Mg2+- dependent AP-endonuclease activity of the WT and mutant (Cys→Ser, Ala and Thr) APE1 proteins. The 43-mer, THF-containing oligonucleotide was incubated with 33 pM APEs for 3 min at 37° C. AP-endonuclease activity measured using a buffer containing 10 mM Mg2+. (a) Lane 1, no protein; lane 2, WT APE1; lanes 3–5, three different batches of C99S mutant; lane 6, C99A mutant; lane 7, C99T mutant; lane 8, C99S/C138S double mutant; lane 9, C99S/C65S/C138S triple mutant. (b) Lane 1, WT APE1; lanes 2, C65S mutant; lane 3, C93S mutant; lanes 4–6, three different batches of C99S mutant; lane 7, C138S mutant; lane 8, C208S mutant; lane 9, C296S mutant; lane 10, C310S mutant.