Suh K, Gabel C A, Bergmann J E
Department of Anatomy and Cell Biology, Columbia University College of Physicians and Surgeons, New York, New York 10032.
J Biol Chem. 1992 Oct 25;267(30):21671-7.
The role of glucosylated oligosaccharides in the biogenesis of the glycoprotein (G protein) of vesicular stomatitis virus was studied in PhaR2.7, a mouse lymphoma cell line deficient in glucosidase II activity. As expected, the great majority of cell-associated G protein remained glucosylated in PhaR2.7, and the G protein was rapidly deglucosylated in BW5147, the parental cell line. Despite these differences in glucosylation, the rates of G protein trimerization and transport to the cell surface were as rapid and efficient in the PhaR2.7 mutant as in BW5147. Surprisingly, greater than 73% of the oligosaccharides on G proteins recovered from released virions were complex-type units. The efficient processing of the G protein oligosaccharides coincided with the efficient removal of glucose residues from its oligosaccharides. After treatment with deoxynojirimycin, an inhibitor of endoplasmic reticulum (ER) glucosidases I and II, the total percentage of G protein-associated high mannose-type oligosaccharides increased more in the parental cells than in the mutant cells. Furthermore, when the G protein was retained in the ER of PhaR2.7 cells by depletion of the cellular ATP pools with carbonyl cyanide m-chlorophenylhydrazone, its oligosaccharides remained glucosylated. Under identical conditions, BW5147 cells removed the glucose residues from > 90% of the retained G protein's oligosaccharides. Thus, PhaR2.7 cells efficiently remove glucose residues from high mannose-type oligosaccharides of selected proteins using a deoxynojirimycin-insensitive enzyme located in a post-ER compartment. The existence of a second mechanism for the deglucosylation of N-linked oligosaccharides provides evidence for the important role of glucose removal in glycoprotein maturation.
在缺乏葡糖苷酶II活性的小鼠淋巴瘤细胞系PhaR2.7中，研究了糖基化寡糖在水疱性口炎病毒糖蛋白（G蛋白）生物合成中的作用。正如预期的那样，在PhaR2.7中，绝大多数与细胞相关的G蛋白保持糖基化状态，而在亲代细胞系BW5147中，G蛋白迅速去糖基化。尽管在糖基化方面存在这些差异，但在PhaR2.7突变体中，G蛋白三聚化和转运到细胞表面的速率与在BW5147中一样迅速且高效。令人惊讶的是，从释放的病毒粒子中回收的G蛋白上超过73%的寡糖是复合型单位。G蛋白寡糖的有效加工与从其寡糖中有效去除葡萄糖残基相吻合。用脱氧野尻霉素（一种内质网葡糖苷酶I和II的抑制剂）处理后，亲代细胞中与G蛋白相关的高甘露糖型寡糖的总百分比增加幅度大于突变细胞。此外，当通过用羰基氰化物间氯苯腙耗尽细胞ATP池将G蛋白保留在PhaR2.7细胞的内质网中时，其寡糖仍保持糖基化。在相同条件下，BW5147细胞从超过90%保留的G蛋白寡糖中去除了葡萄糖残基。因此，PhaR2.7细胞利用位于内质网后区室的一种对脱氧野尻霉素不敏感的酶，有效地从选定蛋白质的高甘露糖型寡糖中去除葡萄糖残基。N-连接寡糖去糖基化的第二种机制的存在为葡萄糖去除在糖蛋白成熟中的重要作用提供了证据。
