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猪瘟病毒石门强毒株和兔化弱毒疫苗株E2蛋白糖基化位点差异分析
引用本文:彭伍平,夏照和,侯强,李娜,孙元,童光志,仇华吉.猪瘟病毒石门强毒株和兔化弱毒疫苗株E2蛋白糖基化位点差异分析[J].病毒学报,2007,23(5):389-393.
作者姓名:彭伍平  夏照和  侯强  李娜  孙元  童光志  仇华吉
作者单位:中国农业科学院,哈尔滨兽医研究所,兽医生物技术国家重点实验室,猪传染病研究室,哈尔滨,150001
基金项目:国家973计划资助(2005CB523202)
摘    要:猪瘟病毒强毒株和兔化弱毒疫苗株E2糖蛋白分别含有5个和6个潜在的糖基化位点,其中986N是兔化弱毒疫苗株所特有的。为了分析二者糖基化位点差异及其影响,将去掉信号肽和跨膜区的猪瘟病毒石门强毒株(Shi-men)和兔化弱毒疫苗株(HCLV)E2基因置于蜂素信号肽序列下游,使其在Sf9细胞内表达重组Shimen-E2和HCLV-E2蛋白。结果显示,重组E2蛋白以二聚体的形式分泌表达于细胞培养液中,但二者分子量存在差异。用endo H和PNGase F对纯化后的重组E2蛋白进行去糖基化处理后,二者分子量大小变成一致,证实石门强毒株和兔化弱毒株E2蛋白分子量大小的差异可能是由于糖基化程度的差异所致。对986N糖基化位点进行定点突变后发现,突变后的Shimen-E2与野生型HCLV-E2分子量大小一致,而突变后的HCLV-E2与野生型Shimen-E2分子量大小一致,表明Shimen-E2和HCLV-E2分子量大小的差异的确是由于986N糖基化位点的差异引起的。

关 键 词:猪瘟病毒  E2囊膜糖蛋白  糖基化位点  杆状病毒表达系统  蜂素信号肽
文章编号:1000-8721(2007)05-0389-05
修稿时间:2007-03-27

Differences in Glycosylation of the E2 Protein between Virulent Shimen Strain and Avirulent C-Strain of Classical Swine Fever Virus
PENG Wu-ping,XIA Zhao-he,HOU Qiang,LI Na,SUN Yuan,TONG Guang-zhi,QIU Hua-ji.Differences in Glycosylation of the E2 Protein between Virulent Shimen Strain and Avirulent C-Strain of Classical Swine Fever Virus[J].Chinese Journal of Virology,2007,23(5):389-393.
Authors:PENG Wu-ping  XIA Zhao-he  HOU Qiang  LI Na  SUN Yuan  TONG Guang-zhi  QIU Hua-ji
Institution:Division of Swine Infectious Diseases, National Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Harbin 150001 ,China
Abstract:The E2 envelope glycoprotein of virulent Shimen strain and avirulent C-strain of Classical swine fever virus(CSFV) has 5 and 6 potential glycosylation sites,respectively,and the potential glycosylation site986N is unique to Cstrain.To study the differences in glycosylation between the virus pair,the E2 genes(removing signal sequence and transmembrane anchor regions) of the two strains fused with the melittin signal sequence were expressed in the Sf9 insect cells.The recombinant E2 proteins were secreted into the medium of Sf9 cells in dimer form with different molecular weight(MW).Deglycosylation of the recombinant E2 proteins by endo H and PNGase F showed that the deglycosalated Shimen-E2 and HCLV-E2 have the same MW,indicating that the different MW between ShimenE2 and HCLV-E2 proteins came from different glycosylation.Site-directed mutagenesis in the potential glycosylation site at 986N demonstrated that the mutated Shimen-E2 protein had the same MW as the wild-type HCLVE2 protein,while the mutated HCLV-E2 had the same MW as the wild-type Shimen-E2 protein.We suggest that the different MW between Shimen-E2 and HCLV-E2 is resulted from the different glycosylation on 986N glycosylation site.
Keywords:Classical swine fever virus  envelope glycoprotein E2  glycosylation site  bacuclovirus expression system  melittin signal
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