Fig. 2.

Dose-response of Src inhibition on FAK activation loop Tyr⁵⁷⁶ and Tyr⁵⁷⁷ modification by Src in vitro. After incubation with varying concentrations of AZD0530 (0.01, 0.1, 1, and 10 μM), recombinant Src was incubated with recombinant FAK for 15 min at 37°C. LC-MRM/MS was then used to absolutely quantify the amount of FAK activation loop modification. A, the amount of modified Tyr⁵⁷⁶/Tyr⁵⁷⁷ FAK activation loop (⋄). B, the amount of FAK activation loop in the mono-phosphorylated pTyr⁵⁷⁶ (□) and bis-phosphorylated pTyr⁵⁷⁶/pTyr⁵⁷⁷ (▵) forms. C, an IC₅₀ value of Src inhibition calculated from the amount of pTyr⁵⁷⁶/pTyr⁵⁷⁷ (▵) determined at each dose. Data points represent the means of three separate experiments, where error bars are ± S.E.M.