Products of Cu(II)-catalyzed oxidation of the N-terminal fragments of alpha-synuclein in the presence of hydrogen peroxide.

Reactive oxygen species (ROS) may provide the covalent modifications of amino acid residues in proteins, formation of protein-protein cross-linkages, and oxidation of the protein backbone resulting in protein fragmentation. In an attempt to elucidate the products of the copper(II)-catalyzed oxidation of the (1-17), (1-28), (1-39) and (1-39)(A30P) fragments of alpha-synuclein, the high performance liquid chromatography (HPLC) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) methods and Cu(II) /hydrogen peroxide as a model oxidizing system were employed. Peptide solution (0.50 mM) was incubated at 37 degrees C for 24 h with metal:peptide:hydrogen peroxide molar ratio 1:1:4 in phosphate buffer, pH 7.4. Oxidation targets for all peptide studied are the methionine residues (M(1), M(5)). Incubation 24 h of the (1-28), (1-39) and (1-39)(A30P) fragments in aerobic conditions lead to the oxidation of one methionine residue to methionine sulfoxide. Reaction of hydrogen peroxide with all fragments of alpha-synuclein resulted in oxidation of two methionine residues (M(1), M(5)) to methionine sulfoxides. For the Cu(II):peptide:hydrogen peroxide 1:1:4 molar ratio systems the further oxidation of methionine residues to sulfone was observed. The cleavage of the peptide bond M(1)-D(2) for all peptides studied was observed as metal binding residues. For the (1-39) and (1-39)(A30P) fragments of alpha-synuclein the molecular ions with lower molecular masses (A(11)-Y(39), E(13)-Y(39)) were also detected.