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- PDB-5ywp: JEV-2H4 Fab complex -

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Basic information

Entry
Database: PDB / ID: 5ywp
TitleJEV-2H4 Fab complex
Components
  • 2H4 heavy chain
  • 2H4 light chain
  • JEV E protein
  • JEV M protein
KeywordsIMMUNE SYSTEM / Flaviviruses / Viral encephalitis / neutralizing monoclonal antibodies / structural analysis / viral entry
Function / homology
Function and homology information


symbiont-mediated suppression of host JAK-STAT cascade via inhibition of host TYK2 activity / flavivirin / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT2 activity / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT1 activity / double-stranded RNA binding / viral capsid / nucleoside-triphosphate phosphatase / mRNA (guanine-N7)-methyltransferase / methyltransferase cap1 / clathrin-dependent endocytosis of virus by host cell ...symbiont-mediated suppression of host JAK-STAT cascade via inhibition of host TYK2 activity / flavivirin / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT2 activity / symbiont-mediated suppression of host JAK-STAT cascade via inhibition of STAT1 activity / double-stranded RNA binding / viral capsid / nucleoside-triphosphate phosphatase / mRNA (guanine-N7)-methyltransferase / methyltransferase cap1 / clathrin-dependent endocytosis of virus by host cell / mRNA (nucleoside-2'-O-)-methyltransferase activity / host cell surface / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / RNA helicase activity / host cell endoplasmic reticulum membrane / host cell perinuclear region of cytoplasm / protein dimerization activity / symbiont-mediated suppression of host type I interferon-mediated signaling pathway / RNA helicase / induction by virus of host autophagy / RNA-directed RNA polymerase / viral RNA genome replication / serine-type endopeptidase activity / RNA-dependent RNA polymerase activity / virus-mediated perturbation of host defense response / fusion of virus membrane with host endosome membrane / viral envelope / host cell nucleus / virion attachment to host cell / structural molecule activity / virion membrane / ATP hydrolysis activity / proteolysis / extracellular region / ATP binding / membrane / metal ion binding
Similarity search - Function
: / Flavivirus capsid protein C superfamily / RNA-directed RNA polymerase, thumb domain, Flavivirus / Flavivirus RNA-directed RNA polymerase, thumb domain / Flavivirus non-structural protein NS2B / Flavivirus envelope glycoprotein E, stem/anchor domain / : / Flavivirus NS3 helicase, C-terminal helical domain / Genome polyprotein, Flavivirus / Flavivirus non-structural protein NS4A ...: / Flavivirus capsid protein C superfamily / RNA-directed RNA polymerase, thumb domain, Flavivirus / Flavivirus RNA-directed RNA polymerase, thumb domain / Flavivirus non-structural protein NS2B / Flavivirus envelope glycoprotein E, stem/anchor domain / : / 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. / 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, flavivirus / Flavivirus envelope glycoprotein M / 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 / DNA/RNA polymerase superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Genome polyprotein / Genome polyprotein
Similarity search - Component
Biological speciesJapanese encephalitis virus
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.6 Å
AuthorsQiu, X.D. / Lei, Y.F. / Yang, P. / Gao, Q. / WANG, N. / Cao, L. / Yuan, S. / Wang, X.X. / Xu, Z.K. / Rao, Z.H.
Funding support China, 1items
OrganizationGrant numberCountry
National Key Research and Development Program of China2016YFC1200400 China
CitationJournal: Nat Microbiol / Year: 2018
Title: Structural basis for neutralization of Japanese encephalitis virus by two potent therapeutic antibodies.
Authors: Xiaodi Qiu / Yingfeng Lei / Pan Yang / Qiang Gao / Nan Wang / Lei Cao / Shuai Yuan / Xiaofang Huang / Yongqiang Deng / Wenyu Ma / Tianbing Ding / Fanglin Zhang / Xingan Wu / Junjie Hu / Shan- ...Authors: Xiaodi Qiu / Yingfeng Lei / Pan Yang / Qiang Gao / Nan Wang / Lei Cao / Shuai Yuan / Xiaofang Huang / Yongqiang Deng / Wenyu Ma / Tianbing Ding / Fanglin Zhang / Xingan Wu / Junjie Hu / Shan-Lu Liu / Chengfeng Qin / Xiangxi Wang / Zhikai Xu / Zihe Rao /
Abstract: Japanese encephalitis virus (JEV), closely related to dengue, Zika, yellow fever and West Nile viruses, remains neglected and not well characterized . JEV is the leading causative agent of ...Japanese encephalitis virus (JEV), closely related to dengue, Zika, yellow fever and West Nile viruses, remains neglected and not well characterized . JEV is the leading causative agent of encephalitis, and is responsible for thousands of deaths each year in Asia. Humoral immunity is essential for protecting against flavivirus infections and passive immunization has been demonstrated to be effective in curing disease. Here, we demonstrate that JEV-specific monoclonal antibodies, 2F2 and 2H4, block attachment of the virus to its receptor and also prevent fusion of the virus. Neutralization of JEV by these antibodies is exceptionally potent and confers clear therapeutic benefit in mouse models. A single 20 μg dose of these antibodies resulted in 100% survival and complete clearance of JEV from the brains of mice. The 4.7 Å and 4.6 Å resolution cryo-electron microscopy structures of JEV-2F2-Fab and JEV-2H4-Fab complexes, together with the crystal structure of 2H4 Fab and our recent near-atomic structure of JEV , unveil the nature and location of epitopes targeted by the antibodies. Both 2F2 and 2H4 Fabs bind quaternary epitopes that span across three adjacent envelope proteins. Our results provide a structural and molecular basis for the application of 2F2 and 2H4 to treat JEV infection.
History
DepositionNov 29, 2017Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0May 2, 2018Provider: repository / Type: Initial release

