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PDBsum entry 3g5y

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protein Protein-protein interface(s) links
Immune system PDB id
3g5y
Jmol
Contents
Protein chains
213 a.a. *
210 a.a. *
16 a.a. *
Waters ×247
* Residue conservation analysis
PDB id:
3g5y
Name: Immune system
Title: Antibodies specifically targeting a locally misfolded region associated egfr
Structure: 175 light chain. Chain: a. Engineered: yes. 175 heavy chain. Chain: b. Engineered: yes. Egfr peptide. Chain: e. Engineered: yes
Source: Mus musculus. Mouse. Organism_taxid: 10090. Expressed in: mus musculus. Expression_system_taxid: 10090. Expression_system_cell: hybridoma cell. Synthetic: yes. Other_details: peptide synthesis
Resolution:
1.59Å     R-factor:   0.199     R-free:   0.250
Authors: T.P.J.Garrett,A.W.Burgess
Key ref:
T.P.Garrett et al. (2009). Antibodies specifically targeting a locally misfolded region of tumor associated EGFR. Proc Natl Acad Sci U S A, 106, 5082-5087. PubMed id: 19289842 DOI: 10.1073/pnas.0811559106
Date:
05-Feb-09     Release date:   09-Feb-10    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
No UniProt id for this chain
Struc: 213 a.a.
Protein chain
No UniProt id for this chain
Struc: 210 a.a.
Protein chain
Pfam   ArchSchema ?
P00533  (EGFR_HUMAN) -  Epidermal growth factor receptor
Seq:
Struc:
 
Seq:
Struc:
 
Seq:
Struc:
1210 a.a.
16 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: Chain E: E.C.2.7.10.1  - Receptor protein-tyrosine kinase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: ATP + a [protein]-L-tyrosine = ADP + a [protein]-L-tyrosine phosphate
ATP
+ [protein]-L-tyrosine
= ADP
+ [protein]-L-tyrosine phosphate
Molecule diagrams generated from .mol files obtained from the KEGG ftp site

 

 
    reference    
 
 
DOI no: 10.1073/pnas.0811559106 Proc Natl Acad Sci U S A 106:5082-5087 (2009)
PubMed id: 19289842  
 
 
Antibodies specifically targeting a locally misfolded region of tumor associated EGFR.
T.P.Garrett, A.W.Burgess, H.K.Gan, R.B.Luwor, G.Cartwright, F.Walker, S.G.Orchard, A.H.Clayton, E.C.Nice, J.Rothacker, B.Catimel, W.K.Cavenee, L.J.Old, E.Stockert, G.Ritter, T.E.Adams, P.A.Hoyne, D.Wittrup, G.Chao, J.R.Cochran, C.Luo, M.Lou, T.Huyton, Y.Xu, W.D.Fairlie, S.Yao, A.M.Scott, T.G.Johns.
 
  ABSTRACT  
 
Epidermal Growth Factor Receptor (EGFR) is involved in stimulating the growth of many human tumors, but the success of therapeutic agents has been limited in part by interference from the EGFR on normal tissues. Previously, we reported an antibody (mab806) against a truncated form of EGFR found commonly in gliomas. Remarkably, it also recognizes full-length EGFR on tumor cells but not on normal cells. However, the mechanism for this activity was unclear. Crystallographic structures for Fab:EGFR(287-302) complexes of mAb806 (and a second, related antibody, mAb175) show that this peptide epitope adopts conformations similar to those found in the wtEGFR. However, in both conformations observed for wtEGFR, tethered and untethered, antibody binding would be prohibited by significant steric clashes with the CR1 domain. Thus, these antibodies must recognize a cryptic epitope in EGFR. Structurally, it appeared that breaking the disulfide bond preceding the epitope might allow the CR1 domain to open up sufficiently for antibody binding. The EGFR(C271A/C283A) mutant not only binds mAb806, but binds with 1:1 stoichiometry, which is significantly greater than wtEGFR binding. Although mAb806 and mAb175 decrease tumor growth in xenografts displaying mutant, overexpressed, or autocrine stimulated EGFR, neither antibody inhibits the in vitro growth of cells expressing wtEGFR. In contrast, mAb806 completely inhibits the ligand-associated stimulation of cells expressing EGFR(C271A/C283A). Clearly, the binding of mAb806 and mAb175 to the wtEGFR requires the epitope to be exposed either during receptor activation, mutation, or overexpression. This mechanism suggests the possibility of generating antibodies to target other wild-type receptors on tumor cells.
 
  Selected figure(s)  
 
Figure 3.
Crystal structures of EGFR 287–302 bound to the Fab fragments. (A) Cartoons of Fab806 (Upper), with the light chain (red), heavy chain (blue), bound peptide (yellow), and the superposed EGFR[287–302] from EGFR (purple) and Fab175 (Lower), with the light chain (yellow), heavy chain (green), bound peptide (lilac), and EGFR[287–302] (purple). (B) Detailed stereoview of 175 Fab complex, viewed orthogonal to A, looking into the antigen-binding site. Peptide backbones are shown as cartoon and Cα traces and the interacting side chains as sticks (same coloring as in A). Atoms are colored as follows: O, red; N, blue; and S, orange; C is as for the main chain and the side chain hydrogen bonds are dashed. Water molecules buried upon complex formation are red spheres. (C) Superposition of EGFR (turquoise surface with the 806 epitope displayed as a purple Cα trace) with the Fab175:peptide complex showing spatial overlap (same coloring as in B).
Figure 4.
Location of the mAb806/175 epitope in EGFR. (A) Cα trace of the ectodomain for untethered EGFR1–621. L1 is at the top with L1 and L2 in teal, CR1 and CR2 in blue, and TGF-α in olive. The mAb806/175 epitope is drawn in red, with the preceding loop (residues 271–286) in orange and the disulfide bonds in yellow. The model was constructed by docking the EGFR-ECD CR2 domain from the tethered conformation (36) onto the structure of an untethered EGFR monomer in the presence of its ligand (24). (B) Untethered EGFR ectodomain (as in A) showing the spacial overlap with the mAb175 Fab (solvent contact surface with the light chain in yellow and the heavy chain in green). The Fab was positioned by superimposing the EGFR[287–302] peptide on the corresponding region of EGFR. The CR1 domain is almost completely buried within the Fab. (C) Detail of A, viewed from the right, showing how the preceding disulfide-bonded loop blocks access to the epitope.
 
  Figures were selected by an automated process.