Transient-state kinetics of apurinic/apyrimidinic (AP) endonuclease 1 acting on an authentic AP site and commonly used substrate analogs: the effect of diverse metal ions and base mismatches.

ABSTRACT: Apurinic/apyrimidinic endonuclease 1 (APE1) is an Mg²⁺-dependent enzyme responsible for incising the DNA backbone 5' to an apurinic/apyrimidinic (AP) site. Here, we use rapid quench flow (RQF) techniques to provide a comprehensive kinetic analysis of the strand-incision activity (k(chemistry)) of APE1 acting on an authentic AP site along with two widely used analogs, a reduced AP site and a tetrahydrofuran (THF) site. In the presence of biologically relevant Mg²⁺, APE1 incises all three substrates at a rate faster than the resolution of the RQF, ≥700 s⁻¹. To obtain quantitative values of k(chemistry) and to facilitate a comparison of the authentic substrate versus the substrate analogs, we replaced Mg²⁺ with Mn²⁺ or Ni²⁺ or introduced a mismatch 5' to the lesion site. Both strategies were

SUBMITTER: Schermerhorn KM 

PROVIDER: S-EPMC3859843 | biostudies-literature | 2013 Oct

REPOSITORIES: biostudies-literature