Lesion (in)tolerance reveals insights into DNA replication fidelity

Abstract

The initial encounter of an unrepaired DNA lesion is likely to be with a replicative DNA polymerase, and the outcome of this event determines whether an error-prone or error-free damage avoidance pathway is taken. To understand the atomic details of this critical encounter, we have determined the crystal structures of the pol α family RB69 DNA polymerase with DNA containing the two most prevalent, spontaneously generated premutagenic lesions, an abasic site and 2′-deoxy-7,8-dihydro-8-oxoguanosine (8-oxodG). Identification of the interactions between these damaged nucleotides and the active site provides insight into the capacity of the polymerase to incorporate a base opposite the lesion. A novel open, catalytically inactive conformation of the DNA polymerase has been identified in the complex with a primed abasic site template. This structure provides the first molecular characterization of the DNA synthesis barrier caused by an abasic site and suggests a general mechanism for polymerase fidelity. In contrast, the structure of the ternary 8-oxodG:dCTP complex is almost identical to the replicating complex containing unmodified DNA, explaining the relative ease and fidelity by which this lesion is bypassed.