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- PDB-3j8d: Cryoelectron microscopy of dengue-Fab E104 complex at pH 5.5 -

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Basic information

Entry
Database: PDB / ID: 3j8d
TitleCryoelectron microscopy of dengue-Fab E104 complex at pH 5.5
Components
  • Envelope protein E
  • antibody E111 Fab fragment
  • glycoprotein DIII
KeywordsVIRUS/IMMUNE SYSTEM / Dengue virus / DENV2 Fab E104 / Low pH / fusion trimer / VIRUS-IMMUNE SYSTEM complex
Function / homology
Function and homology information


symbiont-mediated suppression of host JAK-STAT cascade via inhibition of host TYK2 activity / host cell mitochondrion / flavivirin / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT2 activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of MAVS activity / : / viral capsid / nucleoside-triphosphate phosphatase / double-stranded RNA binding / protein complex oligomerization ...symbiont-mediated suppression of host JAK-STAT cascade via inhibition of host TYK2 activity / host cell mitochondrion / flavivirin / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT2 activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of MAVS activity / : / viral capsid / nucleoside-triphosphate phosphatase / double-stranded RNA binding / protein complex oligomerization / monoatomic ion channel activity / mRNA (guanine-N7)-methyltransferase / methyltransferase cap1 / clathrin-dependent endocytosis of virus by host cell / mRNA (nucleoside-2'-O-)-methyltransferase activity / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / RNA helicase activity / host cell endoplasmic reticulum membrane / host cell perinuclear region of cytoplasm / protein dimerization activity / RNA helicase / induction by virus of host autophagy / RNA-directed RNA polymerase / viral RNA genome replication / RNA-dependent RNA polymerase activity / serine-type endopeptidase activity / fusion of virus membrane with host endosome membrane / viral envelope / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / host cell nucleus / structural molecule activity / virion attachment to host cell / virion membrane / ATP hydrolysis activity / proteolysis / extracellular region / ATP binding / membrane / metal ion binding
Similarity search - Function
: / : / Flavivirus envelope glycoprotein E, stem/anchor domain / RNA-directed RNA polymerase, thumb domain, Flavivirus / Flavivirus RNA-directed RNA polymerase, thumb domain / Flavivirus capsid protein C superfamily / Flavivirus non-structural protein NS2B / Flavivirus NS3 helicase, C-terminal helical domain / Genome polyprotein, Flavivirus / Flavivirus non-structural protein NS4A ...: / : / Flavivirus envelope glycoprotein E, stem/anchor domain / RNA-directed RNA polymerase, thumb domain, Flavivirus / Flavivirus RNA-directed RNA polymerase, thumb domain / Flavivirus capsid protein C superfamily / Flavivirus non-structural protein NS2B / Flavivirus NS3 helicase, C-terminal helical domain / Genome polyprotein, Flavivirus / Flavivirus non-structural protein NS4A / Flavivirus non-structural protein NS2B / Flavivirus capsid protein C / Flavivirus non-structural protein NS4B / mRNA cap 0/1 methyltransferase / Flavivirus capsid protein C / Flavivirus non-structural protein NS4B / Flavivirus non-structural protein NS4A / Flavivirus NS2B domain profile. / mRNA cap 0 and cap 1 methyltransferase (EC 2.1.1.56 and EC 2.1.1.57) domain profile. / Flavivirus non-structural protein NS2A / Flavivirus non-structural protein NS2A / Flavivirus NS3, petidase S7 / Peptidase S7, Flavivirus NS3 serine protease / Flavivirus NS3 protease (NS3pro) domain profile. / Envelope glycoprotein M, flavivirus / Flavivirus envelope glycoprotein M / RNA-directed RNA polymerase, flavivirus / Flavivirus RNA-directed RNA polymerase, fingers and palm domains / Flavivirus non-structural Protein NS1 / Flavivirus non-structural protein NS1 / Envelope glycoprotein M superfamily, flavivirus / Flavivirus polyprotein propeptide / Flavivirus polyprotein propeptide superfamily / Flavivirus polyprotein propeptide / Flaviviral glycoprotein E, central domain, subdomain 1 / Flaviviral glycoprotein E, central domain, subdomain 2 / Flavivirus envelope glycoprotein E, Stem/Anchor domain / Flavivirus glycoprotein E, immunoglobulin-like domain / Flavivirus envelope glycoprotein E, Stem/Anchor domain superfamily / Flavivirus glycoprotein, immunoglobulin-like