title: Advances in Structural and Single-Molecule Methods for Investigating DNA Lesion Bypass and Repair Polymerases.
abstract: Innovative advances in X-ray crystallography and single-molecule biophysics have yielded unprecedented insight into the mechanisms of DNA lesion bypass and damage repair. Time-dependent X-ray crystallography has been successfully applied to view the bypass of 8-oxo-7,8-dihydro-2'-deoxyguanine (8-oxoG), a major oxidative DNA lesion, and the incorporation of the triphosphate form, 8-oxo-dGTP, catalyzed by human DNA polymerase β. Significant findings of these studies are highlighted here, and their contributions to the current mechanistic understanding of mutagenic translesion DNA synthesis (TLS) and base excision repair are discussed. In addition, single-molecule Förster resonance energy transfer (smFRET) techniques have recently been adapted to investigate nucleotide binding and incorporation opposite undamaged dG and 8-oxoG by Sulfolobus solfataricus DNA polymerase IV (Dpo4), a model Y-family DNA polymerase. The mechanistic response of Dpo4 to a DNA lesion and the complex smFRET technique are described here. In this perspective, we also describe how time-dependent X-ray crystallography and smFRET can be used to achieve the spatial and temporal resolutions necessary to answer some of the mechanistic questions that remain in the fields of TLS and DNA damage repair.
authors.affiliation: Department of Chemistry and Biochemistry and The Ohio State Biochemistry Program, The Ohio State University , Columbus, Ohio 43210, United States.
authors.forename: Austin T
authors.initials: AT
authors.lastname: Raper
authors.affiliation: Department of Chemistry and Biochemistry and The Ohio State Biochemistry Program, The Ohio State University , Columbus, Ohio 43210, United States.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Reed
authors.affiliation: Department of Chemistry and Biochemistry and The Ohio State Biochemistry Program, The Ohio State University , Columbus, Ohio 43210, United States.
authors.forename: Varun V
authors.initials: VV
authors.lastname: Gadkari
authors.affiliation: Department of Chemistry and Biochemistry and The Ohio State Biochemistry Program, The Ohio State University , Columbus, Ohio 43210, United States.
authors.forename: Zucai
authors.initials: Z
authors.lastname: Suo
mesh_terms: D018360:Crystallography, X-Ray; D004249:DNA Damage; D004260:DNA Repair; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry*; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans
chemical_list: D004259:DNA-Directed DNA Polymerase