Oxidative DNA damage

Molecular Life Sciences (2014)


Reaction of DNA with reactive oxygen species


Life in an aerobic environment comes with a cost, like that of oxidative damage to several kinds of macromolecules. Partially reduced oxygen species (superoxide anion, hydrogen peroxide) react with transition metals to generate hydroxyl radicals. Nitric oxide, peroxynitrite, and other species can also be involved in DNA damage. Ionizing radiation also forms hydroxyl radical. The most abundant product arising from DNA oxidation is 8-oxo-7,8-dihydroguanne, but a plethora of oxidation products is also formed, and miscoding properties of all have not been characterized. In addition to the damage to the DNA bases, the individual five carbons of the deoxyribose sugar ring can all be attacked by radicals, yielding products with split sugar rings and in some cases DNA fragmentation.


A considerable amount of literature exists on oxidative damage to DNA, and this section is not intended to be comprehensive (see Cadet et al 2010; DeMott and Dedon 2010). More than 80 oxidized DNA bases have been reported in the literature; З19 of these have been reported in cellular DNA. A considerable number of oxidation products of the sugar ring (20-deoxyribose) are also known. Many of the oxidized bases may be artifacts in the sense that they are not relevant to biological systems. Another issue is that some of the analytical systems have been notorious for introducing oxidative artifacts during the analyses. Nevertheless, there is general agreement that (i) a load of DNA lesions of 1/106 bases resulting from oxidative stress reactions is common in biological settings and (ii) levels of some of the lesions can increase dramatically under certain conditions of stress.

Chemistry and consequences

Numerous oxidants are involved as reaction oxygen species and determine the course of which products are formed. The term “reactive oxygen species” (ROS) includes superoxide anion (O2—), hydrogen peroxide (H2O2), hydroxyl radical (OH), and singlet oxygen 1O2 (1Dg state). g-radiation is a source of OH·, in that it produces homolytic cleavage of H2O as it passes through biological systems. OH · can also be generated in the transition metal-dependent reaction of O2— and H2O2 (Fig. 1a).

Oxidative DNA damage 1

Fig. 1.  (a) The Haber-Weiss and Fenton reactions. Together these lead to hydroxyl radical production. (b) 1-Electron oxidations of (a) deoxyguanosine (dGuo) and (b) thymidine (dThd) (Cadet et al 2010)

Lesions known to be formed in the process of g-radiation (in human monocyte cells) include cis- and trans-5,6-dihydroxy-5,6-dihydrothymidine (thymidine glycol), 5-(hydroxymethyl)- 20-deoxyuridine, 5-formyl-20-deoxyuridine, 8-oxo-7,8-dihydro-20-deoxyguanosine (8-oxo dGuo), 2-6-diamino-4-hydroxy-5-formamidopyrimidine (FAPY dGuo), 8-oxo-7,8-dihydro- 20-deoxyadenosine (8-oxo dAdo), and 4,6-deamino-5-formamidopyrimidine (FAPY dAdo) (Cadet et al. 2010). Singlet oxygen can be produced by “type II” photosensitizers (some used in therapy) and some enzymes, e.g., myeloperoxidase.

In addition, myeloperoxidase (elevated in infiammation-related disease states) generates HOCl. HOCl chlorinates bases, yielding 5-chlorocytosine, 8-chloroguanine, and 8-chloroadenine.

1-Electron oxidation is the result of ionizing radiation, electron abstraction by “type I” photosen- sitizers, high-intensity ultraviolet (UV) damage, and oxidants such as KBrO3 and CO3— (Cadet et al 2010). The products can react with a proton or H2O to generate various products (Fig. 1b).

Most attention has been given to oxidized bases derived from reaction with OH·. Reaction with thymidine produces a variety of products (Fig. 2a). Reaction of OH · with deoxyguanosine also yields a mixture of products (Fig. 2b, c). Similarly, reaction of OH · with deoxyadenosine produces products similar to those found with deoxyguanosine (Fig. 2d).

Some other unusual oxidation reactions have been reported. One example is the (deoxyribose) C40 oxidation of deoxycytidine, which leads to an exocyclic deoxycytidine adduct (Fig. 3).

Oxidative DNA damage 2

Fig. 2. Reactions of hydroxyl radical (Cadet et al. 2010). (a) Reactions of thymidine and hydroxy radical. (b) Reaction of deoxyguanosine with hydroxyl radical. (c) Reaction of deoxyguanosine with hydroxyl radical and further reaction with superoxide anion. (d) Reaction of deoxyadenosine with hydroxyl radical

In addition, hydrogen atom abstraction of a purine nucleoside leads to 50,8-cycloadducts of deoxyguanosine and deoxyadenosine (Cadet et al. 2010), although only traces of these adducts have been characterized in cellular DNA.

Oxidative DNA damage 3

Fig. 3. Reaction of deoxycytidine with a neighboring base propenal in DNA to general a crosslink, followed by enzymatic digestion (Cadet et al. 2010)

In addition to oxidized bases, oxidation of the sugar ring can be extensive and complex. Oxidative damage to the deoxyribose ring generally results in strand cleavage of some type. Most reactions generate a mixture of types of damage, and the analytical chemistry is complex. However, some reagents react selectively with individual products, e.g., hydrazine with a 20-deoxypentos-4-ulose abasic site (Fig. 4; DeMott and Dedon 2010), and recent methods in liquid chromatography-mass spectrometry (LC-MS) have provided not only characterization and quantitation of individual products but also permitted the determination of sites of damage within oligonucleotides (Chowdhury and Guengerich 2009).

