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- PDB-8rh0: Trimeric HSV-1F gB ectodomain in postfusion conformation with thr... -

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Basic information

Entry
Database: PDB / ID: 8rh0
TitleTrimeric HSV-1F gB ectodomain in postfusion conformation with three bound HDIT102 Fab molecules.
Components
  • (HDIT102 Fab heavy chain) x 2
  • Envelope glycoprotein B
KeywordsVIRAL PROTEIN / ectodomain / post-fusion / fab molecule / trimeric
Function / homology
Function and homology information


host cell Golgi membrane / host cell endosome membrane / symbiont entry into host cell / viral envelope / virion attachment to host cell / host cell plasma membrane / virion membrane / membrane
Similarity search - Function
Herpesvirus Glycoprotein B ectodomain / Herpesvirus Glycoprotein B / Herpesvirus Glycoprotein B, PH-like domain 1 / Herpesvirus Glycoprotein B, PH-like domain 2 / Herpesvirus Glycoprotein B / Herpesvirus Glycoprotein B PH-like domain / Herpesvirus Glycoprotein B, PH-like domain 2 superfamily
Similarity search - Domain/homology
Envelope glycoprotein B
Similarity search - Component
Biological speciesHuman alphaherpesvirus 1 (Herpes simplex virus type 1)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.44 Å
AuthorsKalbermatter, D. / Seyfizadeh, N. / Imhof, T. / Ries, M. / Mueller, C. / Jenner, L. / Blumenschein, E. / Yendrzheyevskiy, A. / Moog, K. / Eckert, D. ...Kalbermatter, D. / Seyfizadeh, N. / Imhof, T. / Ries, M. / Mueller, C. / Jenner, L. / Blumenschein, E. / Yendrzheyevskiy, A. / Moog, K. / Eckert, D. / Engel, R. / Diebolder, P. / Chami, M. / Krauss, J. / Schaller, T. / Arndt, M.
Funding support1items
OrganizationGrant numberCountry
Other private
CitationJournal: J Biomed Sci / Year: 2024
Title: Development of a highly effective combination monoclonal antibody therapy against Herpes simplex virus.
Authors: Narges Seyfizadeh / David Kalbermatter / Thomas Imhof / Moritz Ries / Christian Müller / Leonie Jenner / Elisabeth Blumenschein / Alexandra Yendrzheyevskiy / Frank Grün / Kevin Moog / ...Authors: Narges Seyfizadeh / David Kalbermatter / Thomas Imhof / Moritz Ries / Christian Müller / Leonie Jenner / Elisabeth Blumenschein / Alexandra Yendrzheyevskiy / Frank Grün / Kevin Moog / Daniel Eckert / Ronja Engel / Philipp Diebolder / Mohamed Chami / Jürgen Krauss / Torsten Schaller / Michaela Arndt /
Abstract: BACKGROUND: Infections with Herpes simplex virus (HSV)-1 or -2 usually present as mild chronic recurrent disease, however in rare cases can result in life-threatening conditions with a large spectrum ...BACKGROUND: Infections with Herpes simplex virus (HSV)-1 or -2 usually present as mild chronic recurrent disease, however in rare cases can result in life-threatening conditions with a large spectrum of pathology. Monoclonal antibody therapy has great potential especially to treat infections with virus resistant to standard therapies. HDIT101, a humanized IgG targeting HSV-1/2 gB was previously investigated in phase 2 clinical trials. The aim of this study was to develop a next-generation therapy by combining different antiviral monoclonal antibodies.
METHODS: A lymph-node derived phage display library (LYNDAL) was screened against recombinant gB from Herpes simplex virus (HSV) -1 and HDIT102 scFv was selected for its binding characteristics using ...METHODS: A lymph-node derived phage display library (LYNDAL) was screened against recombinant gB from Herpes simplex virus (HSV) -1 and HDIT102 scFv was selected for its binding characteristics using bio-layer interferometry. HDIT102 was further developed as fully human IgG and tested alone or in combination with HDIT101, a clinically tested humanized anti-HSV IgG, in vitro and in vivo. T-cell stimulating activities by antigen-presenting cells treated with IgG-HSV immune complexes were analyzed using primary human cells. To determine the epitopes, the cryo-EM structures of HDIT101 or HDIT102 Fab bound to HSV-1F as well as HSV-2G gB protein were solved at resolutions < 3.5 Å.
RESULTS: HDIT102 Fab showed strong binding to HSV-1F gB with Kd of 8.95 × 10 M and to HSV-2G gB with Kd of 3.29 × 10 M. Neutralization of cell-free virus and inhibition of cell-to-cell ...RESULTS: HDIT102 Fab showed strong binding to HSV-1F gB with Kd of 8.95 × 10 M and to HSV-2G gB with Kd of 3.29 × 10 M. Neutralization of cell-free virus and inhibition of cell-to-cell spread were comparable between HDIT101 and HDIT102. Both antibodies induced internalization of gB from the cell surface into acidic endosomes by binding distinct epitopes in domain I of gB and compete for binding. CryoEM analyses revealed the ability to form heterogenic immune complexes consisting of two HDIT102 and one HDIT101 Fab bound to one gB trimeric molecule. Both antibodies mediated antibody-dependent phagocytosis by antigen presenting cells which stimulated autologous T-cell activation. In vivo, the combination of HDIT101 and HDIT102 demonstrated synergistic effects on survival and clinical outcome in immunocompetent BALB/cOlaHsd mice.
CONCLUSION: This biochemical and immunological study showcases the potential of an effective combination therapy with two monoclonal anti-gB IgGs for the treatment of HSV-1/2 induced disease conditions.
History
DepositionDec 14, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 19, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Envelope glycoprotein B
B: Envelope glycoprotein B
C: Envelope glycoprotein B
H: HDIT102 Fab heavy chain
L: HDIT102 Fab heavy chain
E: HDIT102 Fab heavy chain
G: HDIT102 Fab heavy chain
D: HDIT102 Fab heavy chain
F: HDIT102 Fab heavy chain


