Huang L L, Shang F, Nowell T R, Taylor A
Laboratory for Nutrition and Vision Research, USDA Human Nutrition Research Center on Aging at Tufts University, Boston, MA 02111, USA.
Exp Eye Res. 1995 Jul;61(1):45-54. doi: 10.1016/s0014-4835(95)80057-3.
There is a growing consensus that altered proteins are more susceptible to degradation than native proteins. The enhancement of degradation of damaged proteins may be of significance since it prevents the accumulation of damaged proteins in cells. Several proteolytic pathways have been discovered in the lens. These include ATP-independent, ATP-dependent and ATP/ubiquitin-dependent proteolytic pathways. However, the extent of involvement of these proteolytic pathways in degradation of damaged proteins is not well described. alpha-Crystallin was oxidized by exposure to 0.03-3.2 mol.OH (mol protein)-1. Modifications to the oxidized alpha-crystallin and proteolytic susceptibility of the oxidized alpha-crystallin were studied. Exposure to > 0.32 mol.OH per mole of subunit produced aggregates and fragments of alpha-crystallin. Changes in isoelectric points of the proteins were observed after exposure to 0.64 mol.OH (mol protein)-1. The extent of loss of tryptophan and sulfhydryl groups was related to the level of .OH-exposure. Carbonyl content increased progressively with increasing oxidation. When incubated with a supernatant of bovine lens epithelial cells, the .OH-modified proteins were proteolytically degraded up to three times faster than untreated alpha-crystallin. ATP stimulated the degradation of native alpha-crystallin and alpha-crystallin which was exposed to 1.6 mol.OH (mol subunit protein)-1 (alpha 1.6). Sixty-seven per cent and 100% of the ATP-dependent degradation of native alpha-crystallin and alpha 1.6 was ubiquitin-dependent, respectively. The data indicate that alpha-crystallins oxidized by .OH are recognized and degraded rapidly by cytoplasmic proteolytic systems in bovine lens epithelial cells. Both ATP-independent and ATP/ubiquitin-dependent proteolytic pathways are involved in the degradation of native and oxidized alpha-crystallin.
人们越来越普遍地认为，与天然蛋白质相比，发生改变的蛋白质更容易被降解。受损蛋白质降解的增强可能具有重要意义，因为它可防止受损蛋白质在细胞内积累。在晶状体中已发现了几种蛋白水解途径。这些途径包括不依赖ATP的、依赖ATP的以及依赖ATP/泛素的蛋白水解途径。然而，这些蛋白水解途径在受损蛋白质降解中的参与程度尚未得到充分描述。α-晶状体蛋白通过暴露于0.03 - 3.2 mol·OH（每摩尔蛋白质）-1而被氧化。研究了被氧化的α-晶状体蛋白的修饰情况及其蛋白水解敏感性。每摩尔亚基暴露于> 0.32 mol·OH会产生α-晶状体蛋白的聚集体和片段。在暴露于0.64 mol·OH（每摩尔蛋白质）-1后，观察到蛋白质等电点的变化。色氨酸和巯基的损失程度与·OH暴露水平相关。羰基含量随着氧化程度的增加而逐渐增加。当与牛晶状体上皮细胞的上清液一起孵育时，经·OH修饰的蛋白质的蛋白水解降解速度比未处理的α-晶状体蛋白快达三倍。ATP刺激天然α-晶状体蛋白和暴露于1.6 mol·OH（每摩尔亚基蛋白质）-1的α-晶状体蛋白（α1.6）的降解。天然α-晶状体蛋白和α1.6的依赖ATP的降解分别有67%和100%是依赖泛素的。数据表明，被·OH氧化的α-晶状体蛋白在牛晶状体上皮细胞中被细胞质蛋白水解系统识别并迅速降解。不依赖ATP的和依赖ATP/泛素的蛋白水解途径都参与了天然和氧化的α-晶状体蛋白的降解。