Oxidation of the sugar occurs via initial hydrogen abstraction at any of the five carbons. In some cases, multiple pathways can proceed even from hydrogen atom abstraction at a single carbon, and the course of the oxidation is a function of which oxidant is used. More complex oxidants such as bleomycin-based systems are relatively selective, but Fenton-based methods yield complex mix- tures (Chowdhury and Guengerich 2009). A summary of the reactions associated with hydrogen abstraction at each carbon of the deoxyribose ring is shown in Fig. 4 (DeMott and Dedon 2010).

Oxidation at C10 yields an abasic site plus a lactone ring. In this case, the strand is not cleaved.

C20 oxidation yields an erythrose abasic site, but, as in the case of C10 oxidation, the strand remains intact in the absence or further processing. However, the changes in the chemistry of the rings (both with C10 and C20 oxidation) do not allow for efficient repair of the abasic site because of the nature of the damage.

In the cases of oxidation at the C30, C40, and C50 positions, the DNA strand is broken. C30 oxidation can yield either (i) a 3-phosphoglycoaldehyde plus base propenoate or (ii) a 30-keto-2- deoxynucleotide, which can undergo b/d elimination to 2-methylene-3(2H) furanone.

Oxidation at C40 yields either (i) a deoxypentos-4-ulose abasic site (the strand remains intact, as with C0  and C20  oxidations) or (ii) a 30-phosphoglycolate and either (a) a base propenal or (b) malondialdehyde plus a free base.

C50 oxidation yields either (i) a nucleoside 50-aldehyde, which can undergo b/d elimination to furfural, or (ii) a 30-formylphosphate plus a 20-phosphoryl-1,4-dioxo-2-butane, which can undergo b-elimination to trans-butene dialdehyde.

Reactive nitrogen species (RNS) are also oxidants and known to oxidize several bases (Fig. 5) (DeMott and Dedon 2010). These will be treated at further length under section g and include nitric oxide (NO·), peroxynitrite (ONO2—, the product of reaction of NO · with O2—), and peroxycarbonate (ONO2CO2—, the product of reaction of ONO2— and CO2). ONO2— yields primarily deoxyribose oxidation, and ONO2CO2— yields primarily the oxidation and nitration of guanine (DeMott and Dedon 2010). N2O3 is primarily a deaminating species.

Oxidative DNA damage 4

Fig. 4. Oxidation reactions proceeding from hydrogen abstraction at each of the five carbons of deoxyribose in DNA (DeMott and Dedon 2010)

As mentioned earlier, many more base oxidation products have been reported from model systems, etc. than identified in cellular DNA, and the analytical chemistry is challenging. For example, oxanine (or oxanonsine) has never been detected in a biological system (DeMott and Dedon 2010). Another controversial area is secondary oxidation products of bases. The oxidation of 8-oxoG to spirohydantoins has been characterized in model chemical systems but only once in cells that being in (DNA glycosylase) Nei-deficient Escherichia coli treated with CrVI (Slade et al. 2005) and questions have been raised regarding the experimental approach (Cadet et al 2010). Traces of the adduct and the related guanidinodihydantoin were reported in a Rag2—/— mouse model (Mangerich et al. 2012).

Oxidative DNA damage 5

Fig. 5. Reactions of DNA bases (dGuo and deoxyadenosine, dAdo) with N2O3 (DeMott and Dedon 2010)

In conclusion, the course of oxidative damage is a function of the oxidant. Many types of base and sugar damage have been characterized, although it is not clear that all are relevant in vivo. The analytical chemistry is associated with the analysis of the oxidation products which is challenging, and caution is advised before embarking on studies in this area. Finally, the point should be made that organisms have a variety of means of suppressing oxidative stress and even dealing with some of the types of damaged DNA, e.g., 8-oxoguanine.


Cadet J, Douki T, Ravanat J-L (2010) Oxidatively generated damage to isolated and cellular DNA. In: Broyde S, Geacintov NE (eds) The chemical biology of DNA damage. Wiley-VCH Verlag, Weinheim, pp 53–80

Chowdhury G, Guengerich FP (2009) Tandem mass spectrometry-based detection of C40-oxidized abasic sites at specific positions in DNA fragments. Chem Res Toxicol 22:1310–1319

DeMott MS, Dedon PC (2010) Chemistry of infiammation and DNA damage: biological impact of reactive nitrogen species. In: Broyde S, Geacintov NE (eds) The chemical biology of DNA damage. Wiley-VCH Verlag, Weinheim, pp 21–80

Mangerich A, Knutson CG, Parry NM et al (2012) Infection-induced colitis in mice causes dynamic and tissue-specific changes in stress response and DNA damage leading to colon cancer. Proc Natl Acad Sci U S A 109:E1820–E1829

Slade PG, Hailer MK, Martin BD et al (2005) Guanine-specific oxidation of double-stranded DNA by Cr(VI) and ascorbic acid forms spiroiminodihydantoin and 8-oxo-20-deoxyguanosine. Chem Res Toxicol 18:1140–1149