Quantitation of Phosphorylation Preferences of EGFR Tyrosines in Response to HB‐EGF and EGF

In response to extracellular ligands, some receptors in the plasma membrane undergo phosphorylation, the enzymatic transfer of a phosphate group to a specific amino acid such as tyrosine. This chemical reaction is critically important in biological processes and thus quantitative approaches to monitor phosphorylation and understand how it is affected by the identity of the activated ligand are being sought. Here, we examine the utility of the recently introduced experimental platform of osmotically‐derived plasma membrane vesicles in quantifying the phosphorylation preferences of the EGFR ligand HB‐EGF. We introduce improvements in the platform’s detection method and in the quantitation of ligand bias, expanding the toolkit for studies of the physical chemistry behind receptor signaling.