Acetylation of p73 in 293T cell

293T cells ectopically expressing FLAG-tagged p73 were treated with AGK2 and p73 was immunoprecipitated. The precipitants were resuspended in deacetylation buffer (50 mM Tris-HCl pH 8.0, 140 mM NaCl, 2 mM MgCl 2 ) and incubated with recombinant SIRT2 protein (10 U). The reaction was terminated by boiling with SDS sample buffer and acetylation levels were determined by immunoblotting.