Bourdon E, Loreau N, Blache D
INSERM U498, Biochimie des Lipoprotéines et Interactions Vasculaires, Université de Bourgogne, Dijon, France.
FASEB J. 1999 Feb;13(2):233-44. doi: 10.1096/fasebj.13.2.233.
Epidemiological data consistently show that reduced levels of serum albumin, which is the most abundant protein in plasma, are associated with an increased mortality risk. Various biological properties evidenced by direct effects of the albumin molecule may explain its beneficial effects. The present work aimed to investigate in vitro whether glycation or free radicals or both factors would affect the antioxidant properties of bovine serum albumin (BSA). Glycation was performed by long-term incubations (60 days) of BSA with increasing concentrations of glucose (up to 500 mmol/l) at 37 degreesC. Minimally oxidized BSA was obtained after controlled incubations of dialyzed BSA samples with a water-soluble free radical generator [2,2' azo-bis(2-amidinopropane) HCl]. The glycation-mediated modifications and the free radical-induced conformational changes of BSA were monitored using intrinsic fluorescence measurements of the tryptophan residues and acrylamide as a quenching agent. Thiol groups, Amadori glycophore contents, and boronate binding were also measured. We found that the changes observed in the conformation of the BSA molecule were associated with modifications of its antioxidant properties. The latter were studied by the copper-mediated oxidation of human low density lipoproteins and the free radical-induced blood hemolysis test. Our data support the concept that oxidative-induced BSA modifications are important determinants in the antioxidant properties of BSA. Glycated BSA still behaved as an antioxidant but became pro-oxidant in the presence of copper, probably by generating oxygenated species. These data confirm the key role of metals ions in this process. Although these results warrant further in vivo investigations, we propose that, considering the poor glucose control found in diabetics as well as the key role of oxidative stress in vascular complications, glycation-mediated and free radical-induced impairment of the antioxidant properties of albumin might be important parameters in vascular complications encountered in diabetes.
流行病学数据始终表明，血清白蛋白水平降低与死亡风险增加相关，血清白蛋白是血浆中含量最丰富的蛋白质。白蛋白分子的直接作用所展现出的各种生物学特性或许可以解释其有益作用。本研究旨在体外探究糖基化、自由基或这两种因素是否会影响牛血清白蛋白（BSA）的抗氧化特性。通过在37℃下将BSA与浓度不断增加的葡萄糖（高达500 mmol/l）长期孵育（60天）来进行糖基化。在用一种水溶性自由基发生器[2,2'-偶氮二(2-脒基丙烷)盐酸盐]对透析后的BSA样品进行控制性孵育后，获得轻度氧化的BSA。使用色氨酸残基的固有荧光测量以及丙烯酰胺作为猝灭剂来监测BSA的糖基化介导修饰和自由基诱导的构象变化。还测量了巯基、阿马多里糖基含量和硼酸酯结合情况。我们发现，BSA分子构象的变化与其抗氧化特性的改变相关。后者通过铜介导的人低密度脂蛋白氧化和自由基诱导的血液溶血试验进行研究。我们的数据支持这样一种观点，即氧化诱导的BSA修饰是BSA抗氧化特性的重要决定因素。糖基化的BSA仍表现为抗氧化剂，但在有铜存在的情况下可能通过产生活氧物种而变成促氧化剂。这些数据证实了金属离子在这一过程中的关键作用。尽管这些结果有待进一步的体内研究，但我们提出，考虑到糖尿病患者血糖控制不佳以及氧化应激在血管并发症中的关键作用，糖基化介导和自由基诱导的白蛋白抗氧化特性损害可能是糖尿病患者血管并发症的重要参数。
