Abstract
Publisher Summary The epidermal growth factor (EGF) receptor is a 1186-amino acid transmembrane phosphorylated glycoprotein that mediates the biological effects of EGF. This chapter discusses a single-step immunoaffinity chromatography procedure using a monoclonal anti-EGF receptor antibody, which is a competitive inhibitor of EGF binding. Desorption with urea gives the quantitative recovery of denatured protein; competitive elution with EGF yields homogeneous active protein. This procedure provides a convenient single-step, high-yield purification of EGF receptor protein. The major limitation is that the active enzyme is eluted with EGF, and it is not suitable for the kinetic or mechanistic analysis of EGF activation. However, it is useful for other kinetic analyses, such as the analysis to understand the effects of self-phosphorylation.
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