Skip to main content
. Author manuscript; available in PMC: 2010 Oct 2.
Published in final edited form as: DNA Repair (Amst). 2009 Aug 4;8(10):1215–1224. doi: 10.1016/j.dnarep.2009.07.003

Fig. 6.

Fig. 6

Effect of RPS3 on in vitro ubiquitination of p53 by MDM2. For the in vitro ubiquitination assay, 500 ng of purified p53 (8 pmol) was mixed with 2 μg of recombinant MDM2 (wild type, 20 pmol) or MDM2 C464S (E3 ubiquitin ligase defective) proteins and added to the ubiquitination reaction containing E1 (4.5 μM), E2 (130 μM) and 1 mg of ubiquitin. The reactions were then incubated with increasing concentrations (2.5, 25, and 250 ng; 5 pmol) of RPS3. p53 ubiquitination was examined by resolving the reactions on SDS-PAGE, transferring on to PVDF membrane and immunoblotting with p53 DO-1 antibody. Light and dark (above and below) exposures of the western blot show p53 laddering, representing ubiquitination.