title: Identification of the C4'-oxidized abasic site as the most abundant 2-deoxyribose lesion in radiation-damaged DNA using a novel HPLC-based approach.
abstract: A novel analytical high-performance liquid chromatography (HPLC)-based method of quantification of the yields of C4'-oxidized abasic sites, 1, in oxidatively damaged DNA has been elaborated. This new approach is based on efficient conversion of 1 into N-substituted 5-methylene-Δ(3)-pyrrolin-2-ones, 2, upon treatment of damaged DNA with primary amines in neutral or slightly acidic solutions with subsequent quantification of 2 by HPLC. The absolute and relative radiation-chemical yields of 1 in irradiated DNA solutions were re-evaluated using this method. The yields were compared with those of other 2-deoxyribose degradation products including 5-methylene-2(5H)-furanone, malondialdehyde, and furfural resulting from the C1', C4' and C5'-oxidations, respectively. The yield of free base release (FBR) determined in the same systems was employed as an internal measure of the total oxidative damage to the 2-deoxyribose moiety. Application of this technique identifies 1 as the most abundant sugar lesion in double-stranded (ds) DNA irradiated under air in solution (36% FBR). In single-stranded (ss) DNA this product is second by abundance (33% FBR) after 2-deoxyribonolactones (C1'-oxidation; 43% FBR). The production of nucleoside-5'-aldehydes (C5'-oxidation; 14% and 5% FBR in dsDNA and ssDNA, respectively) is in the third place. Taken together with the parallel reaction channel that converts C4'-radicals into malondialdehyde and 3'-phosphoglycolates, our results identify the C4'-oxidation as a prevalent pathway of oxidative damage to the sugar-phosphate backbone (50% or more of all 2-deoxyribose damages) in indirectly damaged DNA.
authors.affiliation: a Department of Chemistry, East Tennessee State University, Johnson City, Tennessee.
authors.forename: Marina
authors.initials: M
authors.lastname: Roginskaya
authors.forename: Reza
authors.initials: R
authors.lastname: Mohseni
authors.forename: Terence J
authors.initials: TJ
authors.lastname: Moore
authors.forename: William A
authors.initials: WA
authors.lastname: Bernhard
authors.forename: Yuriy
authors.initials: Y
authors.lastname: Razskazovskiy
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004247:DNA / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004249:DNA Damage; D003855:Deoxyribose / Q000378:metabolism*; D005720:Gamma Rays / Q000009:adverse effects*; D010084:Oxidation-Reduction / Q000528:radiation effects
chemical_list: D003855:Deoxyribose; D004247:DNA