N-EPSILON-(CARBOXYMETHYL)LYSINE IS A DOMINANT ADVANCED GLYCATION END-PRODUCT (AGE) ANTIGEN IN TISSUE PROTEINS

Advanced glycation end products (AGEs) and glycoxidation products are formed during Maillard or browning reactions between sugars and protei ns and are implicated in the pathophysiology of aging and the complica tions of diabetes. To determine the structure of AGEs, antibodies were prepared to protein browned by incubation with glucose and used in EL ISA assays to measure AGEs formed in model reactions between bovine se rum albumin (BSA) or N-alpha-acetyllysine and glucose, fructose, or gl yoxal. AGEs were formed from glucose and fructose only under oxidative conditions, but from glyoxal under both oxidative and antioxidative c onditions. Gel permeation chromatographic analysis indicated that a si milar AGE was formed in reactions of N-alpha-acetyllysine with glucose , fructose, and glyoxal and that this AGE co-eluted with authentic N-a lpha-acetyl-N-epsilon-(carboxymethyl)lysine. Amino acid analysis of AG E proteins revealed a significant content of N-epsilon-(carboxymethyl) lysine (CML). In ELISA assays using polyclonal antibodies against AGE proteins, CML-BSA (similar to 25 mol of CML/mol of BSA), prepared by c hemical modification of BSA, was a potent inhibitor of the recognition of AGE proteins and of AGEs in human lens proteins. We conclude that AGEs are largely glycoxidation products and that CML is a major AGE re cognized in tissue proteins by polyclonal antibodies to AGE proteins.