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- PDB-8tfe: Crystal structure of Fab fragment of anti-HCV E2 antibody (CBH-7) -

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Basic information

Entry
Database: PDB / ID: 8tfe
TitleCrystal structure of Fab fragment of anti-HCV E2 antibody (CBH-7)
Components
  • CBH-7 Heavy chain
  • CBH-7 Light chain
KeywordsIMMUNE SYSTEM / Antibody / E2 glycoprotein / Fab fragment
Function / homologyCITRATE ANION
Function and homology information
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.62 Å
AuthorsShahid, S. / Mariuzza, R.A.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)R01 AI132213 United States
CitationJournal: Commun Biol / Year: 2025
Title: Cryo-EM structures of HCV E2 glycoprotein bound to neutralizing and non-neutralizing antibodies determined using bivalent Fabs as fiducial markers.
Authors: Salman Shahid / Sharanbasappa S Karade / S Saif Hasan / Rui Yin / Liqun Jiang / Yanxin Liu / Nathaniel Felbinger / Liudmila Kulakova / Eric A Toth / Zhen-Yong Keck / Steven K H Foung / ...Authors: Salman Shahid / Sharanbasappa S Karade / S Saif Hasan / Rui Yin / Liqun Jiang / Yanxin Liu / Nathaniel Felbinger / Liudmila Kulakova / Eric A Toth / Zhen-Yong Keck / Steven K H Foung / Thomas R Fuerst / Brian G Pierce / Roy A Mariuzza /
Abstract: Global elimination of hepatitis C virus (HCV) will require an effective cross-genotype vaccine. The HCV E2 envelope glycoprotein is the main target of neutralizing antibodies but also contains ...Global elimination of hepatitis C virus (HCV) will require an effective cross-genotype vaccine. The HCV E2 envelope glycoprotein is the main target of neutralizing antibodies but also contains epitopes that elicit non-neutralizing antibodies which may provide protection through Fc effector functions rather than direct neutralization. We determined cryo-EM structures of a broadly neutralizing antibody, a moderately neutralizing antibody, and a non-neutralizing antibody bound to E2 to resolutions of 3.8, 3.3, and 3.7 Å, respectively. Whereas the broadly neutralizing antibody targeted the front layer of E2 and the non-neutralizing antibody targeted the back layer, the moderately neutralizing antibody straddled both front and back layers, and thereby defined a new neutralizing epitope on E2. The small size of complexes between conventional (monovalent) Fabs and E2 (~110 kDa) presented a challenge for cryo-EM. Accordingly, we engineered bivalent versions of E2-specific Fabs that doubled the size of Fab-E2 complexes and conferred highly identifiable shapes to the complexes that facilitated particle selection and orientation for image processing. This study validates bivalent Fabs as new fiducial markers for cryo-EM analysis of small proteins such as HCV E2 and identifies a new target epitope for vaccine development.
History
DepositionJul 10, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 17, 2024Provider: repository / Type: Initial release
Revision 1.1Nov 13, 2024Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature / Item: _pdbx_entry_details.has_protein_modification
Revision 1.2Jul 9, 2025Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: CBH-7 Heavy chain
B: CBH-7 Light chain
C: CBH-7 Heavy chain
D: CBH-7 Light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)106,2289
Polymers105,7914
Non-polymers4375
Water14,088782
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)79.935, 101.912, 109.936
Angle α, β, γ (deg.)90.00, 100.24, 90.00
Int Tables number5
Space group name H-MC121

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Components

#1: Antibody CBH-7 Heavy chain


Mass: 27417.719 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell (production host): HEK293 / Cell line (production host): HEK293 / Production host: Homo sapiens (human)
#2: Antibody CBH-7 Light chain


Mass: 25477.658 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell (production host): HEK293 / Cell line (production host): HEK293 / Production host: Homo sapiens (human)
#3: Chemical
ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL


Mass: 62.068 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C2H6O2 / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-FLC / CITRATE ANION


Mass: 189.100 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H5O7 / Feature type: SUBJECT OF INVESTIGATION
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 782 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.08 Å3/Da / Density % sol: 40.94 %
Crystal growTemperature: 295 K / Method: vapor diffusion, hanging drop
Details: 0.1M Sodium phosphate dibasic pH 4.2, 0.2 M NaCl, and 20% PEG8000

