Noncontiguous regions in the extracellular domain of EGF receptor define ligand-binding specificity.
Murine epidermal growth factor (EGF) binds with approximately 250-fold higher binding affinity to the human EGF receptor (EGFR) than to the chicken EGFR. This difference in binding affinity enabled the identification of a major ligand-binding domain for EGF by studying the binding properties of various chicken/human EGFR chimera expressed in transfected cells lacking endogenous EGFR. It was shown that domain III of EGFR is a major ligand-binding region. Here, we analyze the binding properties of novel chicken/human chimera to further delineate the contact sequences in domain III and to assess the role of other regions of EGFR for their contribution to the display of high-affinity EGF binding. The chimeric receptors include chicken EGFR containing domain I of the human EGFR, chicken receptor containing domain I and III of the human EGFR, and two chimeric chicken EGFR containing either the amino terminal or the carboxy terminal halves of domain III of human EGFR, respectively. In addition, the binding of various human-specific anti-EGFR monoclonal antibodies that interfere with EGF binding is also compared. It is concluded that noncontiguous regions of the EGFR contribute additively to the binding of EGF. Each of the two halves of domain III has a similar contribution to the binding energy, and the sum of both is close to that of the entire domain III. This suggests that the folding of domain III juxtaposes sequences that together constitute the ligand-binding site. Domain I also provides a contribution to the binding energy, and the added contributions of both domain I and III to the binding energy generate the high-affinity binding site typical of human EGFR.
Lax
I
I
Rorer Biotechnology, King of Prussia, Pennsylvania 19406.
Fischer
R
R
Ng
C
C
Segre
J
J
Ullrich
A
A
Givol
D
D
Schlessinger
J
J
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002645:Chickens; D003001:Cloning, Molecular; D004247:DNA; D004815:Epidermal Growth Factor / Q000378:metabolism*; D000939:Epitopes; D066246:ErbB Receptors / Q000276:immunology / Q000378:metabolism*; D005110:Extracellular Space / Q000378:metabolism; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D016211:Transforming Growth Factor alpha / Q000378:metabolism
D000939:Epitopes; D008024:Ligands; D016211:Transforming Growth Factor alpha; D004815:Epidermal Growth Factor; D004247:DNA; D066246:ErbB Receptors