title: Chemical repair of base lesions, AP-sites, and strand breaks on plasmid DNA in dilute aqueous solution by ascorbic acid.
abstract: We quantified the damage yields produced in plasmid DNA by γ-irradiation in the presence of low concentrations (10-100 μM) of ascorbic acid, which is a major antioxidant in living systems, to clarify whether it chemically repairs radiation damage in DNA. The yield of DNA single strand breaks induced by irradiation was analyzed with agarose gel electrophoresis as conformational changes in closed circular plasmids. Base lesions and abasic sites were also observed as additional conformational changes by treating irradiated samples with glycosylase proteins. By comparing the suppression efficiencies to the induction of each DNA lesion, in addition to scavenging of the OH radicals derived from water radiolysis, it was found that ascorbic acid promotes the chemical repair of precursors of AP-sites and base lesions more effectively than those of single strand breaks. We estimated the efficiency of the chemical repair of each lesion using a kinetic model. Approximately 50-60% of base lesions and AP-sites were repaired by 10 μM ascorbic acid, although strand breaks were largely unrepaired by ascorbic acid at low concentrations. The methods in this study will provide a route to understanding the mechanistic aspects of antioxidant activity in living systems.
authors.affiliation: Department of Nuclear Engineering and Management, School of Engineering, The University of Tokyo, 7-3-1 Hongo, Tokyo 113-8656, Japan.
authors.forename: Kuniki
authors.initials: K
authors.lastname: Hata
authors.forename: Ayumi
authors.initials: A
authors.lastname: Urushibara
authors.forename: Shinichi
authors.initials: S
authors.lastname: Yamashita
authors.forename: Naoya
authors.initials: N
authors.lastname: Shikazono
authors.forename: Akinari
authors.initials: A
authors.lastname: Yokoya
authors.forename: Yosuke
authors.initials: Y
authors.lastname: Katsumura
mesh_terms: D000975:Antioxidants / Q000737:chemistry; D001205:Ascorbic Acid / Q000737:chemistry*; D053904:DNA Breaks, Single-Stranded / Q000187:drug effects*; D004260:DNA Repair; D004269:DNA, Bacterial / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects*; D004305:Dose-Response Relationship, Drug; D004587:Electrophoresis, Agar Gel; D004926:Escherichia coli / Q000235:genetics; D005720:Gamma Rays; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects*; D011837:Radiation-Protective Agents / Q000737:chemistry; D012996:Solutions / Q000378:metabolism; D014867:Water / Q000378:metabolism
chemical_list: D000975:Antioxidants; D004269:DNA, Bacterial; D011837:Radiation-Protective Agents; D012996:Solutions; D014867:Water; D001205:Ascorbic Acid