Theoretical massNumber of molelcules
Total (without water)516,4919
Polymers516,4919
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein Envelope glycoprotein B / gB


Mass: 100449.375 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human alphaherpesvirus 1 (Herpes simplex virus type 1)
Gene: gB, UL27 / Cell line (production host): HEK293-E6 / Production host: Homo sapiens (human) / References: UniProt: P06436
#2: Antibody HDIT102 Fab heavy chain


Mass: 49310.387 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293-E6 / Production host: Homo sapiens (human)
#3: Antibody HDIT102 Fab heavy chain


Mass: 22403.775 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293-E6 / Production host: Homo sapiens (human)

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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

ComponentName: Trimeric HSV-1F gB ectodomain in post-fusion conformation with three bound HDIT102 Fab molecules
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.523 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293-E6
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
1150 mMSodium ChlorideNaCl1
250 mMTrisC4H11NO31
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 85 % / Chamber temperature: 283.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm
Image recordingAverage exposure time: 4 sec. / Electron dose: 1.15 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 6611

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Processing

EM software
IDNameVersionCategory
1RELION4particle selection
2SerialEM3.8image acquisition
4CTFFIND4.1.14CTF correction
7CCP4 package1.6model fitting
9RELION4initial Euler assignment
10RELION4final Euler assignment
11RELION4classification
12RELION43D reconstruction
13CCP4 package1.6model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1059496
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.44 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 208280 / Algorithm: BACK PROJECTION / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT
Atomic model buildingPDB-ID: 2gum

2gum


Accession code: 2gum / Source name: PDB / Type: experimental model
RefinementResolution: 3.44→192.45 Å / Cor.coef. Fo:Fc: 0.848 / SU B: 18.753 / SU ML: 0.297 / ESU R: 0.347
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflection
Rwork0.36172 --
obs0.36172 322272 100 %
Solvent computationSolvent model: PARAMETERS FOR MASK CACLULATION
Displacement parametersBiso mean: 26.672 Å2
Refinement stepCycle: 1 / Total: 19143
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
ELECTRON MICROSCOPYr_bond_refined_d0.0060.01119629
ELECTRON MICROSCOPYr_bond_other_d0.0120.01617112
ELECTRON MICROSCOPYr_angle_refined_deg1.1061.65226688
ELECTRON MICROSCOPYr_angle_other_deg0.5371.5539825
ELECTRON MICROSCOPYr_dihedral_angle_1_deg4.55752409
ELECTRON MICROSCOPYr_dihedral_angle_2_deg3.91510180
ELECTRON MICROSCOPYr_dihedral_angle_3_deg15.053103078
ELECTRON MICROSCOPYr_dihedral_angle_4_deg
ELECTRON MICROSCOPYr_chiral_restr
ELECTRON MICROSCOPYr_gen_planes_refined0.0040.0222890
ELECTRON MICROSCOPYr_gen_planes_other0.0020.024218
ELECTRON MICROSCOPYr_nbd_refined
ELECTRON MICROSCOPYr_nbd_other
ELECTRON MICROSCOPYr_nbtor_refined
ELECTRON MICROSCOPYr_nbtor_other
ELECTRON MICROSCOPYr_xyhbond_nbd_refined
ELECTRON MICROSCOPYr_xyhbond_nbd_other
ELECTRON MICROSCOPYr_metal_ion_refined
ELECTRON MICROSCOPYr_metal_ion_other
ELECTRON MICROSCOPYr_symmetry_vdw_refined
ELECTRON MICROSCOPYr_symmetry_vdw_other
ELECTRON MICROSCOPYr_symmetry_hbond_refined
ELECTRON MICROSCOPYr_symmetry_hbond_other
ELECTRON MICROSCOPYr_symmetry_metal_ion_refined
ELECTRON MICROSCOPYr_symmetry_metal_ion_other
ELECTRON MICROSCOPYr_mcbond_it1.972.6489672
ELECTRON MICROSCOPYr_mcbond_other1.9692.6489672
ELECTRON MICROSCOPYr_mcangle_it3.3293.96612069
ELECTRON MICROSCOPYr_mcangle_other3.3293.96612070
ELECTRON MICROSCOPYr_scbond_it2.1762.8369957
ELECTRON MICROSCOPYr_scbond_other2.1762.8369958
ELECTRON MICROSCOPYr_scangle_it
ELECTRON MICROSCOPYr_scangle_other3.7714.14914620
ELECTRON MICROSCOPYr_long_range_B_refined6.90348.37572359
ELECTRON MICROSCOPYr_long_range_B_other6.90348.37572360
ELECTRON MICROSCOPYr_rigid_bond_restr
ELECTRON MICROSCOPYr_sphericity_free
ELECTRON MICROSCOPYr_sphericity_bonded
LS refinement shellResolution: 3.44→3.529 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0 0 -
Rwork0.653 23800 -
obs--100 %

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