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. 2014 Apr 21;289(22):15810–15819. doi: 10.1074/jbc.M114.572081

FIGURE 2.

FIGURE 2.

Sumoylation of RanGAP1 and TDG at 22 °C. A and B, in vitro sumoylation experiments performed for modification of RanGAP1-NΔ419 and TDG by SUMO-1 (A) or SUMO-2 (B) and monitored by electrophoresis. The first lane of each gel contains pure TDG, and the second lane contains pure SUMO-1-modified TDG (TDG∼SUMO-1 or TDG-S). Reactions were initiated by adding ATP (2.5 mm) to buffer containing E1 (0.1 μm), E2 (1 μm), SUMO-1 or SUMO-2 (10 μm), and either RanGAP1-NΔ419 or TDG (2 μm). C, in vitro sumoylation of TDG with SUMO-2 over a 24-h time course. Samples were extracted at the time points given. D, in vitro sumoylation of TDG with SUMO-1 and SUMO-2 using SUMO concentrations of <10 μm. SUMO concentrations for a given assay are listed above each set of time points. Samples were extracted from the reaction and quenched at the indicated times (given in minutes), and the reaction progress was analyzed by SDS-PAGE under reducing conditions.