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- PDB-3j7g: Electron cryo-microscopy of human papillomavirus 16 and H16.V5 Fa... -

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Basic information

Entry
Database: PDB / ID: 3j7g
TitleElectron cryo-microscopy of human papillomavirus 16 and H16.V5 Fab fragments
ComponentsL1
KeywordsVIRUS / pentamer of human papillomavirus / QV16-V5 complex / virus-Fab complex / neutralization antibody / maturation
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
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 13.6 Å
AuthorsLee, H. / Hafenstein, S.
CitationJournal: J Virol / Year: 2015
Title: A cryo-electron microscopy study identifies the complete H16.V5 epitope and reveals global conformational changes initiated by binding of the neutralizing antibody fragment.
Authors: Hyunwook Lee / Sarah A Brendle / Stephanie M Bywaters / Jian Guan / Robert E Ashley / Joshua D Yoder / Alexander M Makhov / James F Conway / Neil D Christensen / Susan Hafenstein /
Abstract: Human papillomavirus 16 (HPV16) is a worldwide health threat and an etiologic agent of cervical cancer. To understand the antigenic properties of HPV16, we pursued a structural study to elucidate HPV ...Human papillomavirus 16 (HPV16) is a worldwide health threat and an etiologic agent of cervical cancer. To understand the antigenic properties of HPV16, we pursued a structural study to elucidate HPV capsids and antibody interactions. The cryo-electron microscopy (cryo-EM) structures of a mature HPV16 particle and an altered capsid particle were solved individually and as complexes with fragment of antibody (Fab) from the neutralizing antibody H16.V5. Fitted crystal structures provided a pseudoatomic model of the virus-Fab complex, which identified a precise footprint of H16.V5, including previously unrecognized residues. The altered-capsid-Fab complex map showed that binding of the Fab induced significant conformational changes that were not seen in the altered-capsid structure alone. These changes included more ordered surface loops, consolidated so-called "invading-arm" structures, and tighter intercapsomeric connections at the capsid floor. The H16.V5 Fab preferentially bound hexavalent capsomers likely with a stabilizing effect that directly correlated with the number of bound Fabs. Additional cryo-EM reconstructions of the virus-Fab complex for different incubation times and structural analysis provide a model for a hyperstabilization of the capsomer by H16.V5 Fab and showed that the Fab distinguishes subtle differences between antigenic sites.
IMPORTANCE: Our analysis of the cryo-EM reconstructions of the HPV16 capsids and virus-Fab complexes has identified the entire HPV.V5 conformational epitope and demonstrated a detailed neutralization ...IMPORTANCE: Our analysis of the cryo-EM reconstructions of the HPV16 capsids and virus-Fab complexes has identified the entire HPV.V5 conformational epitope and demonstrated a detailed neutralization mechanism of this clinically important monoclonal antibody against HPV16. The Fab bound and ordered the apical loops of HPV16. This conformational change was transmitted to the lower region of the capsomer, resulting in enhanced intercapsomeric interactions evidenced by the more ordered capsid floor and "invading-arm" structures. This study advances the understanding of the neutralization mechanism used by H16.V5.
History
DepositionJun 24, 2014Deposition site: RCSB / Processing site: RCSB
Revision 1.0Nov 26, 2014Provider: repository / Type: Initial release
Revision 1.1Dec 3, 2014Group: Database references
Revision 1.2Mar 18, 2015Group: Database references
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

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
  • Imaged by Jmol
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  • Deposited structure unit
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  • Simplified surface model + fitted atomic model
  • EMDB-5994
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Structure viewerMolecule:
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Assembly

Deposited unit
A: L1
B: L1
C: L1
D: L1
E: L1


Theoretical massNumber of molelcules
Total (without water)254,3675
Polymers254,3675
Non-polymers00
Water0
1
A: L1
B: L1
C: L1
D: L1
E: L1
x 60


Theoretical massNumber of molelcules
Total (without water)15,262,027300
Polymers15,262,027300
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
A: L1
B: L1
C: L1
D: L1
E: L1
x 5


