Phorbol ester-induced threonine phosphorylation of the human epidermal growth factor receptor occurs within the EGF binding domain.

Partial proteolysis with trypsin has been used to map the sites of phorbol ester-induced phosphorylation of the epidermal growth factor (EGF) receptor. Both 12-O-tetradecanoylphorbol 13-acetate (TPA) and EGF stimulate phosphorylation of the EGF receptor in intact human carcinoma cells. Under the conditions examined, EGF is more effective than TPA in stimulating phosphorylation of a 45 kDa intracellular receptor domain, while TPA is more effective than EGF in inducing phosphorylation of a 120 kDa transmembrane EGF-binding domain. The phosphorylation of the 120 kDa peptide occurs primarily on threonine residues. Two-dimensional peptide mapping indicates that the two major phosphopeptides found in the 120 kDa receptor fragment correspond to the major new phosphopeptides found in intact EGF receptor following treatment with TPA. Thus, the major sites of TPA-induced threonine phosphorylation reside in the 120 kDa binding domain of the EGF receptor.