3U6R

Three dimensional structure of broadly neutralizing anti - Hepatitis C virus (HCV) glycoprotein E2 single chain FV fragment 1:7


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.67 Å
  • R-Value Free: 0.232 
  • R-Value Work: 0.190 
  • R-Value Observed: 0.193 

wwPDB Validation   3D Report Full Report


This is version 1.2 of the entry. See complete history


Literature

High-level secretion of recombinant monomeric murine and human single-chain Fv antibodies from Drosophila S2 cells.

Gilmartin, A.A.Lamp, B.Rumenapf, T.Persson, M.A.Rey, F.A.Krey, T.

(2012) Protein Eng Des Sel 25: 59-66

  • DOI: https://doi.org/10.1093/protein/gzr058
  • Primary Citation of Related Structures:  
    3U6R

  • PubMed Abstract: 

    Single-chain variable fragment (scFvs) antibodies are small polypeptides (∼26 kD) containing the heavy (V(H)) and light (V(L)) immunoglobulin domains of a parent antibody connected by a flexible linker. In addition to being frequently used in diagnostics and therapy for an increasing number of human diseases, scFvs are important tools for structural biology as crystallization chaperones. Although scFvs can be expressed in many different organisms, the expression level of an scFv strongly depends on its particular amino acid sequence. We report here a system allowing for easy and efficient cloning of (i) scFvs selected by phage display and (ii) individual heavy and light chain sequences from hybridoma cDNA into expression plasmids engineered for secretion of the recombinant fragment produced in Drosophila S2 cells. We validated the method by producing five scFvs derived from human and murine parent antibodies directed against various antigens. The production yields varied between 5 and 12 mg monomeric scFv per liter of supernatant, indicating a relative independence on the individual sequences. The recombinant scFvs bound their cognate antigen with high affinity, comparable with the parent antibodies. The suitability of the produced recombinant fragments for structural studies was demonstrated by crystallization and structure determination of one of the produced scFvs, derived from a broadly neutralizing antibody against the major glycoprotein E2 of the hepatitis C virus. Structural comparison with the Protein Data Bank revealed the typical spatial organization of V(H) and V(L) domains, further validating the here-reported expression system.


  • Organizational Affiliation

    Départment de Virologie, Institut Pasteur, Unité de Virologie Structurale, CNRS URA 3015, Paris, France.


Macromolecules
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Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Antibody 1:7 (Heavy chain)A [auth H],
C [auth A]
149Homo sapiensMutation(s): 0 
Entity Groups  
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Sequence Annotations
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  • Reference Sequence
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Entity ID: 2
MoleculeChains Sequence LengthOrganismDetailsImage
Antibody 1:7 (Light chain)B [auth L],
D [auth B]
143Homo sapiensMutation(s): 0 
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
Sequence Annotations
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  • Reference Sequence
Small Molecules
Ligands 1 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
SO4
Query on SO4

Download Ideal Coordinates CCD File 
E [auth H]SULFATE ION
O4 S
QAOWNCQODCNURD-UHFFFAOYSA-L
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.67 Å
  • R-Value Free: 0.232 
  • R-Value Work: 0.190 
  • R-Value Observed: 0.193 
  • Space Group: C 2 2 21
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 105.485α = 90
b = 105.744β = 90
c = 105.595γ = 90
Software Package:
Software NamePurpose
XDSdata scaling
PHASERphasing
BUSTERrefinement
XDSdata reduction
SCALAdata scaling

Structure Validation

View Full Validation Report



Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2012-01-11
    Type: Initial release
  • Version 1.1: 2012-10-17
    Changes: Database references
  • Version 1.2: 2023-09-13
    Changes: Data collection, Database references, Derived calculations, Refinement description