Differential glycation of rat alpha-, beta- and gamma-crystallins

Crystallin glycation seems to play an important role in the development of diabetic cataract. In order to understand the role of glycation in cataractogenesis, levels of glycation of different crystallins were determined by in vitro glycation of rat lens soluble fraction with 50 mM glucose or glucose-6-phosphate (G6P) for up to 5 days and in streptozotocin-diabetic rats during various stages of cataract development. All samples were reduced with [3H]NaBH4 and the tritium incorporation was taken as a measure of glycation. Proteins were routinely separated by molecular sieve HPLC. In vitro studies with glucose showed that gamma-crystallin was readily glycated and reached a plateau by 3 days, while alpha- and beta-crystallins were glycated slowly initially up to 3 days followed by a steep increase as seen on the fifth day. Incubation with 50 mM G6P resulted in an approximately two fold increase in glycation compared to glucose of all crystallins. In the diabetic animals also gamma-crystallin glycation increased approximately twofold within 15 days after the onset of diabetes and an additional threefold within the next 45 days followed by a slight decrease during the following 90-120 days. Increase in glycation, on the contrary, was very slow up to 30 days for alpha-crystallin and up to 60 days for beta-crystallin, followed by a steep increase during the remainder of the experimental period. The high molecular weight (HMW) aggregates had higher levels of glycation than other proteins; the insoluble HMW aggregates contained higher levels of glycation than the soluble HMW aggregates.(ABSTRACT TRUNCATED AT 250 WORDS)