Figure 1

GST-FLT3S is an effective substrate for FLT3 kinase activity assays. Reactions were carried out with FLT3WT, FLT3D835Y, and FLT3D835H at 1.6, 0.4, and 0.4 μg/ml, respectively. A. Assays performed in the presence of 0.2 mg/ml GST-FLT3S or GST. Tyrosine phosphorylation was detected by using anti-phosphotyrosine antibody. Note that autophosphorylation of FLT3 was also seen. The protein levels of GST-FLT3S and GST were revealed by Coomassie blue staining. B. Comparison of specific activity of wild type and two mutant forms of FLT3 recombinant proteins determined with GST-FLT3S at 0.2 mg/ml. Error bars denote standard deviation. C. Activity assays performed with different concentrations of GST-FLT3S. D. Activity assays performed with GST-FLT3S immobilized on glutathione-Sepharose beads. Fluorescent images were acquired under fluorescent microscope with identical exposure times.