Human transforming growth factor α (TGFα) is a 50-residue mitogenic peptide with a compact structure restrained by three disulfide bonds. Sequential and overlapping synthetic peptides were made to identify epitopes of TGFα using a panel of murine monoclonal antibodies and rabbit polyclonal antibodies. Antibodies were raised against human TGFα from different preparations obtained from either chemical synthesis or recombinant DNA techniques. Two related methodologies were used in these experiments. In the first method, probes were synthesized as peptides immobilized on polyethylene pins by the method of Geysen et al. (Geysen, H.M., Meloen, R.H., and Barteling, S.J. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 3998-4002). Three sets of sequentially overlapping tetrapeptides, hexapeptides, and octapeptides covering the entire length of the human TGFα sequence were synthesized. In the second method, a set of overlapping 8-residue synthetic peptides, freely soluble in solution, were used as probes. By both methods, the nonneutralizing monoclonal antibodies, i.e. those that did not inhibit TGFα in mitogenic assays, recognized two immunodominant regions represented by the NH2-terminal segment (residues 1-9) and the most prominent β-sheet of the molecule (residues 22-31). The NH2 terminus and the β-sheet-(22-31) are in the same face of the molecule as determined by the solution structure. These two immunodominant regions were also recognized by the polyclonal antibodies as well as regions in the COOH terminus as minor epitopes. However, none of the neutralizing monoclonal antibodies recognized any synthetic peptides. Thus, our results suggest that the receptor-binding surface of TGFα does not involve the face represented by the NH2-terminal fragment and the major β-sheet of residues 22-31, but rather, that the opposite face represented by two loops formed by residues 12-20 and 34-43 may be involved in TGFα binding to its receptor.
|Original language||English (US)|
|Number of pages||6|
|Journal||Journal of Biological Chemistry|
|State||Published - 1989|
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