年龄相关性白内障α-晶状体蛋白的分子伴侣功能

目的:研究年龄相关性白内障α-晶状体蛋白的分子伴侣功能。方法:使用SephacrylS-300HR分离健康人透明的和年龄相关性白内障混浊的晶状体皮质和核的αL-晶状体蛋白。采用分光光度计测定αL-晶状体蛋白对过氧化氢酶(catalase,CAT)热凝聚的抑制作用。应用SDS-PAGE分析CAT与αL-晶状体蛋白在热凝聚实验中形成的复合物。结果:人αL-晶状体蛋白可特异性地抑制CAT热凝聚。透明晶状体皮质较核的抑制作用明显;白内障αL-晶状体蛋白的抑制作用明显下降,65岁以下与65岁以上、65岁以上白内障Ⅱ与Ⅳ级核之间αL-晶状体蛋白的抑制作用比较,差异均有显著性;65岁以下Ⅱ与Ⅳ级核之间比较,差异无显著性,但Ⅳ级核的抑制作用降低。SDS-PAGE显示热凝聚实验透明晶状体皮质αL-晶状体蛋白的20kDa带在加热后0、40min和2h可溶部分无显著减少,62kDa的CAT带含量逐渐减少。结论:人α-晶状体蛋白具有抑制CAT热凝聚的分子伴侣功能,白内障晶状体核的α-晶体蛋白具有年龄和混浊程度依赖性伴侣功能的降低,这在白内障形成过程中起重要作用。

The Molecular Chaperone Function of α-crystallin in Age-related Cataract

PURPOSE: To evaluate the molecular chaperone function of alpha-crystallin in age-related cataract. METHODS: alpha L-Crystallin of human clear and cataractous lenses in both the cortex and nucleus were separated by chromatography on Sephacryl S-300HR. The preventive effects of alpha L-crystallin on thermally induced aggregation of catalase were measured spectrophotometrically. The complex formation between a catalase and alpha L-crystallin during the aggregation experimental was detected by SDS-PAGE methods. RESULTS: Human alpha L-crystallin specifically prevented catalase from thermal aggregation. Its preventive ability in cortex was greatly higher than that in nucleus of clear lenses. There was a remarkably decrease of preventive ability in cataractous lenses with statistical differences between the group under 65 years old and that over 65 years old, grade II and grade IV in over 65 years group, whereas alpha L-crystallin showed no statistical change in under 65 years group but its preventive ability was diminished in grade IV. SDS-PAGE showed that the 20 kDa bands had no significant decrease in water-soluble portion after heating 0.40 mim and 2 h, whereas the 65 kD bands were increased. CONCLUSION: alpha-Crystallin can function as a molecular chaperone by preventing thermal aggregation of catalase. A decrease in chaperone function from cataractous lenses has been showed in age-dependent and opacification-dependent fashion, which may play an important role in formation of cataract.