Formation of Nepsilon-(succinyl)lysine in vivo: a novel marker for docosahexaenoic acid-derived protein modification.

Free radical-catalyzed peroxidation of docosa- hexaenoic acid (DHA, C22:6/v-3) generates various lipid peroxidation products that covalently modify biomolecules such as proteins. Under a free radical-generating system, DHA significantly modified lysine residues in bovine serum albumin. Upon incubation of oxidized DHA with an amino- compound pyridoxamine or a lysine-containing peptide, N- propanoyl and N-succinyl adducts were determined to be the major modification products. The hydroperoxide levels in the oxidized DHA closely reflected the formation of the N e -(succinyl)lysine (SUL) upon reaction with the peptide, indicating that the hydroperoxides of DHA represent a po- tential pathway for the formation of SUL. To detect the DHA-derived protein modification in vivo, we developed a monoclonal antibody (mAb2B12) specific to SUL and found that the antibody specifically reacts with the SUL moiety. The formation of SUL was then immunochemically demon- strated in the liver of mice fed with DHA followed by intra- peritoneal injection of carbon tetrachloride (CCl4), a hepatic lipid peroxidation model. Immunoreactive materials with mAb2B12 were observed in the DHA1 CCl4 group, but were not significant in the control, DHA-alone, and CCl4-alone groups. These data suggest that the formation of DHA- derived adducts such as SUL may be implicated in the oxi- dative damage observed in DHA-enriched tissues.—Kawai, Y., H. Fujii, M. Okada, Y. Tsuchie, K. Uchida, and T. Osawa. Formation of N e -(succinyl)lysine in vivo: a novel marker for docosahexaenoic acid-derived protein modification. J.Lipid Res. 2006. 47: 1386-1398.