Abstract
Protein modification caused by polyunsaturated fatty acid peroxidation products was examined by Western blotting using monoclonal antibodies raised against major lipd peroxidation products. Reaction mixture contained model protein (BSA or Rubisco), C18 unsaturated fatty acid methyl ester (methyl oleate (18:1), methyl linoleate (18:2) or methyl linolenate (18:3)) in the presence of a metal-catalyzed oxidation system (Fe(III)/ascorbate/O2). In results, the most effective protein modifier was methyl linolenate which caused protein adducts derived from peroxidation products such as malondialdehyde (OHCCH2CHO), acrolein (CH2:CHCHO) and crotonaldehyde (CH3CH:CHCHO). The monoclonal antibodies reacted with several proteins in cucumber leaves, and internal amino acid sequences analysis showed that they were chloroplast-localizing proteins. Therefore chloroplasts, which contain most abundant linolenate as a membrane component among plant organellar, may be a major site occurring protein modification by polyunsaturated fatty acid peroxidation.
