Nagaraj R H, Monnier V M
Department of Ophthalmology, Case Western Reserve University, Cleveland, OH, USA.
Biochim Biophys Acta. 1995 Nov 15;1253(1):75-84. doi: 10.1016/0167-4838(95)00161-m.
Ascorbate (vitamin C) degradation products can undergo non-enzymatic glycation (Maillard reaction) with proteins to form highly crosslinked structures with brown pigmentation and characteristic fluorescence. Proteins in the body, especially the long-lived proteins develop similar changes during aging and diabetes. Several studies have shown excessive degradation of ascorbate in plasma in diabetes, and in ocular lens during aging and cataract formation. Recent studies have suggested that ascorbate degradation products-mediated glycation plays a role in lens pigmentation and cataract formation. However, the precise chemical nature of ascorbate-specific advanced glycation end-products are not known. Here, we report the purification and characterization of a glycation end-product derived from one of the major degradation products of ascorbate, L-threose. This compound was characterized to be 2-acetamido-6-(3-(1,2-dihydroxyethyl)-2-formyl-4-hydroxymethyl-1- pyrrolyl)hexanoic acid (formyl threosyl pyrrole or FTP) formed by the condensation of epsilon-amino group of lysine with two molecules of threose. Formation of FTP occurred rapidly in the incubation of threose and lysine and reached plateau level within a day. We have developed a sensitive assay for its quantification in proteins based on enzyme digestion followed by HPLC. Ribonuclease A and human lens crystallins incubated with L-threose showed time- and sugar concentration-dependent increases in FTP, reaching 8.2 and 2.48 nmol per mg protein, respectively after one week of incubation. Human plasma proteins showed a peak with identical retention time as that of purified FTP under two different HPLC conditions. FTP may be used as a sensitive marker to assess ascorbate-mediated protein glycation and modifications in aging and diabetes.
抗坏血酸（维生素C）降解产物可与蛋白质发生非酶糖基化反应（美拉德反应），形成具有棕色色素沉着和特征性荧光的高度交联结构。体内的蛋白质，尤其是长寿蛋白，在衰老和糖尿病过程中会发生类似变化。多项研究表明，糖尿病患者血浆中的抗坏血酸过度降解，衰老和白内障形成过程中眼晶状体中的抗坏血酸也过度降解。最近的研究表明，抗坏血酸降解产物介导的糖基化在晶状体色素沉着和白内障形成中起作用。然而，抗坏血酸特异性晚期糖基化终产物的确切化学性质尚不清楚。在此，我们报告了一种源自抗坏血酸主要降解产物之一L-苏糖的糖基化终产物的纯化和表征。该化合物经鉴定为2-乙酰氨基-6-（3-（1,2-二羟乙基）-2-甲酰基-4-羟甲基-1-吡咯基）己酸（甲酰苏糖基吡咯或FTP），它是由赖氨酸的ε-氨基与两分子苏糖缩合而成。在苏糖和赖氨酸的孵育过程中，FTP迅速形成，并在一天内达到稳定水平。我们开发了一种基于酶消化后进行高效液相色谱（HPLC）的灵敏检测方法，用于定量蛋白质中的FTP。与L-苏糖一起孵育的核糖核酸酶A和人晶状体晶状体蛋白显示，FTP随时间和糖浓度增加，孵育一周后，每毫克蛋白质分别达到8.2和2.48纳摩尔。在两种不同的HPLC条件下，人血浆蛋白显示出与纯化的FTP相同保留时间的峰。FTP可作为一个灵敏的标志物，用于评估抗坏血酸介导的蛋白质糖基化以及衰老和糖尿病中的修饰情况。
