Multidomain binding of transforming growth factor alpha to the epidermal growth factor receptor.
Solubilized epidermal growth factor receptor (EGF-R) has been used in an extension of the Geysen epitope mapping protocol in order to provide additional insight into the amino acid residues in human transforming growth factor alpha (hTGF alpha) which are critical to recognition and binding. Overlapping heptapeptides which encompassed the 50 amino acid primary sequence of hTGF alpha were synthesized on a polyethylene solid phase, and the amount of detergent-solubilized EGF-R bound to each peptide was measured using ELISA. EGF-R appeared to bind reproducibly to four heptapeptides cognate to sequences in both the N- and C-domains of hTGF alpha (residues 22-28, 28-34, 36-42, and 44-50). Visualization of these four regions on three-dimensional solution phase structures of hTGF alpha, derived from 1H NMR measurements [Kline, T.-P., Brown, F.K., Brown, S.C., Jeffs, P.W., Kopple, K.D., & Mueller, L. (1990) Biochemistry 29, 7805-7813], indicated that the peptide segments are located on a single face of the protein and suggested the presence of a potential receptor binding cavity. If peptide segments within both the N- and C-domains of hTGF alpha are involved in binding to EGF-R, then this has direct consequences for possible molecular mechanisms by which receptor activation might take place. For example, the observed conformational flexibility in the six NMR-derived hTGF alpha structures due to variations in the main-chain torsion angles of Val-33, in combination with the involvement of residues from both domains in the proposed binding cavity, may imply that receptor activation results from interdomain reorientation in the protein ligand.(ABSTRACT TRUNCATED AT 250 WORDS)
Richter
A
A
CRC Unit of Medical Oncology, Southampton University Medical School, Southampton General Hospital, U.K.
Conlan
J W
JW
Ward
M E
ME
Chamberlin
S G
SG
Alexander
P
P
Richards
N G
NG
Davies
D E
DE
D000595:Amino Acid Sequence; D001665:Binding Sites; D004797:Enzyme-Linked Immunosorbent Assay; D066246:ErbB Receptors / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism; D016211:Transforming Growth Factor alpha / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D010446:Peptide Fragments; D016211:Transforming Growth Factor alpha; D066246:ErbB Receptors