Specific covalent labeling of recombinant protein molecules inside live cells
Specific covalent labeling of recombinant protein molecules inside live cells
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DOI：
10.1126/science.281.5374.269
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发表时间：
1998-07-10
期刊：
影响因子：
56.9
通讯作者：
Tsien, RY
中科院分区：
文献类型：
--
作者：
Griffin, BA;Adams, SR;Tsien, RY
在α螺旋的i、i + 1、i + 4和i + 5位含有四个半胱氨酸的重组蛋白质通过细胞外施用4 ′，5 ′-双（1，3，2-二硫阿索兰-2-基）荧光这种设计的小配体是膜渗透的和非荧光的，直到它以高亲和力和特异性结合到四半胱氨酸结构域，这种原位标记比绿色荧光蛋白增加的质量少得多，并在附着位点以及潜在的光谱和化学性质方面提供更大的通用性。该系统提供了一种对靶蛋白进行轻微修饰的方法，使其可以从活细胞内的许多其他蛋白质中挑选出来，并通过从细胞外添加的小的非荧光染料分子进行荧光染色。
Recombinant proteins containing four cysteines at the i, i + 1, i + 4, and i + 5 positions of an alpha helix were fluorescently labeled in Living cells by extracellular administration of 4',5'-bis(1,3,2-dithioarsolan-2-yl)fluor This designed small Ligand is membrane-permeant and nonfluorescent until it binds with high affinity and specificity to the tetracysteine domain, Such in situ Labeling adds much Less mass than does green fluorescent protein and offers greater versatility in attachment sites as well as potential spectroscopic and chemical properties. This system provides a recipe for slightly modifying a target protein so that it can be singled out from the many other proteins inside Live cells and fluorescently stained by small nonfluorescent dye molecules added from outside the cells.