A Single Nuclease-Resistant Linkage in DNA as a Versatile Tool for the Characterization of DNA Lesions: Application to the Guanine Oxidative Lesion “G+34” Generated by Metalloporphyrin/KHSO5 Reagent
journal contributionposted on 19.11.2012, 00:00 by Agnieszka Tomaszewska, Sophie Mourgues, Piotr Guga, Barbara Nawrot, Geneviève Pratviel
The oxidation of an oligonucleotide containing a single nuclease-resistant phosphodiester link, a stereoisomerically pure methylphosphonate, by manganese (Mn-TMPyP) or iron (Fe-TMPyP) porphyrin associated to KHSO5 allowed the isolation and characterization of a guanine lesion corresponding to an increase of mass of 34 amu as compared to guanine (“G+34”), namely, 5-carboxamido-5-formamido-2-iminohydantoin. Enzymatic digestion of the damaged oligonucleotide afforded, apart from the undamaged nucleotide monomer pool, a unique dinucleotide doubly modified with a methylphosphonate and an oxidized guanine base that is suitable for NMR analysis. The method can be applied to the study of any DNA lesion. More importantly, the method can be extended to the analysis of DNA damage in a sequence context. Any preselected residue in a DNA sequence may be individually analyzed by the easy introduction of a single nuclease-resistant link at the 3′- or 5′-position.