Cleavage of in vitro and in vivo formed lens protein cross‐links by a novel cross‐link breaker
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- 作者：Hollenbach ; Seth ; Thampi ; Prajitha ; Viswanathan ; Tito ; Abraham ; Edathara C.
- 关键词：glycation ; protein cross ; linking ; eye lens ; cross ; link breakers ; thiazolium compounds
- 刊名：Molecular and Cellular Biochemistry
- 出版时间：2003
- 出版年：2003
- 期刊代码：38_03008177
- 类别：cp
- 卷：243
- 期：1/2
- 页码：73-80
- 数据来源：sp
摘要
The purpose of this study was to investigate the effect of N‐phenacyl‐4,5‐dimethylthiazolium bromide (DMPTB), an advanced glycation end product (AGE) cross‐link breaker, on lens protein cross‐links formed in vitro and in vivo . DMPTB was synthesized and its structure confirmed by its NMR spectrum. To show whether DMPTB can inhibit AGE cross‐linking, recombinant human &agr;A‐crystallin was glycated with glucose‐6‐phosphate (G6P) in the presence and absence of DMPTB. Reversal of the already formed cross‐links was studied by treating pre‐glycated &agr;A‐crystallin with DMPTB. The ability of DMPTB to cleave in vivo formed cross‐links was ascertained by treating water‐insoluble protein fractions from diabetic human lenses with this compound. Glycation of &agr;A‐crystallin with G6P showed several high molecular weight (HMW) protein bands on the SDS‐PAGE gel; DMPTB inhibited the formation of these HMW proteins. Molecular sieve HPLC confirmed the inhibition of formation of larger aggregates not separated by SDS‐PAGE. Treatment of pre‐glycated &agr;A‐crystallin with DMPTB gave evidence for the degradation of the already formed cross‐‐linked HMW aggregates. Both molecular sieve HPLC and reverse‐phase HPLC of the water‐insoluble protein fractions from two diabetic human lenses showed that DMPTB could degrade a major portion of the cross‐linked HMW aggregates to lower molecular weight proteins. This suggests that the cross‐linked proteins in human lenses are formed predominantly by the advanced glycation process and cross‐link breakers like DMPTB may have application for the intervention of protein cross‐linking in the eye lens.