Chandrasekher G, Cenedella R J
Department of Biochemistry, Kirksville College of Osteopathic Medicine, MO 63501, USA.
Exp Eye Res. 1995 Jun;60(6):707-17. doi: 10.1016/s0014-4835(05)80012-0.
Plasma membrane contains extrinsic as well as intrinsic proteins. Changes in the extrinsic proteins of lens membrane during human aging and cataract formation have not been investigated in detail. Unlike previous studies which examined lens membrane after being stripped of extrinsic proteins by treatment with chaotropic agents, we have isolated whole or 'native' lens membrane on a sucrose gradient by ultracentrifugation of the total water-insoluble protein. Essentially all of the water-insoluble protein from young to aged to cataractous human lens appeared membrane associated. In young lens (20-37 years old), most of the membrane banded at the 25/45% sucrose interface fraction. This fraction contained relatively little urea-soluble protein and likely represents fiber-cell plasma membrane with its physiologically associated extrinsic and intrinsic proteins. With aging (62-80 years old), about one-third of the membrane, as judged by the distribution of cholesterol, banded at a much higher density (50/58% sucrose fraction). The higher density was due to a great increase in the membrane's relative protein content (protein/cholesterol). Although this extra protein was composed of both urea-insoluble and -soluble fractions, the urea-soluble protein predominated in all lenses. Cataractous lens differed from aged-clear lens in that much more of the total membrane (70-75%) had shifted to the high density and participated in this massive binding of cytosolic proteins. Although alpha-crystallin was the principal extrinsic-membrane protein in young lens, high molecular weight aggregate of modified (acidic) crystallins accounted for the increased extrinsic protein in aging. The extrinsic proteins bound to both clear-aged and cataractous lens membrane were aggregated. In conclusion, examination of human lens native membrane fractions revealed that the association of crystallins with membrane in both aging and cataracts was much greater than previously recognized and most of this increased protein was non-covalently bound to the membrane. Much more of the lens total membrane from cataractous than clear-aged lens was involved in this massive protein association and the protein bound to cataract membrane appeared more highly aggregated.
质膜包含外在蛋白和内在蛋白。人类衰老和白内障形成过程中晶状体膜外在蛋白的变化尚未得到详细研究。与以往通过用离液剂处理去除外在蛋白后检查晶状体膜的研究不同，我们通过对总水不溶性蛋白进行超速离心，在蔗糖梯度上分离出完整的或“天然的”晶状体膜。从年轻到年老再到白内障患者的人晶状体中，基本上所有的水不溶性蛋白都与膜相关。在年轻晶状体（20 - 37岁）中，大部分膜在25/45%蔗糖界面部分形成条带。该部分含有相对较少的尿素可溶性蛋白，可能代表纤维细胞质膜及其生理相关的外在和内在蛋白。随着年龄增长（62 - 80岁），根据胆固醇分布判断，约三分之一的膜在更高密度处形成条带（50/58%蔗糖部分）。更高的密度是由于膜的相对蛋白含量（蛋白/胆固醇）大幅增加。尽管这种额外的蛋白由尿素不溶性和可溶性部分组成，但在所有晶状体中尿素可溶性蛋白占主导。白内障晶状体与年老透明晶状体的不同之处在于，总膜的更多部分（70 - 75%）已转移到高密度区域，并参与了这种大量的细胞溶质蛋白结合。虽然α-晶状体蛋白是年轻晶状体中的主要外在膜蛋白，但修饰（酸性）晶状体蛋白的高分子量聚集体导致了衰老过程中外在蛋白的增加。与年老透明和白内障晶状体膜结合的外在蛋白都发生了聚集。总之，对人晶状体天然膜部分的检查表明，衰老和白内障过程中晶状体蛋白与膜的结合比以前认识到的要多得多，并且这种增加的蛋白大部分与膜非共价结合。与年老透明晶状体相比，白内障晶状体的总膜更多地参与了这种大量的蛋白结合，并且与白内障膜结合的蛋白似乎聚集程度更高。
