Quantification of HER2 phosphorylation and the differences after treatment.

*Phosphorylation sites were identified by HER2 phosphomapping.†Quantitative mass spectrometry (MS) was done to determine the basal level of phosphorylation at each site. The absolute amount of phosphorylated peptide was compared to the total of phosphorylated and non-phosphorylated peptides to calculate the percent phosphorylation (% Phosph.)‡Ten minutes after treatment with the indicated molecule, quantitative MS was done to determine the % phosphorylation. This was subtracted from the % phosphorylation in the basal state to provide the mean difference after treatment reported here. This time point was chosen because maximal phosphorylation of AKT was shown to occur within ten minutes after agonist treatment (Figure 4).