Disruption of the intramolecular interaction between human lung cancer-related EGFR kinase domain and carboxy-terminal tail by intra-Tyr1016 phosphorylation or extra-peptidic competitors.

contextHuman epidermal growth factor receptor (EGFR) controls many key signaling pathways associated with cell growth/survival and is a well-established druggable target of lung cancer. Previous studies have primarily focused on the extracellular ligand-binding domain (LBD) and intracellular kinase domain (KD) of EGFR, while its carboxy-terminal (CT) tail extended from the KD domain still remains largely unexplored to date. Considering that only the first ~ 80 CT residues (termed pkCT-tail) proximal to the KD are important for the kinase autoinhibition, we herein attempted to systematically investigate the structural basis, energetic property, and dynamic behavior of pkCT-tail as well as its role in EGFR activity and function

methodsA 10-mer hotspot segment (residues 1009-1018) was identified to play an important role in mediating the intramolecular binding event of pkCT-tail to the KD domain, which is partially folded into an ordered, one-rounded helical conformation to tightly pack against the docking site of the KD domain. It is revealed that the Tyr1016 residue is a key anchor in the hotspot; its phosphorylation triggers the unbinding event of the hotspot segment, pkCT-tail, and even the whole CT-tail from the KD domain in a reversible manner. Several chemically stapled peptidic competitors were rationally designed as the potent binders of the KD domain and can compete with the native hotspot segment for the docking site. We demonstrated that both the intra-Tyr1016 phosphorylation and extra-peptidic competitors share a similar effect on EGFR, which can disrupt the native intramolecular interaction between the EGFR KD domain and pkCT-tail, thus unlocking the autoinhibitory state of the EGFR kinase. However, the peptidic competitors were observed to only moderately activate the kinase, imparting that the kinase activity is controlled by multiple factors rather than only the pkCT-tail.