The Werner syndrome protein limits the error-prone 8-oxo-dG lesion bypass activity of human DNA polymerase kappa.
Bottom Line: Steady-state kinetic analysis reveals that WRN improves hpol κ-catalyzed dCMP insertion opposite 8-oxo-dG ∼10-fold and extension from dC:8-oxo-dG by 2.4-fold.Stimulation is primarily due to an increase in the rate constant for polymerization (kpol), as assessed by pre-steady-state kinetics, and it requires the RecQ C-terminal (RQC) domain.In support of the functional data, recombinant WRN and hpol κ were found to physically interact through the exo and RQC domains of WRN, and co-localization of WRN and hpol κ was observed in human cells treated with hydrogen peroxide.
Affiliation: Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, AR 72205-7199, USA.Show MeSH
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Mentions: Next, we tested whether the isolated WRN exo domain could preferentially degrade 8-oxo-dG containing bp (Figure 4A). In the absence of hpol κ, the WRN1-333 exo domain exhibits a clear preference for the degradation of dA-containing mis-pairs regardless of whether 8-oxo-dG is present or unmodified dG. The rate of degradation was ∼0.9 nM min−1 for the two dA-containing mis-pairs, whereas the rate of degradation for dC:dG and dC:8-oxo-dG bp was 0.33 and 0.40 nM min−1, respectively (Figure 4B). The rate of WRN-catalyzed dA:8-oxo-dG degradation is ∼2-fold faster than dC:8-oxo-dG.
Affiliation: Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, AR 72205-7199, USA.