Backbone Flexibility Influences Nucleotide Incorporation by Human Translesion DNA Polymerase eta opposite Intrastrand Cross-Linked DNA

摘要

Intrastrand cross-links (IaCL) connecting two purine nucleobases in DNA pose a challenge to high-fidelity replication in the cell. Various repair pathways or polymerase bypass can cope with these lesions. The influence of the phosphodiester linkage between two neighboring 2'-deoxyguanosine (dG) residues attached through the O-6 atoms by an alkylene linker on bypass with human DNA polymerase eta (hPol eta) was explored in vitro. Steady-state kinetics and mass spectrometric analysis of products from nucleotide incorporation revealed that although hPol eta is capable of bypassing the 3'-dG in a mostly error-free fashion, significant misinsertion was observed for the 5'-dG of the IaCL containing a butylene or heptylene linker. The lack of the phosphodiester linkage triggered an important increase in frameshift adduct formation across the 5'-dG by hPol eta, in comparison to the 5'-dG of IaCL DNA containing the phosphodiester group.

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