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Structure visualization

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Assembly

Deposited unit
A: JEV E protein
B: JEV M protein
C: JEV E protein
D: JEV M protein
E: JEV E protein
F: JEV M protein
M: 2H4 light chain
N: 2H4 heavy chain
Q: 2H4 light chain
R: 2H4 heavy chain


Theoretical massNumber of molelcules
Total (without water)279,63310
Polymers279,63310
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area31980 Å2
ΔGint-181 kcal/mol
Surface area105460 Å2

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Components

#1: Protein JEV E protein


Mass: 53508.684 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Source: (natural) Japanese encephalitis virus / References: UniProt: P32886*PLUS
#2: Protein JEV M protein


Mass: 8250.488 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Source: (natural) Japanese encephalitis virus / References: UniProt: P27395*PLUS
#3: Antibody 2H4 light chain


Mass: 23840.303 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#4: Antibody 2H4 heavy chain


Mass: 23337.305 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)

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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: JEV E protein with its mouse antibody H / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Japanese encephalitis virus / Strain: P3
Source (recombinant)Organism: Chlorocebus aethiops (grivet)
Buffer solutionpH: 7.4 / Details: PBS
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in.
VitrificationCryogen name: ETHANE

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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: 3000 nm / Nominal defocus min: 1000 nm / Calibrated defocus min: 1000 nm / Calibrated defocus max: 3000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm
Image recordingAverage exposure time: 8 sec. / Electron dose: 25 e/Å2 / Detector mode: OTHER / Film or detector model: FEI FALCON III (4k x 4k) / Num. of real images: 2500
Image scansWidth: 4000 / Height: 4000

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Processing

SoftwareName: PHENIX / Version: 1.10.1_2155: / Classification: refinement
EM software
IDNameVersionCategory
1EMANparticle selection
4GctfCTF correction
10EMANinitial Euler assignment
11RELION1.4final Euler assignment
12RELION1.4classification
13RELION1.43D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 26229
3D reconstructionResolution: 4.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 26229 / Num. of class averages: 50 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00520038
ELECTRON MICROSCOPYf_angle_d0.91327253
ELECTRON MICROSCOPYf_dihedral_angle_d8.97915871
ELECTRON MICROSCOPYf_chiral_restr0.0513084
ELECTRON MICROSCOPYf_plane_restr0.0063467

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