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Only a subset of E2 enzymes ubiquitinate RFP-p53 in MDM2-dependent manner. Beads saturated with RFP-p53 were incubated with all the human E2 enzymes in the presence of MDM2 (A). Similar reactions were also performed with the indicated E2s in the presence of MDM2ΔRING (B). Following the reactions, beads were washed thoroughly, boiled with sample buffer, and analyzed by Western blot with anti-ubiquitin as well as Ponceau-stained. IB, immunoblot.
