Mapping three guanine oxidation products along DNA following exposure to three types of reactive oxygen species

Abstract

Reactive oxygen and nitrogen species generated during respiration, inflammation, and immune response can damage cellular DNA, contributing to aging, cancer, and neurodegeneration. The ability of oxidized DNA bases to interfere with DNA replication and transcription is strongly influenced by their chemical structures and locations within the genome. In the present work, we examined the influence of local DNA sequence context, DNA secondary structure, and oxidant identity on the efficiency and the chemistry of guanine oxidation in the context of the Kras protooncogene. A novel isotope labeling strategy developed in our laboratory was used to accurately map the formation of 2,2-diamino-4-[(2-deoxy-β-D-erythropentofuranosyl)amino]- 5(2 H)-oxazolone (Z), 8-oxo-7,8-dihydro-2'-deoxyguanosine (OG), and 8-nitroguanine (8-NO-G) lesions along DNA duplexes following photooxidation in the presence of riboflavin, treatment with nitrosoperoxycarbonate, and oxidation in the presence of hydroxyl radicals. Riboflavin-mediated photooxidation preferentially induced OG lesions at 5' guanines within GG repeats, while treatment with nitrosoperoxycarbonate targeted 3'-guanines within GG and AG dinucleotides. Little sequence selectivity was observed following hydroxyl radical-mediated oxidation. However, Z and 8-NO-G adducts were overproduced at duplex ends, irrespective of oxidant identity. Overall, our results indicate that the patterns of Z, OG, and 8-NO-G adduct formation in the genome are distinct and are influenced by oxidant identity and the secondary structure of DNA.

Photo-Induced Depletion of Binding Sites in DNA-PAINT Microscopy

“…Not only do we see a significantly reduced depletion in high irradiance DNA-PAINT microscopy (I0=0.2 kW/cm2) (Figure 2 and Figure 3), but also in low irradiance SI-FCS measurements (I0=0.018 kW/cm2), we observed a close to zero depletion rate (Figure 4c,d and Table 2). The importance of ROS for photo-induced damages following fluorescence excitation has been studied not only in the context of fluorescence photo-bleaching [18,19,22,27,28,29], but also photo-toxicity [30,31] and in particular DNA damage [32,33,47]. ROS have also been shown to contribute to photo-induced unbinding of proteins [48,49].…”

“…Different ROS, most prominently singlet oxygen and superoxide radicals, are supposedly formed from excited fluorescent states [29,54]. The likelihood and type of induced damage is reported to be sequence dependent and enriched at duplex ends [33]. Previous studies suggest that guanine is preferentially damaged, due to its lowest redox potential among the DNA bases [55,56,57].…”

“…Previous studies suggest that guanine is preferentially damaged, due to its lowest redox potential among the DNA bases [55,56,57]. Not only were oxidation products frequently found in guanine repeats [33,55], but also electron hole diffusion along the DNA leads to guanine oxidation distant from the site of single electron transfer [58,59,60]. On the other hand, adenine and thymine are the predominant bases on the docking strand that are closest to the fluorescent dye upon imager binding.…”

“…Triplet states have been found to play a role in photo-bleaching pathways and to promote the generation of reactive oxygen species (ROS) [19,22,23,24,25,26,27,28]. ROS are not only known to bleach fluorescent dyes [18,29] and have phototoxic effects on biological samples [30,31], but also to damage DNA [32,33].…”

Antimicrobial metal-based nanoparticles: a review on their synthesis, types and antimicrobial action

“…This cleavage induced by the nanoparticles was studied in the pBR322 plasmid in the presence of Ag NPs via electrophoresis [168]. The results showed that guanine is the most affected nucleobase due to its low redox potential, and its oxidation produces a wide variety of modifications that ultimately affect DNA function ( Figure 7) [169]. NPs can not only affect bacteria but also other complex multicellular organisms through the induced genetic damage [170].…”