alpha- and beta-casein aggregation induced by riboflavin-sensitized photo-oxidation occurs via di-tyrosine cross-links and is oxygen concentration dependent

Type I photo-oxidation generates Trp-(TrpN center dot) and Tyr-derived (TyrO center dot) radicals in proteins which can dimerize producing cross-links, or alternatively react with O-2. It was therefore hypothesized that the O-2 concentration may have a significant effect on dye-photosensitized reactions. We studied photo-oxidation of alpha- and beta-caseins induced by riboflavin (RF), a photosensitizing vitamin present in milk, under aerobic and anaerobic conditions. Triplet-state RF induced oxidative modifications on both caseins, and significant levels of cross-links. The extent of damage, and the yield of cross-links versus oxidized products, was dependent on the O-2 concentration. In the absence of O-2, the overall extent of damage was decreased, but the yield of cross-linked products was significantly elevated. These cross-links are consistent with inter- and intra-molecular di-Tyr or di-Trp bridges. Alternative cross-links were detected in the presence of O-2, consistent with pathways involving the reaction of protein radicals with O-2 or O-2(center dot-).