Guanine Oxidation in Double-Stranded DNA by Mn-TMPyP/KHSO₅:  5,8-Dihydroxy-7,8-dihydroguanine Residue as a Key Precursor of Imidazolone and Parabanic Acid Derivatives

Abstract

The mechanism of oxidation of guanine residues on double-stranded oligonucleotides (ODNs) by the chemical nuclease Mn-TMPyP/KHSO5 is reported. By using HPLC coupled to an electrospray mass spectrometer (ESI/MS) the different oxidized ODN strands were directly analyzed, and labeling experiments in H2 18O allowed us to propose a two-electron oxidation mechanism for guanine residues engaged in double-stranded DNA. We found that the imidazolone derivative (dIz) was formed by trapping of a guanine-cation by a water molecule. Two reaction intermediates on the pathway of the formation of dIz were observed: 5,8-dihydroxy-7,8-dihydroguanine and an oxidized guanidinohydantoin intermediate. Furthermore, a secondary route of guanine oxidation leading to parabanic acid was also evidenced. The mechanism of the different routes of guanine oxidation in double-stranded DNA has been discussed in detail.

In Vivo Bypass Efficiencies and Mutational Signatures of the Guanine Oxidation Products 2-Aminoimidazolone and 5-Guanidino-4-nitroimidazole

“…The Iz derivative is a major and ubiquitous in vitro product of guanine oxidation. In addition to forming as a result of ONOO Ϫ oxidation (28), this lesion also forms when guanine is oxidized by other reactants including hydroxyl radical (42), Mn-TMPyp/KHSO 5 (a chemical nuclease) (43)(44)(45), and riboflavin (photooxidation) (40 -42, 46). Direct formation of Iz from 8-oxoG has also been observed (40,47).…”

Use of Short Duplexes for the Analysis of the Sequence‐Dependent Cleavage of DNA by a Chemical Nuclease, a Manganese Porphyrin

“…The products of the reaction were identified by their mass compared to the mass of the initial material, with the latter being referred to as M. The major signal on the mass spectra corresponded to the doubly charged species [MÀ2 H] 2À . From previous work it was known that guanine was the only base damaged with Mn-TMPyP/KHSO 5 [32] and the masses of the oligonucleotides carrying one guanine lesion are reported www.chembiochem.org in Scheme 5. Direct DNA cleavage due to a C5' hydroxylation mechanism can be unambiguously identified by the mass of the corresponding DNA fragments.…”

“…The initial GG strand eluted at a retention time of t R = 40 min with a m/z = 909.75 amu and z = 2 ( Table 3). The products of guanine oxidation were the imidazolone (3, Scheme 5; t R = 38 min, m/z = 890.35 amu), [39,40] the dehydroguanidinohydantoin (4; t R = 38 min, m/z = 911.75 amu), [36,42,43] and a proposed N-formylamido-iminohydantoin (5; t R = 32.8 min, m/z = 926.55 amu) [32,33] derivative. On this sequence, which was selected to present an oxidation-sensitive GG site, the guanine [a] DM: Mass of the modified oligonucleotide minus the mass of the initial oligonucleotide.…”

“…The third type of chemistry mediated by Mn-TMPyP/KHSO 5 consists of electron abstraction from a guanine base [28][29][30] and the mechanism of guanine oxidation has been studied in detail. [31][32][33][34][35][36] The resulting guanine oxidation products are shown in Scheme 5.…”

Selective Detection of 5‐Methylcytosine Sites in DNA

“…Further oxidation of 8-oxodG occurs in the presence of air, which leads to piperidine-sensitive cleavage sites. [13] Indeed, as depicted in Figure 2 a, reaction of DNA with V 2 O 5 in the presence of LiBr and in the absence of oxygen resulted in a highly specific reaction at the C5ÀC6 double bond of the 5Me dC base. No reaction at any other nucleobase was detected, even at the rather high temperature of 80 8C required for full conversion.…”