Ansari Nadeem A, Alam Khursheed, Ali Asif
Department of Biochemistry, JN Medical College, Faculty of Medicine, AMU, Aligarh, India.
Hum Immunol. 2009 Jun;70(6):417-24. doi: 10.1016/j.humimm.2009.03.015. Epub 2009 Mar 28.
This study analyzes effect of glycation on proteins rich in lysine residues as hyperglycemia induced protein glycation has been mainly reported in diabetes mellitus at the intrachain lysine residues leading to the formation of Amadori modified proteins. We have studied the effect of glucose on poly-l-lysine (PLL), a homopolymer of lysine residues. Levels of Amadori products in the glycated PLL were evaluated by fructosamine assay and the presence of 5-hydroxymethylfurfural (HMF) in the glycated PLL was analyzed by thiobarbituric acid assay. Fluorescence and FT-IR spectroscopy were applied to characterize the modified PLL. Binding characteristics of experimentally induced antibodies against glycated PLL and the presence of antibodies against glycated PLL in the sera of diabetes patients was evaluated by solid phase enzyme immunoassays. The fructosamine assay showed significantly high yield of early glycation (Amadori) products in the glycated PLL, which was confirmed by increased yield of HMF from Amadori products of glycated PLL. Loss in fluorescence intensity and appearance of a new band corresponding to Amadori products were observed in FT-IR spectrum of the glycated PLL. Glycated PLL was found to be highly immunogenic in rabbits as compared to the native form. Serum antibodies from diabetes patients showed appreciably high recognition of the glycated PLL. The results conclusively show the glycation induced damage to the lysine molecules and specific recognition of Amadori-lysine residues by serum antibodies from diabetes patients. The glycated lysine residues may serve as a diagnostic biomarker for early glycation process in diabetes mellitus.
本研究分析了糖基化对富含赖氨酸残基蛋白质的影响，因为高血糖诱导的蛋白质糖基化主要在糖尿病中报道，发生在链内赖氨酸残基上，导致形成阿马多里修饰蛋白。我们研究了葡萄糖对聚-L-赖氨酸（PLL）（赖氨酸残基的均聚物）的影响。通过果糖胺测定法评估糖基化PLL中阿马多里产物的水平，并通过硫代巴比妥酸测定法分析糖基化PLL中5-羟甲基糠醛（HMF）的存在情况。应用荧光和傅里叶变换红外光谱（FT-IR）对修饰后的PLL进行表征。通过固相酶免疫测定法评估实验诱导的抗糖基化PLL抗体的结合特性以及糖尿病患者血清中抗糖基化PLL抗体的存在情况。果糖胺测定法显示糖基化PLL中早期糖基化（阿马多里）产物的产量显著升高，糖基化PLL的阿马多里产物中HMF产量的增加证实了这一点。在糖基化PLL的FT-IR光谱中观察到荧光强度降低以及出现对应于阿马多里产物的新谱带。与天然形式相比，发现糖基化PLL在兔子中具有高度免疫原性。糖尿病患者的血清抗体对糖基化PLL表现出明显较高的识别度。结果确凿地表明糖基化诱导了赖氨酸分子的损伤，并且糖尿病患者的血清抗体对阿马多里赖氨酸残基具有特异性识别。糖基化的赖氨酸残基可能作为糖尿病早期糖基化过程的诊断生物标志物。
