Kohring G W, Mayer F, Mayer H
Eur J Cell Biol. 1985 May;37:1-6.
Purified restriction endonuclease EcoRI isolated from Escherichia coli BS 5 was used for the production of enzyme-specific IgG antibodies in rabbits. For enzyme localization experiments, paraformaldehyde-glutaraldehyde-fixed cells were embedded and polymerized by a low-temperature procedure using Lowicryl K4M. The immuno electron microscopic protein A-gold technique and an immuno-gold method revealed that 70% of the enzyme-specific labeling were located in the cell envelope whereas 30% were found in the cytoplasm. In metal-shadowed preparations, no indications for the presence of the enzyme could be found on the cell surface; however, on the surface of cell protoplasts enzyme specific labeling could be detected. The results indicate the presence of the major amount of EcoRI in the periplasmic space of the cell where it might be loosely bound or even freely diffusible.
从大肠杆菌BS 5中分离出的纯化限制性内切酶EcoRI用于在兔子体内产生酶特异性IgG抗体。对于酶定位实验，使用低温程序用Lowicryl K4M对多聚甲醛-戊二醛固定的细胞进行包埋和聚合。免疫电子显微镜蛋白A-金技术和免疫金方法显示，70%的酶特异性标记位于细胞膜，而30%位于细胞质中。在金属阴影制备物中，在细胞表面未发现该酶存在的迹象；然而，在细胞原生质体表面可检测到酶特异性标记。结果表明，细胞周质空间中存在大量的EcoRI，它可能松散结合甚至自由扩散。
