Yokomori K, La Monica N, Makino S, Shieh C K, Lai M M
Department of Microbiology, University of Southern California, School of Medicine, Los Angeles 90033.
Virology. 1989 Dec;173(2):683-91. doi: 10.1016/0042-6822(89)90581-3.
We have previously shown that gp65 (E3) is a virion structural protein which varies widely in quantity among different strains of mouse hepatitis virus (MHV). In this study, the biosynthetic pathway and possible biological activities of this protein were examined. The glycosylation of gp65 in virus-infected cells was inhibited by tunicamycin but not by monensin, suggesting that it contains an N-glycosidic linkage. Glycosylation is cotranslational and appears to be complete before the glycoprotein reaches the Golgi complex. Pulse-chase experiments showed that this protein decreased in size after 30 min of chase, suggesting that the carbohydrate chains of gp65 undergo trimming during its transport across the Golgi. This interpretation is supported by the endoglycosidase treatment of gp65, which showed that the peptide backbone of gp65 did not decrease in size after pulse-chase periods. This maturation pathway is distinct from that of the E1 or E2 glycoproteins. Partial endoglycosidase treatment indicated that gp65 contains 9 to 10 carbohydrate side chains; thus, almost all of the potential glycosylation sites of gp65 were glycosylated. In vitro translation studies coupled with protease digestion suggest that gp65 is an integral membrane protein. The presence of gp65 in the virion is correlated with the presence of an acetylesterase activity. No hemagglutinin activity was detected.
我们之前已经表明，gp65（E3）是一种病毒体结构蛋白，在不同株系的小鼠肝炎病毒（MHV）中其数量差异很大。在本研究中，我们检测了该蛋白的生物合成途径及可能的生物学活性。衣霉素可抑制病毒感染细胞中gp65的糖基化，但莫能菌素无此作用，这表明它含有N - 糖苷键。糖基化是共翻译过程，且似乎在糖蛋白到达高尔基体复合体之前就已完成。脉冲追踪实验表明，追踪30分钟后该蛋白大小减小，这表明gp65的糖链在其穿过高尔基体的转运过程中会发生修剪。对gp65进行内切糖苷酶处理支持了这一解释，结果显示脉冲追踪后gp65的肽主链大小并未减小。这种成熟途径与E1或E2糖蛋白的不同。部分内切糖苷酶处理表明gp65含有9至10条糖侧链；因此，gp65几乎所有潜在的糖基化位点都已被糖基化。体外翻译研究结合蛋白酶消化表明gp65是一种整合膜蛋白。病毒体中gp65的存在与乙酰酯酶活性的存在相关。未检测到血凝素活性。
