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Fig. 1. Isolation and identification of phosphotyrosine-dependent Dab1-binding proteins from embryonic mouse brain. (A) Two protein bands are observed to bind specifically to phosphorylated GST-Dab1 by silver staining samples resolved by SDS-PAGE (arrows). Embryonic brain lysates were incubated with immobilized tyrosine-phosphorylated GST-Dab1 (pY, lane 1) or unphosphorylated Dab1 (Y, lane 2) and bound proteins were eluted with 100 mM phenyl phosphate solution. The band at approximately 180 kDa was more distinct in the preparative gels used for protein identification. (B) Dock1, associated with tyrosine-phosphorylated GST-Dab1 (lane 1), but not unphosphorylated GST-Dab1 (lane2), was detected by western blotting samples eluted with sample buffer. (C) Crk was observed to interact with tyrosine-phosphorylated GST-Dab1 (lane 1) but not unphosphorylated GST-Dab1 (lane 2) by western blotting samples eluted with sample buffer.





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