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Y627F

Gab1-. WT. Y627F. D PH. WT. D Grb2. _. _. _. _. _. EGF (ng/ml). 5. 50. 5. 50. 5. 50. 5. 50. 5. 50. pERK1-Myc-His. Gab1-Myc. Gab1-Flag. ERK1-Myc-His. LEGEND OF SUPPLEMENTAL FIGURE S1

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Y627F

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  1. Gab1- WT Y627F DPH WT DGrb2 _ _ _ _ _ EGF (ng/ml) 5 50 5 50 5 50 5 50 5 50 pERK1-Myc-His Gab1-Myc Gab1-Flag ERK1-Myc-His LEGEND OF SUPPLEMENTAL FIGURE S1 HEK293 cells were cotransfected with plasmids encoding His-Myc-tagged ERK1 and the following Myc-tagged Gab1 constructs: wild-type (WT) or mutated on its Shp2 binding site (Y627F) or deleted of its PH domain (∆PH), Flag-tagged Gab1 constructs: wild-type (WT) or mutated on its Grb2-binding sites (∆Grb2), as indicated. After stimulation with EGF (5 or 50 ng/ml) when indicated, cells were lysed and processed for His-tag affinity precipitation followed by anti-pERK1-2 immunoblotting. The corresponding lysates were then subjected to anti-Myc or anti-Flag immunoblotting to control the expression level of each construct. Sampaio et al. Supplemental Figure S1

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