Protein Interactions in the T7 DNA Replisome Facilitate DNA Damage Bypass

Abstract

The DNA replisome inevitably encounters DNA damage during DNA replication. The T7 DNA replisome contains a DNA polymerase (gp5), the processivity factor thioredoxin (trx), a helicase-primase (gp4), and a ssDNA-binding protein (gp2.5). T7 protein interactions mediate this DNA replication. However, whether the protein interactions could promote DNA damage bypass is still little addressed. In this study, we investigated strand-displacement DNA synthesis past 8-oxoG or O⁶ -MeG lesions at the synthetic DNA fork by the T7 DNA replisome. DNA damage does not obviously affect the binding affinities between helicase, polymerase, and DNA fork. Relative to unmodified G, both 8-oxoG and O⁶ -MeG-as well as GC-rich template sequence clusters-inhibit strand-displacement DNA synthesis and produce partial extension products. Relative to the gp4 ΔC-tail, gp4 promotes DNA damage bypass. The presence of gp2.5 also promotes it. Thus, the interactions of polymerase with helicase and ssDNA-binding protein facilitate DNA damage bypass. Accessory proteins in other complicated DNA replisomes also facilitate bypassing DNA damage in similar manner. This work provides new mechanistic information relating to DNA damage bypass by the DNA replisome.

Keywords: DNA damage; DNA replication; protein-protein interactions; replisome.