Fig. 1.

Global Identification of SUMO E3 ligase-dependent substrates using the HuProt™ arrays. A, Schematic describing the global SUMOylation assay design. SUMOylation reactions with high and low concentrations of recombinant E1/E2, paired with SUMO1 or SUMO2, were performed as controls. E3 ligases were spiked into the low E1/E2 conditions and performed with SUMO1 and SUMO2. All experimental and control conditions were performed in triplicate. B, E3 ligase activity was tested in pilot study performed on a specialized SUMOylation protein chip containing 82 control proteins. Low concentrations of E1/E2 allow for direct detection of E3 ligase activity compared with high E1/E2 controls reactions. C, PIAS3 SUMO2 substrates from the HuProt™ microarray are highlighted.