Fig. 4. Effects of kinase domain mutations on the autophosphorylation activity of p94fer. A, wild-type fer (Lane 1), fer KR652/3NQ (Lane 2), and fer{Delta}816–823 (Lane 3) were linked to a HA epitope, overexpressed transiently in COS1 cells, and immunoprecipitated with {alpha}-HA antibody. The precipitated proteins were exposed to Western blot analysis using monoclonal {alpha}-HA antibody (top) or monoclonal {alpha}-phosphotyrosine (bottom). The marked band (bottom) represents an in vivo autophosphorylated p94fer. B, the same analysis as in A was carried out with: wild-type fer (Lane 1), fer Y715F (Lane 2), fer G571R (Lane 3), fer G571A (Lane 4), fer K592R (Lane 5), and fer K592N (Lane 6).