Figure 3.

Myc-PDK1-mediated phosphorylation of (H89)-C is prevented by depletion of Myc-PDK1 with PDK1 antibodies. The S100 fraction of 293 cells transfected with Myc-PDK1 was diluted 5- or 20-fold and then preincubated with or without PDK1 antibodies as described in Materials and Methods. Cross-reacting proteins were removed by centrifugation after the addition of protein G beads. (A) Autoradiography showing phosphorylation of H89-C by Myc-PDK1 with or without preincubation with the PDK1 antibody. (B) Western blot showing the amount of Myc-PDK1 remaining in the cell extracts with and without preincubation with PDK1 antibody. (C) Immunoblot displaying the amount of Myc-PDK1 pulled down by pretreatment with anti PDK1.