Abstract：
Prolonged non-enzymatic glycation of proteins results in the formation of advanced glycation end products (AGEs) that cause several diseases. The glycation of Ribonuclease A (RNase A) at pH 7.4 and 37 °C with ribose, glucose and fructose has been monitored by UV-vis, fluorescence, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and matrix assisted laser desorption ionization spectroscopy-time of flight (MALDI-TOF) methods. The enzymatic activity and DNA binding ability of glycated RNase A was also investigated by an agarose gel-based assay. A precipitation assay examined the ribonucleolytic activity of the glycated enzyme. An increase in incubation time resulted in the formation of high molecular weight AGEs with a decrease in ribonucleolytic activity. Ribose exhibits the highest potency as a glycating agent and showed the greatest reduction in the ribonucleolytic activity of the enzyme. Interestingly, glycated RNase A was unable to bind with the ribonuclease inhibitor (RI) and DNA. The glycated form of the protein was also found to be ineffective in DNA melting unlike native RNase A.
摘要：
蛋白质的长期非酶糖基化导致导致几种疾病的晚期糖基化终产物(AGEs)的形成。核糖核酸酶A（RNaseA）在pH7.4和37°C下用核糖糖化，葡萄糖和果糖已通过UV-vis监测，荧光，十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）和基质辅助激光解吸电离光谱-飞行时间（MALDI-TOF）方法。还通过基于琼脂糖凝胶的测定法研究了糖化RNaseA的酶活性和DNA结合能力。沉淀测定检查了糖化酶的核糖核酸分解活性。孵育时间的增加导致高分子量AGEs的形成，而核糖核酸分解活性降低。核糖表现出作为糖化剂的最高效力，并显示出酶的核糖核酸分解活性的最大降低。有趣的是,糖化核糖核酸酶A不能与核糖核酸酶抑制剂（RI）和DNA结合。与天然RNA酶A不同，还发现蛋白质的糖基化形式在DNA解链中无效。
