Schwartz A L, Trausch J S, Ciechanover A, Slot J W, Geuze H
Edward Mallinckrodt Department of Pediatrics, Washington University School of Medicine, St. Louis, MO.
Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5542-6. doi: 10.1073/pnas.89.12.5542.
As the first enzyme in the ubiquitin system the ubiquitin-activating enzyme E1 plays a pivotal role in all pathways of protein ubiquitination. In an effort to learn more about the cell biology of this pathway, we have purified the 110-kDa enzyme to homogeneity and generated a panel of distinct monoclonal antibodies to it. Using quantitative electron microscopic immunolocalization with these anti-E1 monoclonal antibodies, we find that E1 is abundant both within the cytoplasm and nucleus. Within the cytoplasm, E1 was found throughout the cytoplasmic volume as well as enriched along the cytoplasmic face of the rough endoplasmic reticulum and associated with the dense material along the desmosomal junctions. E1 was also found associated with the cytoplasmic surface of endosomal/lysosomal vacuoles. Interestingly, E1 was also found within the mitochondria. The lumen of rough endoplasmic reticulum, Golgi complex, endosomes, and lysosomes were negative. The specific localization of E1 to distinct subcellular organelles suggests that E1 may play multiple physiological roles within the cell.
作为泛素系统中的首个酶，泛素激活酶E1在蛋白质泛素化的所有途径中都起着关键作用。为了更深入了解该途径的细胞生物学，我们已将110 kDa的酶纯化至同质，并制备了一组针对它的不同单克隆抗体。使用这些抗E1单克隆抗体进行定量电子显微镜免疫定位，我们发现E1在细胞质和细胞核中均大量存在。在细胞质内，E1存在于整个细胞质体积中，并且在粗面内质网的细胞质面富集，并与桥粒连接处的致密物质相关联。还发现E1与内体/溶酶体空泡的细胞质表面相关。有趣的是，在线粒体中也发现了E1。粗面内质网、高尔基体、内体和溶酶体的腔均为阴性。E1在不同亚细胞细胞器中的特异性定位表明E1可能在细胞内发挥多种生理作用。
