The glycation-associated crosslinking of lens proteins by ascorbic acid is not mediated by oxygen free radicals.
The reaction by which ascorbic acid (ASA) causes the glycation and crosslinking of lens proteins displays a rigid requirement for the presence of oxygen, and is inhibited by the presence of glutathione. Oxygen is required to oxidize ASA to dehydroascorbic acid (DHA) and other products which are the active glycating species. No evidence could be found to support a role for oxidative protein crosslinking by a free radical mechanism. Crosslinking was not inhibited by blocking protein sulfhydryl groups with iodoacetamide, nor were the protein crosslinks dissociated by boiling with 2% mercaptoethanol prior to SDS-PAGE. The addition of a variety of oxygen free radical quenchers had no effect on the extent of protein crosslinking. In fact, the removal of oxygen from the reaction mixture had no effect on either protein glycation, protein crosslinking or the modification of lysine residues, provided DHA was used as the glycating agent. All of these activities were inhibited, however, if ASA was the glycating agent. This confirms that oxygen is required only to convert ASA to DHA.
Mason Institute of Opthalmology, University of Missouri, Columbia 65212.
M
M
Prabhakaram
B J
BJ
Ortwerth
D000818:Animals; D001205:Ascorbic Acid / Q000494:pharmacology*; D002417:Cattle; D003459:Crystallins / Q000187:drug effects*; D003683:Dehydroascorbic Acid / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D016166:Free Radical Scavengers; D010100:Oxygen; D011485:Protein Binding; D011489:Protein Denaturation / Q000187:drug effects
D003459:Crystallins; D016166:Free Radical Scavengers; D001205:Ascorbic Acid; D010100:Oxygen; D003683:Dehydroascorbic Acid