Detection of dideoxyosone intermediates of glycation using a monoclonal antibody: characterization of major epitope structures.
Glycation or the Maillard reaction in proteins forms advanced glycation end products (AGEs) that contribute to age- and diabetes-associated changes in tissues. Dideoxyosones, which are formed by the long-range carbonyl shift of the Amadori product, are newly discovered intermediates in the process of AGE formation in proteins. They react with o-phenylenediamine (OPD) to produce quinoxalines. We developed a monoclonal antibody against 2-methylquinoxaline-6-carboxylate coupled to keyhole limpet hemocyanin. The antibody reacted strongly with ribose and fructose (+OPD)-modified RNase A and weakly with glucose and ascorbate (+OPD)-modified RNase A. Reaction with substituted quinoxalines indicated that this antibody favored the 2-methyl group on the quinoxaline ring. We used high performance liquid chromatography to isolate and purify three antibody-reactive products from a reaction mixture of N alpha-hippuryl-L-lysine+ribose+OPD. The two most reactive products were identified as diastereoisomers of N1-benzoylglycyl-N6-(2-hydroxy-3-quinoxalin-2-ylpropyl)lysine and the other less reactive product as N1-benzoylglycyl-N6-[2-hydroxy-2-(3-methylquinoxalin-2-yl)ethyl]lysine. Our study confirms that dideoxyosone intermediates form during glycation and offers a new tool for the study of this important pathway in diabetes and aging.
Puttaiah
Shivaprakash
S
Department of Ophthalmology, Case Western Reserve University, Cleveland, OH 44106, USA.
Zhang
Yuming
Y
Pilch
Heather A
HA
Pfahler
Christoph
C
Oya-Ito
Tomoko
T
Sayre
Lawrence M
LM
Nagaraj
Ram H
RH
D000818:Animals; D000911:Antibodies, Monoclonal; D001205:Ascorbic Acid / Q000737:chemistry; D004151:Dipeptides / Q000737:chemistry*; D000939:Epitopes; D005632:Fructose / Q000737:chemistry; D005947:Glucose / Q000737:chemistry; D017127:Glycation End Products, Advanced / Q000737:chemistry*; D006031:Glycosylation; D006433:Hemocyanins / Q000737:chemistry; D051379:Mice; D011810:Quinoxalines / Q000737:chemistry* / Q000276:immunology; D012259:Ribonuclease, Pancreatic / Q000737:chemistry; D012266:Ribose / Q000737:chemistry; D013237:Stereoisomerism
C509426:2-methylquinoxaline-6-carboxylic acid; D000911:Antibodies, Monoclonal; D004151:Dipeptides; D000939:Epitopes; D017127:Glycation End Products, Advanced; C509428:N(1)-benzoylglycyl-N(6)-(2-hydroxy-2-(3-methylquinoxalin-2-yl)ethyl)lysine; C509427:N(1)-benzoylglycyl-N(6)-(2-hydroxy-3-quinoxalin-2-ylpropyl)lysine; D011810:Quinoxalines; D005632:Fructose; D012266:Ribose; D006433:Hemocyanins; D012259:Ribonuclease, Pancreatic; C032808:keyhole-limpet hemocyanin; D005947:Glucose; D001205:Ascorbic Acid