A NEW METHOD FOR DETECTING ENDOCYTOSED PROTEINS
A NEW METHOD FOR DETECTING ENDOCYTOSED PROTEINS
复制标题
DOI：
10.1002/j.1460-2075.1988.tb03302.x
复制
发表时间：
1988-12-20
期刊：
影响因子：
11.4
通讯作者：
LUTTER, R
中科院分区：
文献类型：
--
作者：
BRETSCHER, MS;LUTTER, R
描述了一种新的试剂DPSgt，它被设计用于在0℃下标记细胞表面蛋白。该试剂易于制备；它是可溶的，在一端含有活性的不均匀酯，在另一端含有可放射性的酪氨酸，在两者之间含有二硫化物。用谷胱甘肽在0℃下切断细胞中的二硫键，可将标签从细胞中去除。当细胞在标记和还原之间被加热到37℃时，被内吞的标记蛋白获得了对还原的抗性。这提供了一种测量表面蛋白内吞作用的简单方法。已经估计了K562细胞和成纤维细胞中转铁蛋白和LDL受体的胞内池。结果表明，细胞内受体位于非还原区室中，K562细胞对平均细胞表面（通过非包被坑过程）的摄取很小。
A new reagent, DPSgt, is described which has been designed to label cell surface proteins at 0.degree.C. The reagent is easily made; it is soluble and contains a reactive impermeant ester at one end, a tyrosine which can be radioiodinated at the other, and a disulphide in-between. The label can be removed from cells by cleaving the disulphide linkage in it with glutathione at 0.degree.C. When cells are warmed to 37.degree.C between labelling and reduction, labelled proteins which are endocytosed acquired resistance to reduction. This provides a simple way of measuring the endocytosis of surface proteins. The intracellular pools of transferrin and LDL receptors in K562 cells and fibroblasts have been estimated. The results indicate that intracellular receptors are in non-reducing compartments, and that uptake of average cell surface (by non-coated pit processes) in K562 cells is small.