Figure 5.

Incision activity of NEIL2 and AcNEIL2. (A) NEIL2 (5 µg) was incubated with either native p300 HAT domain (0.5 µg) (lanes 4 and 5) or heat-inactivated HAT domain (lanes 2 and 3) and 1 mM AcCoA for 45 min at 30°C; 50 and 100 nM of WT NEIL2 (lanes 2 and 3) or AcNEIL2 (lanes 4 and 5) were incubated with 500 nM of 5′ ³²P-labeled 5-OHU·G oligo at 37°C for 20 min, and the cleaved products were analyzed as in Figure 4. Lane 1, no protein. (B) Incision activity of the AcK153R mutant. The K153R mutant (5 µg) was acetylated in vitro and the incision activity was measured as in (A). (C) Western analysis (using anti-AcLys antibody) of WT (lanes 1 and 2) and mutant NEIL2 (lanes 3 and 4) after acetylation as in (A) and (B).