Purification of the Extracellular Domain of the Epidermal Growth Factor Receptor Produced by Recombinant Baculovirus-Infected Insect Cells in a 10-L Reactor

Abstract

The binding of ligand to the extracellular domain of the epidermal growth factor (EGF) receptor is the event that initiates the activation of the tyrosine kinase activity present on the cytoplasmic domain. This activation leads to the phosphorylation of the receptor and other cellular substrates, and results ultimately in stimulation of mitogenesis (1,2). The EGF receptor system is implicated in both the uncontrolled growth of certain carcinomas and the controlled growth that occurs during wound healing (3). In order to develop EGF receptor antagonists that may be used to treat cancer and agonists that may promote wound healing, an understanding of the structure-function relationships of the ligand receptor complex is essential. By studying the soluble extracellular domain of the receptor, the use of detergents is avoided, and biochemical and biophysical analyses are facilitated. We have expressed the extracellular domain of the EGF receptor (EGFR-ED) by infecting insect cells with a recombinant baculovirus that carries a DNA insert encoding for the entire extracellular domain, including the N-terminal signal sequence.