FIGURE 4.

Identification of β-catenin as a target protein of SP141. (A) Biotinylated SP141 (biotin-SP141) was conjugated with avidin beads and then incubated with Panc-1 and AsPC-1 cell lysates in the presence or absence of 2-, 10-, and 20-fold excess of non-biotinylated SP141. The mixtures were blotted for bound β-catenin and MDM2 proteins. (B) Biotin-SP141-conjugated avidin beads were incubated with various concentrations of recombinant His-β-catenin in the presence or absence of non-biotinylated SP141. The bound proteins were detected using anti-His and anti-Biotin antibodies. (C) Computational modeling of SP141 binding to β-catenin. SP141 was rendered in yellow. (D) The predicted binding mode of SP141 with β-catenin. The key residues interacting with SP141 were rendered as sticks. (E) Biotin-SP141-conjugated avidin beads were incubated with the His-tagged recombinant proteins of full length and different regions of β-catenin. The inputs and bound proteins were examined using anti-His antibody. All experiments were repeated at least three times, and similar results were obtained each time.