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- EMDB-8339: HCV E2 bound to AR1B, AR2A, and HCV1 Fabs -

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Basic information

Entry
Database: EMDB / ID: EMD-8339
TitleHCV E2 bound to AR1B, AR2A, and HCV1 Fabs
Map dataSecond RCT reconstruction of HCV E2 in complex with three antibodies
Sample
  • Complex: HCV E2 bound to 3 different neutralizing fragments antigen binding (Fabs)
    • Complex: HCV E2 protein
    • Complex: AR1B, AR2A, and HCV1 Fabs
KeywordsHCV / E2 / neutralizing antibodies / VIRAL PROTEIN-IMMUNE SYSTEM complex
Biological speciesHepatitis C virus / Homo sapiens (human)
Methodsingle particle reconstruction / negative staining / Resolution: 30.0 Å
AuthorsWard AB / Nieusma T
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious DiseasesAI79031 United States
National Institutes of Health/National Institute Of Allergy and Infectious DiseasesAI106005 United States
National Institutes of Health/National Institute Of Allergy and Infectious DiseasesAI123365 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2016
Title: Structural flexibility at a major conserved antibody target on hepatitis C virus E2 antigen.
Authors: Leopold Kong / David E Lee / Rameshwar U Kadam / Tong Liu / Erick Giang / Travis Nieusma / Fernando Garces / Netanel Tzarum / Virgil L Woods / Andrew B Ward / Sheng Li / Ian A Wilson / Mansun Law /
Abstract: Hepatitis C virus (HCV) is a major cause of liver disease, affecting over 2% of the world's population. The HCV envelope glycoproteins E1 and E2 mediate viral entry, with E2 being the main target of ...Hepatitis C virus (HCV) is a major cause of liver disease, affecting over 2% of the world's population. The HCV envelope glycoproteins E1 and E2 mediate viral entry, with E2 being the main target of neutralizing antibody responses. Structural investigations of E2 have produced templates for vaccine design, including the conserved CD81 receptor-binding site (CD81bs) that is a key target of broadly neutralizing antibodies (bNAbs). Unfortunately, immunization with recombinant E2 and E1E2 rarely elicits sufficient levels of bNAbs for protection. To understand the challenges for eliciting bNAb responses against the CD81bs, we investigated the E2 CD81bs by electron microscopy (EM), hydrogen-deuterium exchange (HDX), molecular dynamics (MD), and calorimetry. By EM, we observed that HCV1, a bNAb recognizing the N-terminal region of the CD81bs, bound a soluble E2 core construct from multiple angles of approach, suggesting components of the CD81bs are flexible. HDX of multiple E2 constructs consistently indicated the entire CD81bs was flexible relative to the rest of the E2 protein, which was further confirmed by MD simulations. However, E2 has a high melting temperature of 84.8 °C, which is more akin to proteins from thermophilic organisms. Thus, recombinant E2 is a highly stable protein overall, but with an exceptionally flexible CD81bs. Such flexibility may promote induction of nonneutralizing antibodies over bNAbs to E2 CD81bs, underscoring the necessity of rigidifying this antigenic region as a target for rational vaccine design.
History
DepositionAug 16, 2016-
Header (metadata) releaseAug 31, 2016-
Map releaseOct 26, 2016-
UpdateSep 6, 2023-
Current statusSep 6, 2023Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3.69
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 3.69
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_8339.png

Downloads & links

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Map

FileDownload / File: emd_8339.map.gz / Format: CCP4 / Size: 2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSecond RCT reconstruction of HCV E2 in complex with three antibodies
Voxel sizeX=Y=Z: 4.35 Å
Density
Contour LevelBy AUTHOR: 3.69 / Movie #1: 3.69
Minimum - Maximum-8.729169000000001 - 13.494557
Average (Standard dev.)0.000000005939592 (±0.61436117)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-40-40-40
Dimensions808080
Spacing808080
CellA=B=C: 348.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.354.354.35
M x/y/z808080
origin x/y/z0.0000.0000.000
length x/y/z348.000348.000348.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ416416416
MAP C/R/S123
start NC/NR/NS-40-40-40
NC/NR/NS808080
D min/max/mean-8.72913.4950.000

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Supplemental data

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Sample components

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Entire : HCV E2 bound to 3 different neutralizing fragments antigen bindin...

EntireName: HCV E2 bound to 3 different neutralizing fragments antigen binding (Fabs)
Components
  • Complex: HCV E2 bound to 3 different neutralizing fragments antigen binding (Fabs)
    • Complex: HCV E2 protein
    • Complex: AR1B, AR2A, and HCV1 Fabs

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Supramolecule #1: HCV E2 bound to 3 different neutralizing fragments antigen bindin...

SupramoleculeName: HCV E2 bound to 3 different neutralizing fragments antigen binding (Fabs)
type: complex / ID: 1 / Parent: 0
Molecular weightTheoretical: 50 KDa

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Supramolecule #2: HCV E2 protein

SupramoleculeName: HCV E2 protein / type: complex / ID: 2 / Parent: 1
Source (natural)Organism: Hepatitis C virus

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Supramolecule #3: AR1B, AR2A, and HCV1 Fabs

SupramoleculeName: AR1B, AR2A, and HCV1 Fabs / type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Homo sapiens (human)

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.001 mg/mL
BufferpH: 7.5
StainingType: NEGATIVE / Material: uranyl formate

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Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Average electron dose: 25.0 e/Å2
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

Startup modelType of model: RANDOM CONICAL TILT / Random conical tilt - Tilt angle: 55 degrees
Initial angle assignmentType: NOT APPLICABLE / Software - Name: EMAN (ver. 1)
Final angle assignmentType: NOT APPLICABLE / Software - Name: EMAN (ver. 1)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 30.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: EMAN (ver. 1) / Number images used: 6364

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