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- PDB-7t4s: CryoEM structure of the HCMV Pentamer gH/gL/UL128/UL130/UL131A in... -

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Basic information

Entry
Database: PDB / ID: 7t4s
TitleCryoEM structure of the HCMV Pentamer gH/gL/UL128/UL130/UL131A in complex with NRP2 and neutralizing fabs 8I21 and 13H11
Components
  • (Envelope glycoprotein ...) x 3
  • (Envelope protein ...) x 2
  • Fab 13H11 heavy chain
  • Fab 13H11 light chain
  • Fab 8I21 heavy chain
  • Fab 8I21 light chain
  • Neuropilin-2
KeywordsVIRAL PROTEIN/Immune System / glycoprotein complex / antibody complex / Neuropilin 2 / VIRAL PROTEIN / VIRAL PROTEIN-Immune System complex
Function / homology
Function and homology information


vestibulocochlear nerve structural organization / dorsal root ganglion morphogenesis / ventral trunk neural crest cell migration / sympathetic neuron projection guidance / facioacoustic ganglion development / trigeminal ganglion development / trigeminal nerve structural organization / sensory neuron axon guidance / facial nerve structural organization / branchiomotor neuron axon guidance ...vestibulocochlear nerve structural organization / dorsal root ganglion morphogenesis / ventral trunk neural crest cell migration / sympathetic neuron projection guidance / facioacoustic ganglion development / trigeminal ganglion development / trigeminal nerve structural organization / sensory neuron axon guidance / facial nerve structural organization / branchiomotor neuron axon guidance / gonadotrophin-releasing hormone neuronal migration to the hypothalamus / axon extension involved in axon guidance / sympathetic neuron projection extension / NrCAM interactions / Neurophilin interactions with VEGF and VEGFR / neural crest cell migration involved in autonomic nervous system development / sympathetic ganglion development / vascular endothelial growth factor receptor activity / nerve development / semaphorin receptor complex / semaphorin receptor activity / outflow tract septum morphogenesis / regulation of postsynapse organization / negative chemotaxis / growth factor binding / cytokine binding / semaphorin-plexin signaling pathway / positive regulation of endothelial cell proliferation / positive regulation of endothelial cell migration / axon guidance / cellular response to leukemia inhibitory factor / signaling receptor activity / heparin binding / angiogenesis / host cell endosome / host cell Golgi apparatus / postsynaptic membrane / entry receptor-mediated virion attachment to host cell / cell adhesion / axon / fusion of virus membrane with host plasma membrane / viral envelope / symbiont entry into host cell / host cell plasma membrane / glutamatergic synapse / virion membrane / extracellular region / metal ion binding / identical protein binding / membrane / plasma membrane
Similarity search - Function
Cytomegalovirus UL128 protein / Herpesvirus UL130, cytomegalovirus / HCMV glycoprotein pUL130 / Cytomegalovirus glycoprotein L / Cytomegalovirus glycoprotein L / Betaherpesvirus glycoprotein L (gL) domain profile. / Neuropilin / Neuropilin, C-terminal / C-terminal domain of neuropilin glycoprotein / Herpesvirus glycoprotein H main domain ...Cytomegalovirus UL128 protein / Herpesvirus UL130, cytomegalovirus / HCMV glycoprotein pUL130 / Cytomegalovirus glycoprotein L / Cytomegalovirus glycoprotein L / Betaherpesvirus glycoprotein L (gL) domain profile. / Neuropilin / Neuropilin, C-terminal / C-terminal domain of neuropilin glycoprotein / 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 / Domain in meprin, A5, receptor protein tyrosine phosphatase mu (and others) / : / Coagulation factors 5/8 type C domain (FA58C) signature 2. / MAM domain, meprin/A5/mu / MAM domain / MAM domain profile. / Coagulation factors 5/8 type C domain (FA58C) signature 1. / CUB domain / Domain first found in C1r, C1s, uEGF, and bone morphogenetic protein. / CUB domain / Spermadhesin, CUB domain superfamily / CUB domain profile. / Coagulation factor 5/8 C-terminal domain, discoidin domain / Coagulation factors 5/8 type C domain (FA58C) profile. / F5/8 type C domain / Coagulation factor 5/8 C-terminal domain / Galactose-binding domain-like / Galactose-binding-like domain superfamily / Concanavalin A-like lectin/glucanase domain superfamily / Jelly Rolls / Sandwich / Mainly Beta
