Figure 6. Kinetic parameters without and with ML199 or Compound 1.

Figure 6Kinetic parameters without and with ML199 or Compound 1

Ten pg of APE1 (~28 pM) was incubated without or with 5 μM of the indicated inhibitor at room temperature for 15 min. Increasing concentrations of radiolabeled abasic DNA substrate (i.e. 5, 10, 25, 50, or 100 nM) were then added, and the reactions were incubated at 37 ºC for 5 min before the addition of stop buffer. Intact substrate was separated from incised product on a 15% polyacrylamide denaturing gel, and the percent conversion was determined by standard phosphorimager analysis. Lineweaver – Burk plots of 1/V versus 1/[S] were used to determine KM and kcat (shown). The plotted data points (averages and standard deviations) were derived from 11 independent values for the no inhibitor reactions, and 5 values for each of the inhibitor points.

From: Small Molecule Inhibitors of the Human Apurinic/apyrimidinic Endonuclease 1 (APE1)

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