Reducing sugars that mediate the crosslinks between the amine groups of proteins are decomposed to stable, yellow fluorescent compounds called Advanced Glycation Endproducts (AGEs) via the Maillard (or glycation) reaction. On the pathway to AGEs, alpha-dicarbonyl compounds, such as 3-deoxyglucosone, methylglyoxal, and glyoxal, are detected. These alpha-dicarbonyl compounds are more reactive to the amine groups than the sugars and are often found in diabetes mellitus, cataracts, aging, Alzheimer's disease, and other disease states. In the current research period, the generation and properties of protein-bound free radicals by glyoxal and glycolaldehyde have been investigated under physiological conditions.Bovine serum albumin (BSA) when incubated with glyoxal or glycolaldehyde showed that inter- and/or intramolecular crosslinking occurred in the presence of the carbonyl compounds as revealed by SDS-PAGE analysis. The crosslinked BSAs demonstrated EPR signals, indicating that radical species were generated during the crosslinking process. UV-VIS investigation with Cu,Zn-SOD suggested that superoxide anion radical was also formed during the crosslinking process and the protein-bound free radicals were generated by one-electron transfer from the one-electron reduced form of the free radical to the dioxygen molecule under physiological conditions. It was also found that the radical centers behave like enzyme active centers having oxidation-reduction capabilities. Model reactions between the carbonyl compounds and amine group-containing amino acids have been performed to shed light on the detailed structures of the free radical centers. EPR studies of the model reactions suggest that glyoxal and glycolaldehyde in protein solutions generate pyrazine-cation radical species that form inter- or/and intramolecular crosslinks between N-terminal and N-terminal, N-terminal and the sidechain of the lysine residue, and the sidechains of lysine residues. Additionally, glyoxal may form another kind of protein-bound radical center cross-linking the sidechains of lysine and arginine residues.