domain / Flavivirus glycoprotein central and dimerisation domain / Flavivirus glycoprotein, central and dimerisation domains / Ribosomal RNA methyltransferase, FtsJ domain / FtsJ-like methyltransferase / Flavivirus/Alphavirus glycoprotein, immunoglobulin-like domain superfamily / Flavivirus glycoprotein, central and dimerisation domain superfamily / Flaviviral glycoprotein E, dimerisation domain / DEAD box, Flavivirus / Flavivirus DEAD domain / helicase superfamily c-terminal domain / Immunoglobulin E-set / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / RNA-directed RNA polymerase, catalytic domain / RdRp of positive ssRNA viruses catalytic domain profile. / S-adenosyl-L-methionine-dependent methyltransferase superfamily / Peptidase S1, PA clan, chymotrypsin-like fold / DNA/RNA polymerase superfamily / Peptidase S1, PA clan / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Genome polyprotein / Genome polyprotein / Genome polyprotein
Similarity search - Component
Biological speciesDengue virus 2 Thailand/16681/84
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 26 Å
AuthorsZhang, X.Z. / Sheng, J. / Austin, S.K. / Hoornweg, T. / Smit, J.M. / Kuhn, R.J. / Diamond, M.S. / Rossmann, M.G.
CitationJournal: J Virol / Year: 2015
Title: Structure of acidic pH dengue virus showing the fusogenic glycoprotein trimers.
Authors: Xinzheng Zhang / Ju Sheng / S Kyle Austin / Tabitha E Hoornweg / Jolanda M Smit / Richard J Kuhn / Michael S Diamond / Michael G Rossmann /
Abstract: Flaviviruses undergo large conformational changes during their life cycle. Under acidic pH conditions, the mature virus forms transient fusogenic trimers of E glycoproteins that engage the lipid ...Flaviviruses undergo large conformational changes during their life cycle. Under acidic pH conditions, the mature virus forms transient fusogenic trimers of E glycoproteins that engage the lipid membrane in host cells to initiate viral fusion and nucleocapsid penetration into the cytoplasm. However, the dynamic nature of the fusogenic trimer has made the determination of its structure a challenge. Here we have used Fab fragments of the neutralizing antibody DV2-E104 to stop the conformational change of dengue virus at an intermediate stage of the fusion process. Using cryo-electron microscopy, we show that in this intermediate stage, the E glycoproteins form 60 trimers that are similar to the predicted "open" fusogenic trimer.
IMPORTANCE: The structure of a dengue virus has been captured during the formation of fusogenic trimers. This was accomplished by binding Fab fragments of the neutralizing antibody DV2-E104 to the ...IMPORTANCE: The structure of a dengue virus has been captured during the formation of fusogenic trimers. This was accomplished by binding Fab fragments of the neutralizing antibody DV2-E104 to the virus at neutral pH and then decreasing the pH to 5.5. These trimers had an "open" conformation, which is distinct from the "closed" conformation of postfusion trimers. Only two of the three E proteins within each spike are bound by a Fab molecule at domain III. Steric hindrance around the icosahedral 3-fold axes prevents binding of a Fab to the third domain III of each E protein spike. Binding of the DV2-E104 Fab fragments prevents domain III from rotating by about 130° to the postfusion orientation and thus precludes the stem region from "zipping" together the three E proteins along the domain II boundaries into the "closed" postfusion conformation, thus inhibiting fusion.
History
DepositionOct 13, 2014Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 12, 2014Provider: repository / Type: Initial release
Revision 1.1Dec 24, 2014Group: Database references
Revision 1.2Jul 26, 2017Group: Source and taxonomy / Category: entity_src_nat
Item: _entity_src_nat.common_name / _entity_src_nat.pdbx_ncbi_taxonomy_id / _entity_src_nat.pdbx_organism_scientific
Revision 1.3Jul 18, 2018Group: Data collection / Category: em_software / Item: _em_software.image_processing_id / _em_software.name
Revision 1.4Feb 21, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / pdbx_initial_refinement_model / pdbx_struct_oper_list / struct_ref_seq_dif
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _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 / _pdbx_struct_oper_list.name / _pdbx_struct_oper_list.symmetry_operation / _pdbx_struct_oper_list.type / _struct_ref_seq_dif.details

