Effect of carboxyl terminal truncation on the tyrosine kinase activity of the epidermal growth factor receptor.

The carboxyl terminal domain of the epidermal growth factor receptor (EGFR) is an important regulatory region in mediating the tyrosine kinase-dependent biological effects of EGF. The effect of a 164-amino-acid carboxyl deletion of the EGFR or the EGFR cytoplasmic kinase domain on in vitro tyrosine kinase activity was assessed. C'-terminal truncation of the EGFR resulted in dependence on Mn2+ for full activity. The EGFR kinase domain (kd EGFR) and the C'-terminally truncated kinase domain (kd c'1022 EGFR) also exhibited a strong preference for Mn2+ compared to Mg2+, with kd c'1022 EGFR being completely inactive in the presence of Mg2+ alone. Sphingosine or ammonium sulfate specifically activated both kd EGFR and kd c'1022 EGFR. EGFR and c'1022 EGFR displayed similar EGF-stimulated in vitro tyrosine kinase activities; however, kd EGFR was 5- to 10-fold more active in vitro than kd c'1022 EGFR. Thus, the regulatory contribution of the C'-terminus is most evident when the EGFR ligand binding domain is removed. These results indicate that an intact EGFR C'-terminus is necessary for the protein to assume a fully active conformation.