ABSTRACT
Glycoxidation and lipoxidation reactions contribute to the chemical modification of proteins during the Maillard reaction. Reactive oxygen species, produced during the oxidation of sugars and lipids in these processes, irreversibly oxidize proteins. Methionine is particularly susceptible to oxidation, yielding the oxidation product methionine sulfoxide (MetSO). Here we describe a method for the analysis of MetSO using proteomic techniques. Using these techniques, we measured MetSO formation on the model protein RNase during aerobic incubations with glucose and arachidonate. We also evaluated the susceptibility of MetSO to reduction by NaBH4), a commonly used reductant in the analysis of Maillard reaction products.
Subject(s)
Methionine/analogs & derivatives , Proteome , Aerobiosis , Carbohydrates , Glycosylation , Lipid Peroxidation , Lipids , Maillard Reaction , Mass Spectrometry , Oxidation-Reduction , Reactive Oxygen Species , Ribonucleases/chemistryABSTRACT
RNase A (1 mM) was incubated with glucose (0.4 M) at 37 degrees C for up to 14 days in phosphate buffer (0.2 M, pH 7.4), digested with trypsin and analysed by LC-MS. The major sites of fructoselysine formation were Lys(1), Lys(7), Lys(37) and Lys(41). Three of these sites (Lys(7), Lys(37) and Lys(41)) were also the major sites of N epsilon-(carboxymethyl)lysine formation.