Identification of the high affinity binding site of transforming growth factor-alpha (TGF-alpha) for the chicken epidermal growth factor (EGF) receptor using EGF/TGF-alpha chimeras.
Human epidermal growth factor (hEGF) and human transforming growth factor-alpha (hTGF-alpha) are structurally related growth factors that share relatively little sequence homology. They both exert their biological action by binding to the cell-surface EGF receptor. hEGF and hTGF-alpha bind with similar affinity to the hEGF receptor, but hEGF binds with an approximately 100-fold lower affinity to the chicken EGF receptor compared with hTGF-alpha. To map the region in hTGF-alpha that confers its ability to bind with high affinity to the chicken EGF receptor, 10 hybrids of hEGF and hTGF-alpha were constructed by exchanging domains bordered by the third, fourth, and sixth conserved cysteine residues. The activity of the expressed chimeric proteins was determined by their ability to compete with 125I-labeled mouse EGF for binding to NIH-3T3 cells transfected with the hEGF receptor. Subsequent binding competition studies of NIH-3T3 cells transfected with the chicken EGF receptor showed that chimeras carrying TGF-alpha sequences COOH-terminal of the sixth cysteine have a high affinity for this receptor, similar to hTGF-alpha. In contrast, chimeras with EGF sequences in this COOH-terminal domain have only low binding affinity, similar to hEGF. We conclude that the COOH-terminal linear region of hTGF-alpha is important for its high affinity interaction with the chicken EGF receptor.
Kramer
R H
RH
Department of Cell Biology, University of Nijmegen, The Netherlands.
Lenferink
A E
AE
van Bueren-Koornneef
I L
IL
van der Meer
A
A
van de Poll
M L
ML
van Zoelen
E J
EJ
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002645:Chickens; D004815:Epidermal Growth Factor / Q000737:chemistry*; D066246:ErbB Receptors / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship; D016211:Transforming Growth Factor alpha / Q000737:chemistry / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D016211:Transforming Growth Factor alpha; D004815:Epidermal Growth Factor; D066246:ErbB Receptors