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- EMDB-6809: The CryoEM map of HPV58 VLP in complex with the Fab fragment of a... -

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Basic information

Entry
Database: EMDB / ID: 6809
TitleThe CryoEM map of HPV58 VLP in complex with the Fab fragment of antibody A12A3
Map data
SampleHuman papillomavirusHuman papillomavirus infection:
virus
SourceHuman papillomavirus
Methodsingle particle reconstruction / cryo EM / 9.5 Å resolution
AuthorsLi SW / Li ZH
CitationJournal: MBio / Year: 2017
Title: Crystal Structures of Two Immune Complexes Identify Determinants for Viral Infectivity and Type-Specific Neutralization of Human Papillomavirus.
Authors: Zhihai Li / Daning Wang / Ying Gu / Shuo Song / Maozhou He / Jingjie Shi / Xinlin Liu / Shuangping Wei / Jinjin Li / Hai Yu / Qingbing Zheng / Xiaodong Yan / Timothy S Baker / Jun Zhang / Jason S McLellan / Shaowei Li / Ningshao Xia
Abstract: Persistent, high-risk human papillomavirus (HPV) infection is the primary cause of cervical cancer. Neutralizing antibodies elicited by L1-only virus-like particles (VLPs) can block HPV infection; ...Persistent, high-risk human papillomavirus (HPV) infection is the primary cause of cervical cancer. Neutralizing antibodies elicited by L1-only virus-like particles (VLPs) can block HPV infection; however, the lack of high-resolution structures has limited our understanding of the mode of virus infection and the requirement for type specificity at the molecular level. Here, we describe two antibodies, A12A3 and 28F10, that specifically bind to and neutralize HPV58 and HPV59, respectively, through two distinct binding stoichiometries. We show that the epitopes of A12A3 are clustered in the DE loops of two adjacent HPV58 L1 monomers, whereas 28F10 recognizes the HPV59 FG loop of a single monomer. Via structure-based mutagenesis and analysis of antibody binding, we further identified the residues HPV58 D154, S168, and N170 and HPV59 M267, Q270, E273, Y276, K278, and R283, which play critical roles in virus infection. By substituting these strategic epitope residues into other HPV genotypes, we could then redirect the type-specific binding of the antibodies to these genotypes, thus highlighting the importance of these specific residues, HPV58 R161, S168, and N308 and HPV59 Q270, E273, and D281. Overall, our findings provide molecular insights into potential structural determinants of HPV required for infectivity and type specificity. High-risk human papillomaviruses (HPVs) are considered the major causative pathogens of cancers that affect epithelial mucosa, such as cervical cancer. However, because of the lack of high-resolution structural information on the sites of neutralization, we have yet to determine the precise mode of HPV infection and how different types of HPV cause infection. Our crystal structures in this study have uncovered discrete binding stoichiometries for two different antibodies. We show that one A12A3 Fab binds to the center of one HPV58 pentamer, whereas five 28F10 Fabs bind along the top fringe of one HPV59 pentamer. Furthermore, through targeted epitope analysis, we show that 6 to 7 discontinuous residues of the L1 major capsid protein of HPV are determinants, at least in part, for virus infection and type specificity. This knowledge will help us to unravel the process of HPV infection and can potentially be used to drive the development of therapeutics that target neutralization-sensitive sites.
DateDeposition: Aug 10, 2017 / Header (metadata) release: Oct 25, 2017 / Map release: Oct 25, 2017 / Last update: Oct 25, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 2
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 2
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_6809.map.gz (map file in CCP4 format, 296353 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
420 pix
2.22 Å/pix.
= 931.56 Å
420 pix
2.22 Å/pix.
= 931.56 Å
420 pix
2.22 Å/pix.
= 931.56 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.218 Å
Density
Contour Level:2 (by author), 2 (movie #1):
Minimum - Maximum-7.2638855 - 11.711547
Average (Standard dev.)-3.7027E-10 (1)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions420420420
Origin555
Limit424424424
Spacing420420420
CellA=B=C: 931.56 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.2182.2182.218
M x/y/z420420420
origin x/y/z0.0000.0000.000
length x/y/z931.560931.560931.560
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS555
NC/NR/NS420420420
D min/max/mean-7.26411.712-0.000

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Supplemental data

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Sample components

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Entire Human papillomavirus

EntireName: Human papillomavirus
Details: VLP generated by recombinantly expressed; Fab Fragment generated by proteolytic cleavage of IgG antibody;
Number of components: 1

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Component #1: virus, Human papillomavirus

VirusName: Human papillomavirusHuman papillomavirus infection / Class: VIRUS-LIKE PARTICLE
Details: VLP generated by recombinantly expressed; Fab Fragment generated by proteolytic cleavage of IgG antibody;
Empty: Yes / Enveloped: No / Isolate: OTHER
SpeciesSpecies: Human papillomavirus
Source (engineered)Expression System: Escherichia coli (E. coli)

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionpH: 6.5
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Tecnai F30 / Image courtesy: FEI Company
ImagingMicroscope: FEI TECNAI F30
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 25 e/Å2 / Illumination mode: SPOT SCAN
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: FEI FALCON I (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 3600
3D reconstructionResolution: 9.5 Å / Resolution method: FSC 0.143 CUT-OFF

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