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Structure visualization

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  • Biological unit as complete icosahedral assembly
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  • Biological unit as icosahedral pentamer
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  • Biological unit as icosahedral 23 hexamer
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  • Deposited structure unit
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  • Simplified surface model + fitted atomic model
  • EMDB-6146
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Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
B: glycoprotein DIII
A: antibody E111 Fab fragment
D: antibody E111 Fab fragment
F: glycoprotein DIII
G: Envelope protein E
H: Envelope protein E
I: Envelope protein E


Theoretical massNumber of molelcules
Total (without water)245,9667
Polymers245,9667
Non-polymers00
Water0
1
B: glycoprotein DIII
A: antibody E111 Fab fragment
D: antibody E111 Fab fragment
F: glycoprotein DIII
G: Envelope protein E
H: Envelope protein E
I: Envelope protein E
x 60


Theoretical massNumber of molelcules
Total (without water)14,757,949420
Polymers14,757,949420
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation59
2


  • Idetical with deposited unit
  • icosahedral asymmetric unit
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
B: glycoprotein DIII
A: antibody E111 Fab fragment
D: antibody E111 Fab fragment
F: glycoprotein DIII
G: Envelope protein E
H: Envelope protein E
I: Envelope protein E
x 5


  • icosahedral pentamer
  • 1.23 MDa, 35 polymers
Theoretical massNumber of molelcules
Total (without water)1,229,82935
Polymers1,229,82935
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation4
4
B: glycoprotein DIII
A: antibody E111 Fab fragment
D: antibody E111 Fab fragment
F: glycoprotein DIII
G: Envelope protein E
H: Envelope protein E
I: Envelope protein E
x 6


  • icosahedral 23 hexamer
  • 1.48 MDa, 42 polymers
Theoretical massNumber of molelcules
Total (without water)1,475,79542
Polymers1,475,79542
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation5
5


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit, std point frame
TypeNameSymmetry operationNumber
transform to point frame1
SymmetryPoint symmetry: (Schoenflies symbol: I (icosahedral))

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Components

#1: Protein glycoprotein DIII / Coordinate model: Cα atoms only


Mass: 10616.091 Da / Num. of mol.: 2 / Fragment: SEE REMARK 999
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Dengue virus 2 Thailand/16681/84 / Production host: unidentified (others) / References: UniProt: P17763*PLUS
#2: Antibody antibody E111 Fab fragment / Coordinate model: Cα atoms only


Mass: 46637.730 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#3: Protein Envelope protein E / / Coordinate model: Cα atoms only


Mass: 43819.391 Da / Num. of mol.: 3 / Fragment: UNP residues 281-674
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Dengue virus 2 Thailand/16681/84 / Production host: unidentified (others) / References: UniProt: P12823, UniProt: P29990*PLUS
Sequence detailsTHE MODELED SEQUENCE FOR GLYCOPROTEIN DIII IS FROM DENGUE VIRUS 1 (UNP Q88640).

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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
IDNameTypeDetailsParent-ID
1Dengue virus complexed with Fab E104 at pH 5.5COMPLEX1800
2Dengue virus 2VIRUS1
3Fab E1041
Details of virusEmpty: NO / Enveloped: YES / Host category: VERTEBRATES / Isolate: STRAIN / Type: VIRION
Natural hostOrganism: Homo sapiens
Buffer solutionName: 120 mM NaCl, 20 mM Tris HCl, 1 mM EDTA / pH: 5.5 / Details: 120 mM NaCl, 20 mM Tris HCl, 1 mM EDTA
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Details: Plunged into liquid ethane

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS / Date: Jan 1, 2013
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Camera length: 0 mm
Specimen holderSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 24 e/Å2 / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k)
Image scansNum. digital images: 1000
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthRelative weight: 1

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Processing

EM software
IDNameCategory
1EMfitmodel fitting
2EMAN3D reconstruction
SymmetryPoint symmetry: I (icosahedral)
3D reconstructionResolution: 26 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 528 / Nominal pixel size: 5.2 Å / Actual pixel size: 5.2 Å / Details: (Single particle--Applied symmetry: I) / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: RECIPROCAL / Details: REFINEMENT PROTOCOL--rigid body
Atomic model building
IDPDB-ID 3D fitting-IDAccession codeInitial refinement model-IDSource nameType
11OK8

1ok8

11OK81PDBexperimental model
24FFZ

4ffz

14FFZ2PDBexperimental model
Refinement stepCycle: LAST
ProteinNucleic acidLigandSolventTotal
Num. atoms1990 0 0 0 1990

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