Copper generated reactive oxygen leads to formation of lysine–DNA adducts

This work describes the addition of a lysine derivative to guanine base in a nucleoside, an oligonucleotide, and to a large DNA that occurs via oxidation by copper generated reactive oxygen species. Nucleophiles present during oxidation leads to the formation of adducts. In this work, 2′-deoxyguanosine is oxidized by copper generated reactive oxygen species in the presence of a lysine derivative, Nα-acetyl-lysine methyl ester. Under these conditions the guanidinohydantoin–lysine adduct is observed in a relative yield of 27% when compared to other guanine oxidation products. MS2 strongly supports that lysine is added to the 5-position during the formation of guanidinohydantoin–lysine. A fourteen-nucleotide DNA duplex was oxidized under similar conditions. Digestion showed formation of the same guanidinohydantoin–lysine nucleoside. The reaction was then examined on a 392-nucleotide DNA substrate. Oxidation in the presence of the lysine ester showed adduct formation as stops in a primer extension assay. Adducts predominately formed at a 5′-GGG at position 415. Six of the seven sites that showed reaction greater than 3-fold above background were guanine sites. We conclude from this study that copper can catalyze the formation of DNA–protein adducts and may form in cells with elevated copper and oxidative stress.

Graphical abstractGuanine oxidation has been shown give a large abundance of modifications including DNA–protein crosslinks. Copper generated reactive oxygen species leads to the formation of DNA–lysine crosslinks in a relative yield of 27% when compared to other guanine oxidation products. The crosslink forms in oligonucleotides and a 392-nucleotide DNA. Since several disease states possess elevated copper-levels and oxidative stress, then it is likely that the formation of these types of crosslinks may occur.Figure optionsDownload full-size imageDownload as PowerPoint slide