling / lung development / cell surface receptor protein tyrosine kinase signaling pathway / peptidyl-tyrosine phosphorylation / host cell endosome membrane / cellular response to reactive oxygen species / cell chemotaxis / regulation of actin cytoskeleton organization / cellular response to amino acid stimulus / wound healing / receptor protein-tyrosine kinase / platelet aggregation / positive regulation of fibroblast proliferation / Constitutive Signaling by Aberrant PI3K in Cancer / cell junction / cell migration / PIP3 activates AKT signaling / protein autophosphorylation / PI5P, PP2A and IER3 Regulate PI3K/AKT Signaling / RAF/MAP kinase cascade / host cell Golgi apparatus / in utero embryonic development / entry receptor-mediated virion attachment to host cell / positive regulation of ERK1 and ERK2 cascade / positive regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / receptor complex / protein kinase activity / nuclear body / cilium / positive regulation of cell migration / fusion of virus membrane with host plasma membrane / external side of plasma membrane / positive regulation of cell population proliferation / viral envelope / symbiont entry into host cell / endoplasmic reticulum membrane / protein-containing complex binding / host cell plasma membrane / virion membrane / Golgi apparatus / protein homodimerization activity / protein-containing complex / nucleoplasm / ATP binding / nucleus / membrane / plasma membrane / cytoplasm / cytosol
Similarity search - Function
Herpesvirus UL74, glycoprotein / Herpes UL74 glycoproteins / Platelet-derived growth factor receptor alpha / Cytomegalovirus glycoprotein L / Cytomegalovirus glycoprotein L / Betaherpesvirus glycoprotein L (gL) domain profile. / Herpesvirus glycoprotein H main domain / Herpesvirus glycoprotein H / Herpesvirus glycoprotein H, C-terminal / Herpesvirus glycoprotein H, C-terminal domain superfamily ...Herpesvirus UL74, glycoprotein / Herpes UL74 glycoproteins / Platelet-derived growth factor receptor alpha / Cytomegalovirus glycoprotein L / Cytomegalovirus glycoprotein L / Betaherpesvirus glycoprotein L (gL) domain profile. / Herpesvirus glycoprotein H main domain / Herpesvirus glycoprotein H / Herpesvirus glycoprotein H, C-terminal / Herpesvirus glycoprotein H, C-terminal domain superfamily / Herpesvirus glycoprotein H C-terminal domain / Platelet-derived growth factor receptor Ig-like domain 4 / Tyrosine-protein kinase, receptor class III, conserved site / Receptor tyrosine kinase class III signature. / Immunoglobulin I-set / Immunoglobulin I-set domain / : / Immunoglobulin subtype 2 / Immunoglobulin C-2 Type / Tyrosine-protein kinase, catalytic domain / Tyrosine kinase, catalytic domain / Tyrosine protein kinases specific active-site signature. / Immunoglobulin subtype / Immunoglobulin / Tyrosine-protein kinase, active site / Serine-threonine/tyrosine-protein kinase, catalytic domain / Protein tyrosine and serine/threonine kinase / Ig-like domain profile. / Immunoglobulin-like domain / Immunoglobulin-like domain superfamily / Serine/Threonine protein kinases, catalytic domain / Protein kinase, ATP binding site / Protein kinases ATP-binding region signature. / Immunoglobulin-like fold / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Envelope glycoprotein H / Platelet-derived growth factor receptor alpha / Envelope glycoprotein L / Glycoprotein O
Similarity search - Component
Biological speciesHuman herpesvirus 5 strain AD169
Human betaherpesvirus 5
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.43 Å
AuthorsLiu, J. / Vanarsdall, A.L. / Chen, D. / Johnson, D.C. / Jardetzky, T.S.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01AI150659 United States
CitationJournal: mBio / Year: 2021
Title: Cryo-Electron Microscopy Structure and Interactions of the Human Cytomegalovirus gHgLgO Trimer with Platelet-Derived Growth Factor Receptor Alpha.
Authors: Jing Liu / Adam Vanarsdall / Dong-Hua Chen / Andrea Chin / David Johnson / Theodore S Jardetzky /
Abstract: Human cytomegalovirus (HCMV) is a herpesvirus that produces disease in transplant patients and newborn children. Entry of HCMV into cells relies on gH/gL trimer (gHgLgO) and pentamer (gHgLUL128-131) ...Human cytomegalovirus (HCMV) is a herpesvirus that produces disease in transplant patients and newborn children. Entry of HCMV into cells relies on gH/gL trimer (gHgLgO) and pentamer (gHgLUL128-131) complexes that bind cellular receptors. Here, we studied the structure and interactions of the HCMV trimer, formed by AD169 strain gH and gL and TR strain gO proteins, with the human platelet-derived growth factor receptor alpha (PDGFRα). Three trimer surfaces make extensive contacts with three PDGFRα N-terminal domains, causing PDGFRα to wrap around gO in a structure similar to a human hand, explaining the high-affinity interaction. gO is among the least conserved HCMV proteins, with 8 distinct genotypes. We observed high conservation of residues mediating gO-gL interactions but more extensive gO variability in the PDGFRα interface. Comparisons between our trimer structure and a previously determined structure composed of different subunit genotypes indicate that gO variability is accommodated by adjustments in the gO-PDGFRα interface. We identified two loops within gO that were disordered and apparently glycosylated, which could be deleted without disrupting PDGFRα binding. We also identified four gO residues that contact PDGFRα, which when mutated produced markedly reduced receptor binding. These residues fall within conserved contact sites of gO with PDGFRα and may represent key targets for anti-trimer neutralizing antibodies and HCMV vaccines. Finally, we observe that gO mutations distant from the gL interaction site impact trimer expression, suggesting that the intrinsic folding or stability of gO can impact the efficiency of trimer assembly. HCMV is a herpesvirus that infects a large percentage of the adult population and causes significant levels of disease in immunocompromised individuals and birth defects in the developing fetus. The virus encodes a complex protein machinery that coordinates infection of different cell types in the body, including a trimer formed of gH, gL, and gO subunits. Here, we studied the interactions of the HCMV trimer with its receptor on cells, the platelet derived growth factor receptor α (PDGFRα), to better understand how HCMV coordinates virus entry into cells. Our results add to our understanding of HCMV strain-specific differences and identify sites on the trimer that represent potential targets for therapeutic antibodies or vaccine development.
History
DepositionJul 2, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 8, 2022Provider: repository / Type: Initial release
Revision 1.1Nov 13, 2024Group: Data collection / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Envelope glycoprotein H
B: Envelope glycoprotein L
C: Envelope glycoprotein O
D: Platelet-derived growth factor receptor alpha


