Study of EGFR Signaling/Endocytosis by Site-Directed Mutagenesis

As an essential tool in molecular biology and biochemistry, in vitro mutagenesis is important for mechanistic and functional studies of protein at the cellular level. Through site-directed substitution, insertion, or deletion of potential function residues, we may evaluate the specific biological role of proteins and understand the underlying mechanism. EGF ligand binding will activate EGFR and initiate signaling transduction for cell proliferation, differentiation, and survival; meanwhile, it will also induce the EGFR endocytosis for degradation to maintain cellular homeostasis. During this complicated process, some key residue(s) or sequences of EGFR molecule play essential regulatory roles. We describe in this chapter the inverse PCR-based site-directed mutagenesis protocol used in our studies on EGFR signaling and endocytosis.