Auto-ubiquitination assay performed in which the E3 ligase complex RAD18-RAD6 was incubated with E1 and ubiquitin in the presence/absence of MAGEA4for 1 hour at 37C. All proteins were purified from E.coli. Samples were loaded and run onto a precast SDS PAGE gel, which was stained with Coomasie. Approximately 20ug of RAD18-RAD6 was used for the assay.