Stefek M, Kyselova Z, Rackova L, Krizanova L
Institute of Experimental Pharmacology, Slovak Academy of Sciences, Dubravska cesta 9, 841 04 Bratislava, Slovakia.
Biochim Biophys Acta. 2005 Jun 30;1741(1-2):183-90. doi: 10.1016/j.bbadis.2005.01.006. Epub 2005 Feb 10.
In an attempt to model the processes of free radical-mediated cataractogenesis, we investigated the oxidative modification of rat eye lens proteins by peroxyl radicals generated by thermal decomposition of 2,2'-azobis(2-amidinopropane)hydrochloride (AAPH) under aerobic conditions. When incubated with AAPH, the soluble eye lens proteins precipitated in a time-dependent manner. The insolubilisation was accompanied by the accumulation of protein free carbonyls and the diminution of sulfhydryls, yet the processes were shifted in time. The SDS-PAGE analysis of the AAPH-treated proteins revealed the presence of high molecular weight cross-links and, to a lesser extent, fragments. The aggregation and cross-linking of proteins along with the generation of free carbonyls was significantly inhibited by the chain-breaking antioxidants stobadine and Trolox. On the other hand, the AAPH-initiated sulfhydryl consumption was much less sensitive to the antioxidants studied. The results point to a complex mechanism of peroxyl-radical-mediated modification of eye lens proteins with implications for cataract development and they indicate a potentially protective role of antioxidants.
为了模拟自由基介导的白内障形成过程，我们研究了在有氧条件下，2,2'-偶氮二(2-脒基丙烷)盐酸盐(AAPH)热分解产生的过氧自由基对大鼠眼晶状体蛋白的氧化修饰作用。当与AAPH一起孵育时，可溶性眼晶状体蛋白会随时间沉淀。不溶性化伴随着蛋白质游离羰基的积累和巯基的减少，但这些过程在时间上有所偏移。对经AAPH处理的蛋白质进行SDS-PAGE分析，发现存在高分子量交联物，程度较轻的还有片段。链断裂抗氧化剂司他丁和生育三烯酚显著抑制了蛋白质的聚集和交联以及游离羰基的生成。另一方面，AAPH引发的巯基消耗对所研究的抗氧化剂不太敏感。结果表明过氧自由基介导的眼晶状体蛋白修饰机制复杂，与白内障发展有关，并且表明抗氧化剂具有潜在的保护作用。
