Chemical and Biological Evidence for Base Propenals as the Major Sourceof the Endogenous M1dG Adduct in CellularDNA

Abstract

The endogenous DNA adduct, M 1 dG, has been shown to arise in vitro in reactions of dG with malondialdehyde (MDA), a product of both lipid peroxidation and 4-oxidation of deoxyribose in DNA, and with base propenals also derived from deoxyribose 4-oxidation. We now report the results of cellular studies consistent with base propenals, and not MDA, as the major source of M 1 dG under biological conditions. As a foundation for cellular studies, M 1 dG, base propenals, and MDA were quantified in purified DNA treated with oxidizing agents known to produce deoxyribose 4-oxidation. The results revealed a consistent pattern; Fe 2؉ -EDTA and ␥-radiation generated MDA but not base propenals or M 1 dG, whereas bleomycin and peroxynitrite (ONOO ؊ ) both produced M 1 dG as well as base propenals with no detectable MDA. These observations were then assessed in Escherichia coli with controlled membrane levels of polyunsaturated fatty acids (PUFA). ONOO ؊ treatment (2 mM) of cells containing no PUFA (defined medium with 18:0/stearic acid) produced 6.5 M 1 dG/10 7 deoxynucleotides and no detectable lipid peroxidation products, including MDA, as compared with 3.8 M 1 dG/ 10 7 deoxynucleotides and 0.07 g/ml lipid peroxidation products with control cells grown in a mixture of fatty acids (0.5% PUFA) mimicking Luria-Bertani medium. In cells grown with linoleic acid (18:2), the level of PUFA rose to 54% and the level of MDA rose to 0.14 g/ml, whereas M 1 dG fell to 1.4/10 7 deoxynucleotides. Parallel studies with ␥-radiation revealed levels of MDA similar to those produced by ONOO ؊ but no detectable M 1 dG. These results are consistent with base propenals as the major source of M 1 dG in this model cell system.

Formation of 1,4-Dioxo-2-butene-Derived Adducts of 2‘-Deoxyadenosine and 2‘-Deoxycytidine in Oxidized DNA

“…Many of these products are electrophilic and capable of reacting with local nucleophiles to form both DNA and protein adducts. Examples include the formation of DNAprotein cross-links when repair proteins interact with the deoxyribonolactone abasic site arising from 1′-oxidation of deoxyribose in DNA (1,2), the formation of M 1 dG, the pyrimidopurinone adduct of dG, in reactions with base propenals arising from 4′-oxidation of deoxyribose (3)(4)(5), and the formation of glyoxal adducts of dG from the 3′-phosphoglycolaldehyde residue that arises from 3′-oxidation of deoxyribose (6). We now extend these observations to include DNA adducts derived from electrophilic products of 5′-oxidation of deoxyribose in DNA (7).…”

Metabolism in Vitro and in Vivo of the DNA Base Adduct, M1G

“…The pyrimidopurinone adduct, M 1 dG, forms from reactions of 2′-deoxyguanosine (dG) with either the DNA peroxidation product, base propenal (8)(9)(10), or the lipid peroxidation product, malondialdehyde (10-12) (Scheme 1). M 1 dG is mutagenic in bacterial and mammalian cells (13)(14)(15)(16) and can be extracted from the genomic DNA of humans and animals (17)(18)(19).…”

N-formylation of lysine in histone proteins as a secondary modification arising from oxidative DNA damage

“…2, oxidation of deoxyribose produces a variety of electrophilic products that have the potential to react with nucleophilic sites in cellular DNA, RNA, and proteins. For example, the base propenal product of 4Ј-oxidation of deoxyribose in DNA has been shown to be a major cellular source of the mutagenic endogenous DNA adduct, M 1 G (19). The present work addresses protein adducts formed by the 3Ј-formylphos-phate residue that arises from oxidation of the 5Ј-position of deoxyribose along with a 5Ј-(2-phosphoryl-1,4-dioxobutane) species (Fig.…”