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- PDB-3jba: The U4 antibody epitope on human papillomavirus 16 identified by ... -

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Basic information

Entry
Database: PDB / ID: 3jba
TitleThe U4 antibody epitope on human papillomavirus 16 identified by cryo-EM
Components
  • H16.U4 antibody heavy chain
  • H16.U4 antibody light chain
  • Major capsid protein L1
KeywordsVIRUS/IMMUNE SYSTEM / HPV16 / antibody / U4 / neutralization / Fab / VIRUS-IMMUNE SYSTEM complex
Function / homology
Function and homology information


T=7 icosahedral viral capsid / endocytosis involved in viral entry into host cell / host cell nucleus / structural molecule activity / virion attachment to host cell
Similarity search - Function
Major capsid L1 (late) protein, Papillomavirus / Major capsid L1 (late) superfamily, Papillomavirus / L1 (late) protein / Double-stranded DNA virus, group I, capsid
Similarity search - Domain/homology
Major capsid protein L1 / Major capsid protein L1
Similarity search - Component
Biological speciesHuman papillomavirus type 16
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 12 Å
AuthorsGuan, J. / Bywaters, S.M. / Brendle, S.A. / Lee, H. / Ashley, R.E. / Christensen, N.D. / Hafenstein, S.
CitationJournal: J Virol / Year: 2015
Title: The U4 Antibody Epitope on Human Papillomavirus 16 Identified by Cryo-electron Microscopy.
Authors: Jian Guan / Stephanie M Bywaters / Sarah A Brendle / Hyunwook Lee / Robert E Ashley / Neil D Christensen / Susan Hafenstein /
Abstract: The human papillomavirus (HPV) major structural protein L1 composes capsomers that are linked together through interactions mediated by the L1 C terminus to constitute a T=7 icosahedral capsid. H16. ...The human papillomavirus (HPV) major structural protein L1 composes capsomers that are linked together through interactions mediated by the L1 C terminus to constitute a T=7 icosahedral capsid. H16.U4 is a type-specific monoclonal antibody recognizing a conformation-dependent neutralizing epitope of HPV thought to include the L1 protein C terminus. The structure of human papillomavirus 16 (HPV16) complexed with H16.U4 fragments of antibody (Fab) was solved by cryo-electron microscopy (cryo-EM) image reconstruction. Atomic structures of virus and Fab were fitted into the corresponding cryo-EM densities to identify the antigenic epitope. The antibody footprint mapped predominately to the L1 C-terminal arm with an additional contact point on the side of the capsomer. This footprint describes an epitope that is presented capsid-wide. However, although the H16.U4 epitope suggests the presence of 360 potential binding sites exposed in the capsid valley between each capsomer, H16.U4 Fab bound only to epitopes located around the icosahedral five-fold vertex of the capsid. Thus, the binding characteristics of H16.U4 defined in this study showed a distinctive selectivity for local conformation-dependent interactions with specific L1 invading arms between five-fold related capsomers.
IMPORTANCE: Human papillomavirus 16 (HPV16) is the most prevalent oncogenic genotype in HPV-associated anogenital and oral cancers. Here we use cryo-EM reconstruction techniques to solve the ...IMPORTANCE: Human papillomavirus 16 (HPV16) is the most prevalent oncogenic genotype in HPV-associated anogenital and oral cancers. Here we use cryo-EM reconstruction techniques to solve the structures of the HPV16 capsid complexes using H16.U4 fragment of antibody (Fab). Different from most other antibodies directed against surface loops, H16.U4 monoclonal antibody is unique in targeting the C-terminal arm of the L1 protein. This monoclonal antibody (MAb) is used throughout the HPV research community in HPV serological and vaccine development and to define mechanisms of HPV uptake. The unique binding mode of H16.U4 defined here shows important conformation-dependent interactions within the HPV16 capsid. By targeting an important structural and conformational epitope, H16.U4 may identify subtle conformational changes in different maturation stages of the HPV capsid and provide a key probe to analyze the mechanisms of HPV uptake during the early stages of virus infection. Our analyses precisely define important conformational epitopes on HPV16 capsids that are key targets for successful HPV prophylactic vaccines.
History
DepositionAug 11, 2015Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 7, 2015Provider: repository / Type: Initial release
Revision 1.1Nov 18, 2015Group: Database references
Revision 1.2Jul 18, 2018Group: Data collection / Category: em_software / Item: _em_software.image_processing_id / _em_software.name

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Structure visualization

Movie
  • 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-6424
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  • Superimposition on EM map
  • EMDB-6424
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Structure viewerMolecule:
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Assembly

Deposited unit
L: H16.U4 antibody light chain
H: H16.U4 antibody heavy chain
B: Major capsid protein L1
A: Major capsid protein L1
E: Major capsid protein L1
D: Major capsid protein L1
C: Major capsid protein L1
F: Major capsid protein L1