Similarity search - Domain/homology
Envelope glycoprotein H / Neuropilin-2 / UL128 / UL130 / Envelope glycoprotein L / UL131A
Similarity search - Component
Biological speciesHuman betaherpesvirus 5
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsKschonsak, M. / Johnson, M.C. / Schelling, R. / Green, E.M. / Rouge, L. / Ho, H. / Patel, N. / Kilic, C. / Kraft, E. / Arthur, C.P. ...Kschonsak, M. / Johnson, M.C. / Schelling, R. / Green, E.M. / Rouge, L. / Ho, H. / Patel, N. / Kilic, C. / Kraft, E. / Arthur, C.P. / Rohou, A.L. / Comps-Agrar, L. / Martinez-Martin, N. / Perez, L. / Payandeh, J. / Ciferri, C.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Sci Adv / Year: 2022
Title: Structural basis for HCMV Pentamer receptor recognition and antibody neutralization.
Authors: Marc Kschonsak / Matthew C Johnson / Rachel Schelling / Evan M Green / Lionel Rougé / Hoangdung Ho / Nidhi Patel / Cem Kilic / Edward Kraft / Christopher P Arthur / Alexis L Rohou / ...Authors: Marc Kschonsak / Matthew C Johnson / Rachel Schelling / Evan M Green / Lionel Rougé / Hoangdung Ho / Nidhi Patel / Cem Kilic / Edward Kraft / Christopher P Arthur / Alexis L Rohou / Laetitia Comps-Agrar / Nadia Martinez-Martin / Laurent Perez / Jian Payandeh / Claudio Ciferri /
Abstract: Human cytomegalovirus (HCMV) represents the viral leading cause of congenital birth defects and uses the gH/gL/UL128-130-131A complex (Pentamer) to enter different cell types, including epithelial ...Human cytomegalovirus (HCMV) represents the viral leading cause of congenital birth defects and uses the gH/gL/UL128-130-131A complex (Pentamer) to enter different cell types, including epithelial and endothelial cells. Upon infection, Pentamer elicits the most potent neutralizing response against HCMV, representing a key vaccine candidate. Despite its relevance, the structural basis for Pentamer receptor recognition and antibody neutralization is largely unknown. Here, we determine the structures of Pentamer bound to neuropilin 2 (NRP2) and a set of potent neutralizing antibodies against HCMV. Moreover, we identify thrombomodulin (THBD) as a functional HCMV receptor and determine the structures of the Pentamer-THBD complex. Unexpectedly, both NRP2 and THBD also promote dimerization of Pentamer. Our results provide a framework for understanding HCMV receptor engagement, cell entry, antibody neutralization, and outline strategies for antiviral therapies against HCMV.
History
DepositionDec 10, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Mar 23, 2022Provider: repository / Type: Initial release
Revision 1.1Oct 30, 2024Group: Data collection / Refinement description / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_3d_fitting_list / em_admin / pdbx_entry_details / pdbx_initial_refinement_model / pdbx_modification_feature
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type / _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Envelope glycoprotein H
B: Envelope glycoprotein L
C: Envelope protein UL128
D: Envelope glycoprotein UL130
E: Envelope protein UL131A
F: Neuropilin-2
H: Fab 8I21 heavy chain
J: Fab 13H11 heavy chain
K: Fab 13H11 light chain
I: Fab 8I21 light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)387,92720
Polymers385,89610
Non-polymers2,03110
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Envelope glycoprotein ... , 3 types, 3 molecules ABD

