Phosphorylation of EBP50 by PKC or Cdc2 inhibits its ability to bind two PDZ ligands simultaneously. (A) Binding between untagged full-length EBP50 and 6xHis-EPI64 was tested by precipitating the 6xHis tag with Talon beads. (B) EBP50 was preincubated with soluble 6xHis-EPI64 and complexes recovered on FERM beads. (C) EBP50 was preincubated with soluble 6xHis-EPI64 and its ability to simultaneously bind GST–β-catenin tail coupled to glutathione agarose was tested. For A–C, 20% of the bound fraction was run on the gels.