Covalent adducts arising from the decomposition products of lipid hydroperoxides in the presence of cytochrome c

Polyunsaturated fatty acids can be converted to lipid hydroperoxides through nonenzymatic and enzymatic pathways. The prototypic ω-6 lipid hydroperoxide 13-hydroperoxy-octadecadienoic acid (13-HPODE) homolytically decomposes to form highly reactive α,β-unsaturated aldehydes, such as 9,12-dioxo-10(E)-dodecenoic acid (DODE), 4-oxo-2(E)-nonenal (ONE), 4,5-epoxy-2(E)-decenal (EDE), and 4-hydroxy-2(E)-nonenal (HNE), that can form covalent adducts with DNA. Both 4-oxo-2(E)-nonenal and 4-hydroxy-2(E)-nonenal can also modify proteins to form products that can potentially serve as biomarkers of lipid hydroperoxide-mediated macromolecule damage. In this study, cytochrome c was used to identify and individually characterize the modification sites for each of these aldehydes and also determine the most abundant adduct formed following the decomposition of 13-HPODE. The adducts were characterized by ESI-TOF/MS analysis of the intact proteins and by a combination of ESI-ion-trap/MSn and quadrupole-TOF/MS/MS analysis of...