  • icosahedral pentamer
  • 1.27 MDa, 25 polymers
Theoretical massNumber of molelcules
Total (without water)1,271,83625
Polymers1,271,83625
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation4
4
A: L1
B: L1
C: L1
D: L1
E: L1
x 6


  • icosahedral 23 hexamer
  • 1.53 MDa, 30 polymers
Theoretical massNumber of molelcules
Total (without water)1,526,20330
Polymers1,526,20330
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
L1 / L1 protein / Major capsid protein L1


Mass: 50873.422 Da / Num. of mol.: 5 / Fragment: UNP residues 47-500 / Source method: isolated from a natural source / Source: (natural) Human papillomavirus type 16 / References: UniProt: Q4VRM0, 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
IDNameTypeDetailsParent-ID
1Mature HPV16 quasivirus capsid complexed with H16.V5 FabsCOMPLEXThree hundred H16.V5 Fabs bind to one HPV16 capsid0
2Human papillomavirus 16PapillomaviridaeVIRUS1
3H16.V5 Fab1
Molecular weightValue: 42 MDa / Experimental value: NO
Details of virusEmpty: NO / Enveloped: NO / Host category: VERTEBRATES / Isolate: OTHER / Type: VIRION
Natural hostOrganism: Homo sapiens
Buffer solutionName: 137 mM NaCl, 2.7 mM KCl, 10 mM Na2HPO4, 1.8 mM KH2PO4 / pH: 7.4
Details: 137 mM NaCl, 2.7 mM KCl, 10 mM Na2HPO4, 1.8 mM KH2PO4
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: glow-discharged holey carbon Quantifoil grids
VitrificationInstrument: GATAN CRYOPLUNGE 3 / Cryogen name: ETHANE / Temp: 102 K / Humidity: 90 %
Details: Blot for 0.7 seconds before plunging into liquid ethane (GATAN CRYOPLUNGE 3).
Method: Blot for 0.7 seconds before plunging

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Electron microscopy imaging

MicroscopyModel: JEOL 2100 / Date: Oct 30, 2013
Electron gunElectron source: LAB6 / Accelerating voltage: 200 kV / Illumination mode: SPOT SCAN
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 80000 X / Nominal defocus max: 3990 nm / Nominal defocus min: 690 nm / Cs: 2 mm
Specimen holderSpecimen holder model: GATAN LIQUID NITROGEN / Temperature: 95 K / Tilt angle max: 0 ° / Tilt angle min: 0 °
Image recordingElectron dose: 15 e/Å2 / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k)
Image scansNum. digital images: 411
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
CTF correctionDetails: Each particle
SymmetryPoint symmetry: I (icosahedral)
3D reconstructionMethod: Cross-common lines / Resolution: 13.6 Å / Resolution method: FSC 0.5 CUT-OFF / Num. of particles: 2075 / Nominal pixel size: 1.48 Å / Actual pixel size: 1.48 Å
Details: Semi-automatic particle selection was performed using e2boxer.py to obtain the particle coordinates, followed by particle boxing, linearization, normalization, and apodization of the images ...Details: Semi-automatic particle selection was performed using e2boxer.py to obtain the particle coordinates, followed by particle boxing, linearization, normalization, and apodization of the images using Robem. Defocus and astigmatism values to perform contrast transfer function (CTF) correction were assessed using Robem for the extracted particles. The icosahedrally averaged reconstructions were initiated using a random model generated with setup_rmc and reached 14 A resolution estimated at a Fourier Shell Correlation (FSC) of 0.5. For the last step of refinement, the final maps were CTF-corrected using a B factor of 200 A2. (Single particle--Applied symmetry: I)
Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL / Details: REFINEMENT PROTOCOL--flexible
Atomic model buildingPDB-ID: 3OAE

3oae
PDB Unreleased entry


Accession code: 3OAE / Source name: PDB / Type: experimental model
Refinement stepCycle: LAST
ProteinNucleic acidLigandSolventTotal
Num. atoms16610 0 0 0 16610

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