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

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journal contribution
posted on 29.12.2015 by Derek K. O’Flaherty, F. Peter Guengerich, Martin Egli, Christopher J. Wilds
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 O6 atoms by an alkylene linker on bypass with human DNA polymerase η (hPol η) was explored in vitro. Steady-state kinetics and mass spectrometric analysis of products from nucleotide incorporation revealed that although hPol η 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 η, in comparison to the 5′-dG of IaCL DNA containing the phosphodiester group.