Lesion (in)tolerance of a DNA polymerase at the molecular level.

Eva Freisinger, University of Zurich, Switzerland

High-fidelity DNA polymerases accomplish the accurate replication of DNA and are crucial for the maintenance of a functional genome. The encounter of modified or damaged DNA generally either leads to disengagement of the polymerase from the primer/template strand allowing for the repair machinery to proceed, or the enzyme may be able to bypass the DNA lesion itself (translesion synthesis) with a lesion and polymerase specific fidelity. Understanding these events is crucial for elucidating the mechanism by which DNA damage can result in miscoding, leading to mutagenesis and carcinogenesis. We were able to obtain crystal structures of a pol a family DNA polymerase from bacteriophage RB69 in complex with DNA containing the two most prevalent, spontaneously generated premutagenic lesions, an abasic site and 2'-deoxy-7,8-dihydro-8-oxoguanosine (8-oxodG).[1] While the complex with 8-oxodG resembles closely the structure with unmodified DNA, the complex with the abasic site features the polymerase in the catalytically inactive 'open' conformation. The latter structure is a remarkable new finding as so far this 'open' conformation was only observed for the enzyme in the apo state or in editing mode, but never with a DNA template bound to the polymerase active site. The new insights gained from the presented structures permit us for the first time to explain at the molecular level (a) the relative ease and fidelity by which the 8-oxodG lesion is bypassed and (b) the blocking effect of an abasic site to DNA synthesis, herewith increasing our knowledge of miscoding, a first step in mutagenesis.

[1] Freisinger E, Grollman AP, Miller H, Kisker C. EMBO J. 2004, 23(7):1494-1505.