Figure 5

Methodology used to identify in vivo kinase substrates using FSBA and quantitative MS. In vivo substrates are identified based on the higher phosphopeptide intensity in MS for both the vehicle-treated (blue) and kinase-added (green) samples. A higher phosphopeptide abundance in MS for the vehicle-treated (blue) sample relative to the inhibitor-treated (red) sample indicates a phosphorylation site regulated by the kinase of interest in vivo. A higher phosphopeptide abundance for the kinase-added (green) sample relative to the inhibitor-treated (red) sample indicates a phosphorylation site directly targeted by the kinase in vitro.