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

O’Flaherty, Derek K., Guengerich, F. Peter, Egli, Martin, Wilds, Christopher J.
Biochemistry 2015 v.54 no.51 pp. 7449-7456
DNA, DNA-directed DNA polymerase, chemical elements, crosslinking, humans, mass spectrometry
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.