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bioRxiv · 10.1101/2024.05.06.592774

Structural and biochemical characterization of LIG1 during mutagenic nick sealing of oxidatively damaged ends at the final step of DNA repair

Almohdar, D.·Balu, K. E.·Ratcliffe, J.·Tang, Q.·Parwal, T.·Caglayan, M.
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Abstract

DNA ligase 1 (LIG1) joins broken strand-breaks in the phosphodiester backbone to finalize DNA repair pathways. We previously reported that LIG1 fails on nick repair intermediate with 3-oxidative damage incorporated by DNA polymerase (pol) {beta} at the downstream steps of base excision repair (BER) pathway. Here, we determined X-ray structures of LIG1/nick DNA complexes containing 3-8oxodG and 3-8oxorG opposite either a templating Cytosine or Adenine and demonstrated that the ligase active site engages with mutagenic repair intermediates during steps 2 and 3 of the ligation reaction referring to the formation of DNA-AMP intermediate and a final phosphodiester bond, respectively. Furthermore, we showed the mutagenic nick sealing of DNA substrates with 3-8oxodG:A and 3-8oxorG:A by LIG1 wild-type, immunodeficiency disease-associated variants, and DNA ligase 3 (LIG3) in vitro. Finally, we observed that LIG1 and LIG3 seal resulting nick after an incorporation of 8oxorGTP:A by pol{beta} and AP-Endonuclease 1 (APE1) can clean oxidatively damaged ends at the final steps. Overall, our findings uncover a mechanistic insight into how LIG1 discriminates DNA or DNA/RNA junctions including oxidative damage and a functional coordination between the downstream enzymes, pol{beta}, APE1, and BER ligases, to process mutagenic repair intermediates to maintain repair efficiency.

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Almohdar, D., Balu, K. E., Ratcliffe, J., Tang, Q., Parwal, T., Caglayan, M.. 2024-05-06. Structural and biochemical characterization of LIG1 during mutagenic nick sealing of oxidatively damaged ends at the final step of DNA repair. https://doi.org/10.1101/2024.05.06.592774

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