Fig. (2). SUMOylation Site in GSK 3β (A) The purified GST-GSK 3β wt or GST-GSK 3β SUMO mutant (K292R) fusion protein was used as the substrate protein in the SUMOy-lation assay as described in the Materials and Methods section. The SUMOylation of GSK 3β wt was detected as a high molecular weight protein band (left lane), whereas its SUMO mutant was totally inhibited, as shown (right lane). (B) Ha –GSK 3β wt or Ha –GSK 3β SUMO mutant was transfected to COS-1 cells and immunoprecipitated with Ha mouse monoclonal antibody. The immunoprecipitants were sub-jected to the western bolt with SUMO-1, as described in the Materials and Methods section. The SUMOylation of GSK 3β wt was indicated as several high molecular weight protein bands (left lane), whereas its SUMO mutant was totally inhibited (right lane). To monitor the GSK 3β protein expression, the immunoprecipitants were subjected to the western bolt with GSK 3β polyclonal antibody (bottom).