N-糖基化对乙型脑炎病毒感染与增殖的影响.docVIP

  N-糖基化对乙型脑炎病毒感染与增殖的 影响# 张子重，宰俊杰，宋云峰，陈焕春** 5 10 15 （华中农业大学动物医学院，武汉 430070） 摘要：N-糖基化对于病毒的感染与增殖中均具有重要作用，切除病毒表面糖链的方法已成为 登革热病毒等的潜在治疗方法。为阐明日本乙型脑炎病毒（Japanese encephalitis virus， JEV）表面的糖链在其感染和增殖过程中的作用，本研究用肽 N-糖苷酶 F（Peptide N-Glycosidase F, PNGase F）和内切糖苷酶 H（Endoglycosidase H, Endo H）处理乙型脑 炎病毒，将酶切后的病毒用 Western blot 验证糖基化酶切的效果，同时将酶切的 JEV 接种 BHK-21 细胞进行蚀斑试验，观察蚀斑的大小及病毒滴度的变化情况。同时用衣霉素处理 BHK-21 细胞并同步接种 JEV，进行蚀斑试验，观察蚀斑大小的变化与病毒增殖情况。结果显 示，用 Endo H 和 PNGase F 处理 JEV 后，病毒形成的蚀斑均显著小于未处理的病毒对照，且 病毒增殖滴度显著降低，说明病毒粒子表面的糖基化修饰对于病毒的侵染具有重要作用。衣 霉素处理细胞后，病毒产生的蚀斑大小没有明显差异，但增殖滴度却显著降低，说明糖基化 在 JEV 增殖过程中也具有重要的功能。本研究说明 JEV 的糖基化对于其感染和增殖均具有重 要的作用，消除或抑制病毒的糖基化可作为抗病毒治疗的一种潜在的手段。 关键词：日本乙型脑炎病毒；糖苷酶；N-糖基化；感染；增殖 中图分类号：S855.3 20 Effects of N-Linked Glycosylation on the Infection and Proliferation of Japanese encephalitis virus (JEV) ZHANG Zizhong, ZAI Junjie, SONG Yunfeng, CHEN Huanchun (College of Veterinary Medicine, Huazhong Agricultural University, WuHan 430070) 25 30 35 Abstract: N-linked glycosylation site is critical for the infection and proliferation of most virus, elimination or inhibition of glycosylation is becoming a potent therapy of some virus such as dengue virus. In this study, endo-glycosidase enzymes Peptide N-Glycosidase F (PNGase F) and Endoglycosidase H (Endo H) were used to digest JEV to remove the the N-glycosylation on the virus particles surface. Western blot analysis showed that the glycan chains of JEV particles was successfully removed. To observe the changes in JEV infectivity and proliferation, plaque assay was performed using treated JEV on BHK-21 cells. In another exprement, BHK-21 cells were treated with tunicamycin and infected with JEV. The results showed that JEV digested by PNGase F or Endo H formed smaller plaques than control group, while JEV titer was reduced significantly. The plaques formed by JEV in cells treated with tunicamycin were not observed significant differences in size, but JEV titer was also reduced significantly. From these results we conformed that N-g