Figure 4.

Ubiquitin acceptor Lys in APE1. (A) A549 was transiently transfected with cDNA encoding ‘KallR’ (all Lys residues in APE1 were altered to Arg) as an internal control for transfection (Supplementary Figure S4), along with wtAPE1 (1), T233E (2), T233E K24/25/27R (3) and T233E ND41 APE1 (N-terminal deletion of 41 amino acid residues). (Top) Ubiquitinated APE1; Bot) intact APE1, KallR and ND41 are shown. KallR mutant APE1 migrates slightly faster than the intact APE1 in SDS/PAGE, due to the difference in amino-acid composition from the wtAPE1 (Supplementary Figure S4). (B) Relative intensities of ubiquitinated APE1 were calculated from three independent experiments as in (A), using KallR signals to normalize the transfection efficiency.