Theoretical massNumber of molelcules
Total (without water)346,0848
Polymers346,0848
Non-polymers00
Water0
1
L: H16.U4 antibody light chain
H: H16.U4 antibody heavy chain
B: Major capsid protein L1
A: Major capsid protein L1
E: Major capsid protein L1
D: Major capsid protein L1
C: Major capsid protein L1
F: Major capsid protein L1
x 60


Theoretical massNumber of molelcules
Total (without water)20,765,039480
Polymers20,765,039480
Non-polymers00
Water0
TypeNameSymmetry operationNumber
point symmetry operation60
2


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit
TypeNameSymmetry operationNumber
point symmetry operation1
3
L: H16.U4 antibody light chain
H: H16.U4 antibody heavy chain
B: Major capsid protein L1
A: Major capsid protein L1
E: Major capsid protein L1
D: Major capsid protein L1
C: Major capsid protein L1
F: Major capsid protein L1
x 5


  • icosahedral pentamer
  • 1.73 MDa, 40 polymers
Theoretical massNumber of molelcules
Total (without water)1,730,42040
Polymers1,730,42040
Non-polymers00
Water0
TypeNameSymmetry operationNumber
point symmetry operation5
4
L: H16.U4 antibody light chain
H: H16.U4 antibody heavy chain
B: Major capsid protein L1
A: Major capsid protein L1
E: Major capsid protein L1
D: Major capsid protein L1
C: Major capsid protein L1
F: Major capsid protein L1
x 6


  • icosahedral 23 hexamer
  • 2.08 MDa, 48 polymers
Theoretical massNumber of molelcules
Total (without water)2,076,50448
Polymers2,076,50448
Non-polymers00
Water0
TypeNameSymmetry operationNumber
point symmetry operation6
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: Antibody H16.U4 antibody light chain


Mass: 12699.254 Da / Num. of mol.: 1 / Fragment: Fab variable domain / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#2: Antibody H16.U4 antibody heavy chain


Mass: 12711.025 Da / Num. of mol.: 1 / Fragment: Fab variable domain / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse)
#3: Protein
Major capsid protein L1


Mass: 53445.617 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human papillomavirus type 16 / Gene: L1 / Plasmid: SV40 / Cell line (production host): HEK293TT / Production host: Homo sapiens (human) / References: UniProt: C9E771, UniProt: P03101*PLUS

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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
IDNameTypeParent-ID
1Quasi-HPV16 complexed with H16.U4 FabCOMPLEX0
2Human papillomavirusHuman papillomavirus infectionVIRUS1
3H16.U4 antibody1
Molecular weightValue: 32.8 MDa / Experimental value: NO
Details of virusEmpty: NO / Enveloped: NO / Host category: VERTEBRATES / Isolate: SEROTYPE / Type: VIRUS-LIKE PARTICLE
Natural hostOrganism: Homo sapiens
Buffer solutionName: 1 M NaCl, 200 mM Tris / pH: 7.4 / Details: 1 M NaCl, 200 mM Tris
SpecimenConc.: 1.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: glow-discharged holey carbon support grid
VitrificationInstrument: GATAN CRYOPLUNGE 3 / Cryogen name: ETHANE / Temp: 102 K / Humidity: 90 % / Details: Plunged into liquid ethane (GATAN CRYOPLUNGE 3).

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Electron microscopy imaging

MicroscopyModel: JEOL 2100 / Date: Jul 22, 2014
Electron gunElectron source: LAB6 / Accelerating voltage: 200 kV / Illumination mode: SPOT SCAN
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 50000 X / Nominal defocus max: 5430 nm / Nominal defocus min: 1600 nm / Cs: 2 mm
Specimen holderSpecimen holder model: GATAN LIQUID NITROGEN / Temperature: 95 K
Image recordingElectron dose: 15 e/Å2 / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k)
Image scansNum. digital images: 151
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthRelative weight: 1

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Processing

EM software
IDNameVersionCategory
1Situsmodel fitting
2UCSF Chimeramodel fitting
3Auto3DEM3D reconstruction
4EMAN23D reconstruction
5RELION3D reconstruction
CTF correctionDetails: each particle
SymmetryPoint symmetry: I (icosahedral)
3D reconstructionMethod: Cross-common lines / Resolution: 12 Å / Resolution method: FSC 0.5 CUT-OFF / Num. of particles: 5806 / Nominal pixel size: 2.33 Å / Actual pixel size: 2.33 Å
Details: (Single particle details: The particles were selected using semi-automatic program e2boxer.py.) (Single particle--Applied symmetry: I)
Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL / Details: REFINEMENT PROTOCOL--rigid body
Atomic model buildingPDB-ID: 3J6R

3j6r


Pdb chain-ID: F
Refinement stepCycle: LAST
ProteinNucleic acidLigandSolventTotal
Num. atoms13253 0 0 0 13253

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