#1: Protein Envelope glycoprotein H / gH


Mass: 87311.273 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human betaherpesvirus 5 / Gene: UL75, gH / Production host: Homo sapiens (human) / References: UniProt: F5H9T3
#2: Protein Envelope glycoprotein L / gL


Mass: 30846.492 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human betaherpesvirus 5 / Gene: UL115, gL / Production host: Homo sapiens (human) / References: UniProt: Q71DN9
#4: Protein Envelope glycoprotein UL130 / UL130


Mass: 28866.811 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human betaherpesvirus 5 / Gene: UL130 / Production host: Homo sapiens (human) / References: UniProt: Q38M07

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Envelope protein ... , 2 types, 2 molecules CE

#3: Protein Envelope protein UL128 / UL128


Mass: 19746.023 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human betaherpesvirus 5 / Gene: UL128 / Production host: Homo sapiens (human) / References: UniProt: Q38LY2
#5: Protein Envelope protein UL131A / Protein UL131A / UL131A


Mass: 15011.043 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human betaherpesvirus 5 / Gene: UL131A, HHV5wtgp112 / Production host: Homo sapiens (human) / References: UniProt: Q8AZ45

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Antibody , 4 types, 4 molecules HJKI

#7: Antibody Fab 8I21 heavy chain


Mass: 26816.311 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)
#8: Antibody Fab 13H11 heavy chain


Mass: 26600.086 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)
#9: Antibody Fab 13H11 light chain


Mass: 25780.020 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)
#10: Antibody Fab 8I21 light chain


Mass: 25995.080 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Escherichia coli (E. coli)

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Protein / Sugars / Non-polymers , 3 types, 11 molecules F

#11: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 9 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0
#12: Chemical ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Ca
#6: Protein Neuropilin-2 / Vascular endothelial cell growth factor 165 receptor 2


Mass: 98923.211 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: NRP2, VEGF165R2 / Production host: Homo sapiens (human) / References: UniProt: O60462

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Details

Has ligand of interestN
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

ComponentName: Pentameric complex of HCMV proteins gH, gL, UL128, UL130, UL131A bound to Neuropilin 2 and fabs of human neutralizing antibodies 8I21 and 13H11
Type: COMPLEX / Entity ID: #1-#10 / Source: MULTIPLE SOURCES
Molecular weightValue: 0.36 MDa / Experimental value: NO
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
1300 mMsodium chlorideNaCl1
225 mMHEPESHEPES1
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Sample was mildly crosslinked with 0.025% glutaraldehyde, incubated for 10 min at room temperature and quenched with 9 mM TRIS pH 7.5
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R0.6/1
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K / Details: blot for 3.5 seconds before plunging

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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 magnification: 165000 X / Nominal defocus max: 1500 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 10 sec. / Electron dose: 52 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 4 / Num. of real images: 21357
Details: Images were collected in movie-mode at 4 frames/second.
Image scansMovie frames/image: 40

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Processing

EM software
IDNameVersionCategory
1Gautomatch0.56particle selection
2SerialEM3.7.11image acquisition
4CTFFIND4.1.13CTF correction
7Coot0.89model fitting
9cisTEM1.02initial Euler assignment
10cisTEM1.02final Euler assignment
11RELION3.1classification
12cisTEM1.023D reconstruction
13PHENIX1.19.2model refinement
Image processingDetails: Movie frames were corrected for motion and aligned. Images with a CTF fit resolution of 6.0 A or better were selected for particle picking.
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 2532206 / Details: template-matching particle picking
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 2252924
Details: A composite map was generated from the three individual focused 3D maps.
Num. of class averages: 44 / Symmetry type: POINT
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
12QQO

2qqo

12QQO1PDBexperimental model
25VOB

5vob

15VOB2PDBexperimental model

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