Figure 2

Multimerization assays with the Cupidin deletion mutants. Cross-linking assay using the chemical cross-linker DMP. Bacterially expressed GST-proteins were digested by thrombin, and the resulting untagged proteins were incubated in the presence (+) or absence (-) of DMP followed by Western analysis using anti-Cupidin (CPD C) antibody. One asterisk indicates the migration of multimers, and two asterisks indicate that of monomers. (B) Co-immunoprecipitation assay using extracts from cells in which the three distinct constructs shown in the upper panels were heterologously overexpressed in COS7 cells. I, Input; C, IP samples using non-immune serum; F, IP samples using anti-Flag antibody. Western blot analysis was performed using anti-Cupidin antibody, and the multimerization efficacies were calculated from the band signal intensities, and shown in the graph at the bottom.