Theoretical massNumber of molelcules
Total (without water)220,8814
Polymers220,8814
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area18050 Å2
ΔGint-105 kcal/mol
Surface area62210 Å2

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Components

#1: Protein Envelope glycoprotein H / gH


Mass: 80435.430 Da / Num. of mol.: 1 / Fragment: UNP Residues 41-718
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human herpesvirus 5 strain AD169 / Strain: AD169 / Gene: gH, UL75 / Production host: Homo sapiens (human) / References: UniProt: P12824
#2: Protein Envelope glycoprotein L / gL


Mass: 27138.018 Da / Num. of mol.: 1 / Fragment: UNP Residues 31-278
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human herpesvirus 5 strain AD169 / Strain: AD169 / Gene: gL, UL115 / Plasmid: pTT5 / Cell line (production host): HEK293-6E / Production host: Homo sapiens (human) / References: UniProt: P16832
#3: Protein Envelope glycoprotein O / Glycoprotein O / UL74 / UL74 protein


Mass: 53937.715 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human betaherpesvirus 5 / Gene: UL74 / Production host: Homo sapiens (human) / References: UniProt: Q8AZ32
#4: Protein Platelet-derived growth factor receptor alpha / PDGF-R-alpha / PDGFR-alpha / Alpha platelet-derived growth factor receptor / Alpha-type platelet- ...PDGF-R-alpha / PDGFR-alpha / Alpha platelet-derived growth factor receptor / Alpha-type platelet-derived growth factor receptor / CD140 antigen-like family member A / CD140a antigen / Platelet-derived growth factor alpha receptor / Platelet-derived growth factor receptor 2 / PDGFR-2


Mass: 59370.016 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PDGFRA, PDGFR2, RHEPDGFRA / Production host: Homo sapiens (human)
References: UniProt: P16234, receptor protein-tyrosine kinase
Has protein modificationY

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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

Component
IDNameTypeEntity IDParent-IDSource
1HCMV trimer in complex with receptor PDGFRalphaCOMPLEXall0RECOMBINANT
2human platelet-derived growth factor receptor alpha extracellular domainCOMPLEX#41RECOMBINANT
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
110.22 MDaNO
21NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
21Human herpesvirus 5 strain AD16910360
32Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
21Homo sapiens (human)9606
32Homo sapiens (human)9606
Buffer solutionpH: 7.5 / Details: 300mM NaCl and 20mM HEPES7.5
SpecimenConc.: 1.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 96 % / Chamber temperature: 293 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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2400 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 75 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
EM imaging opticsEnergyfilter slit width: 20 eV

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.19.2_4158refinement
PHENIX1.19.2_4158refinement
EM software
IDNameCategory
4cryoSPARCCTF correction
9cryoSPARCinitial Euler assignment
CTF correctionType: NONE
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.43 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 345997 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 94.19 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.002212293
ELECTRON MICROSCOPYf_angle_d0.524116745
ELECTRON MICROSCOPYf_chiral_restr0.04081921
ELECTRON MICROSCOPYf_plane_restr0.00392142
ELECTRON MICROSCOPYf_dihedral_angle_d4.29921649

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