Tweto J, Doyle D
J Biol Chem. 1976 Feb 10;251(3):872-82.
The turnover of the plasma membrane proteins of hepatoma tissue culture cells was examined by three different methods--loss of polypeptides labeled in situ by lactoperoxidase-catalyzed iodination, loss of membrane polypeptides labeled with amino acid precursors, and loss from the membrane of fucose-labeled polypeptides. In both logarithmically growing and density-inhibited cells the proteins of the membrane are degraded with a half-life of about 100 hours. This is longer than the half-life of total cell protein, 50 to 60 hours, and longer than the doubling time of the cells, about 30 hours. Similar values for the rate of degradation of the membrane proteins were obtained by each of the three techniques. The same fucose-labeled polypeptides are present in the microsomal and the plasma membrane fractions of hepatoma tissue culture cells as analyzed by electrophoresis in dodecyl sulfate-acrylamide gels. But the fucose-labeled polypeptides were lost from the microsomal fraction at a faster rate than from the plasma membrane. Autoradiographic and double labeling techniques using 125I and 131I, or [3H]leucine and [14C]leucine were used to measure the relative rates of degradation of the proteins in the plasma membrane. All of the leucine-labeled polypeptides and the iodinated polypeptides had similar rates of degradation. These results support a model for the biogenesis of the plasma membrane in which the proteins are incorporated and removed in large structural units.
通过三种不同方法检测了肝癌组织培养细胞质膜蛋白的周转情况——通过乳过氧化物酶催化碘化原位标记的多肽的损失、用氨基酸前体标记的膜多肽的损失以及岩藻糖标记的多肽从膜上的损失。在对数生长期和密度抑制期的细胞中，膜蛋白均以约100小时的半衰期被降解。这比总细胞蛋白的半衰期（50至60小时）长，也比细胞的倍增时间（约30小时）长。通过这三种技术中的每一种都获得了膜蛋白降解速率的相似值。如在十二烷基硫酸钠-丙烯酰胺凝胶中进行电泳分析所示，肝癌组织培养细胞的微粒体和质膜部分中存在相同的岩藻糖标记的多肽。但是岩藻糖标记的多肽从微粒体部分的损失速率比从质膜的损失速率更快。使用125I和131I或[3H]亮氨酸和[14C]亮氨酸的放射自显影和双重标记技术用于测量质膜中蛋白质的相对降解速率。所有亮氨酸标记的多肽和碘化多肽都有相似的降解速率。这些结果支持了一种质膜生物发生模型，即蛋白质在大的结构单元中被整合和去除。