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 23-ID-B / Wavelength: 0.979 Å
DetectorType: DECTRIS EIGER X 16M / Detector: PIXEL / Date: Oct 7, 2021
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.979 Å / Relative weight: 1
ReflectionResolution: 1.62→50 Å / Num. obs: 107383 / % possible obs: 98.4 % / Redundancy: 6.4 % / Biso Wilson estimate: 19.42 Å2 / CC1/2: 0.981 / CC star: 0.995 / Rmerge(I) obs: 0.09 / Rpim(I) all: 0.038 / Rrim(I) all: 0.098 / Net I/σ(I): 27.75
Reflection shellResolution: 1.62→1.65 Å / Redundancy: 5.6 % / Rmerge(I) obs: 0.544 / Num. unique obs: 5314 / CC1/2: 0.911 / CC star: 0.976 / Rpim(I) all: 0.238 / Rrim(I) all: 0.595 / % possible all: 98.5

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Processing

Software
NameVersionClassification
REFMAC5.8.0267refinement
HKL-3000data reduction
SCALAdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 6X9X

6x9x


Resolution: 1.62→46.14 Å / Cor.coef. Fo:Fc: 0.966 / Cor.coef. Fo:Fc free: 0.951 / SU B: 2.222 / SU ML: 0.074 / Cross valid method: THROUGHOUT / ESU R: 0.093 / ESU R Free: 0.095 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflectionSelection details
Rfree0.21384 5362 5 %RANDOM
Rwork0.17695 ---
obs0.17881 102019 98.13 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso mean: 23.222 Å2
Baniso -1Baniso -2Baniso -3
1-2.84 Å2-0 Å2-0.37 Å2
2---1.2 Å2-0 Å2
3----1.41 Å2
Refinement stepCycle: 1 / Resolution: 1.62→46.14 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6449 0 29 782 7260
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0110.0136704
X-RAY DIFFRACTIONr_bond_other_d0.0010.0176087
X-RAY DIFFRACTIONr_angle_refined_deg1.6491.6419145
X-RAY DIFFRACTIONr_angle_other_deg1.441.56914091
X-RAY DIFFRACTIONr_dihedral_angle_1_deg7.3155885
X-RAY DIFFRACTIONr_dihedral_angle_2_deg33.10622.878271
X-RAY DIFFRACTIONr_dihedral_angle_3_deg13.626151038
X-RAY DIFFRACTIONr_dihedral_angle_4_deg20.7941526
X-RAY DIFFRACTIONr_chiral_restr0.0740.2906
X-RAY DIFFRACTIONr_gen_planes_refined0.0090.027687
X-RAY DIFFRACTIONr_gen_planes_other0.0010.021501
X-RAY DIFFRACTIONr_nbd_refined
X-RAY DIFFRACTIONr_nbd_other
X-RAY DIFFRACTIONr_nbtor_refined
X-RAY DIFFRACTIONr_nbtor_other
X-RAY DIFFRACTIONr_xyhbond_nbd_refined
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_metal_ion_refined
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined
X-RAY DIFFRACTIONr_symmetry_vdw_other
X-RAY DIFFRACTIONr_symmetry_hbond_refined
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_symmetry_metal_ion_refined
X-RAY DIFFRACTIONr_symmetry_metal_ion_other
X-RAY DIFFRACTIONr_mcbond_it2.0772.2843498
X-RAY DIFFRACTIONr_mcbond_other2.0752.2833497
X-RAY DIFFRACTIONr_mcangle_it2.9323.4194376
X-RAY DIFFRACTIONr_mcangle_other2.9333.424377
X-RAY DIFFRACTIONr_scbond_it3.022.5233206
X-RAY DIFFRACTIONr_scbond_other3.0212.5233205
X-RAY DIFFRACTIONr_scangle_it
X-RAY DIFFRACTIONr_scangle_other4.4633.6654763
X-RAY DIFFRACTIONr_long_range_B_refined6.07127.8697173
X-RAY DIFFRACTIONr_long_range_B_other6.07127.8717174
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 1.62→1.664 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.291 357 -
Rwork0.255 7323 -
obs--95.01 %

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