兔出血症病毒衣壳蛋白P区二聚体的表达及其与受体结合能力分析

doi:10.11843/j.issn.0366-6964.2016.02.015

畜牧兽医学报 ›› 2016, Vol. 47 ›› Issue (2): 325-330.doi: 10.11843/j.issn.0366-6964.2016.02.015

兔出血症病毒衣壳蛋白P区二聚体的表达及其与受体结合能力分析

胡波，范志宇，王芳^*，魏后军，宋艳华，仇汝龙，徐为中，薛家宾

(江苏省农业科学院兽医研究所•农业部兽用生物制品工程技术重点实验室•国家兽用生物制品工程技术研究中心，南京 210014)

收稿日期:2015-06-20 出版日期:2016-02-23 发布日期:2016-02-23
通讯作者: 王芳，研究员，E-mail：rwangfang@126.com
作者简介:胡波（1982- ），男，江苏南京人，助理研究员，硕士，主要从事畜禽疫病防控与兽医生物技术研究，Tel：025-84390337，E-mail：hadisi2002@163.com
基金资助:
现代农业产业技术体系建设专项资金（CARS-44）；江苏省农业科技自主创新资金项目［CX (13) 5029］

Expression of the P Dimer of Rabbit Hemorrhagic Disease Virus Capsid Protein and Analysis of Its Binding Ability to Receptor

HU Bo，FAN Zhi-yu，WANG Fang^*，WEI Hou-jun，SONG Yan-hua，QIU Ru-long，XU Wei-zhong，XUE Jia-bin

(Institute of Veterinary Medicine，Jiangsu Academy of Agricultural Sciences/Key Laboratory of Veterinary Biological Engineering and Technology of Ministry of Agriculture /National Center for Engineering Research of Veterinary Bio-products，Nanjing 210014，China)

Received:2015-06-20 Online:2016-02-23 Published:2016-02-23

摘要/Abstract

摘要：

为检测兔出血症病毒（RHDV）衣壳蛋白（VP60）P区形成二聚体的能力及其与HBGAs受体结合的能力，作者以pMD19T-VP60为模板扩增包含铰链区H的P区基因（HP）并克隆至原核表达载体pET28a（+）中，转化大肠杆菌BL21（DE3）菌株，经IPTG诱导表达，获得HP蛋白。经SDS-PAGE和Western blot检测，结果显示，HP蛋白主要以包涵体的形式高效表达，经HisTrap亲和柱纯化后可形成二聚体结构。通过HBGAs受体结合试验表明，P区与完整病毒衣壳相似，能与唾液中的HBGAs受体发生结合。本研究为进一步开展RHDV衣壳蛋白与受体的相互作用研究奠定基础。

Abstract:

Rabbit hemorrhagic disease virus (RHDV) is a member of the Caliciviridae family (Lagovirus genus).Similar to human norovirus (Caliciviridae，Norovirus genus)，RHDV binds histo-blood group antigens (HBGAs) and this is thought to be important for infection.The aim of this study was to determine whether the P domain of RHDV capsid protein (VP60) could form dimers and to analyze its binding ability to HBGAs receptor.The P domain gene containing hinge (HP) was amplified and cloned into pET-28a (+) expression vector and introduced into Escherichia coli BL21 (DE3).The recombinant HP protein，confirmed by SDS-PAGE and Western blot，was effectively expressed in form of inclusion bodies by inducing with IPTG.The recombinant protein was then purified with HisTrap affinity chromatography，which could form dimers after purification.The HBGAs binding assay showed that the P domain bound H type HBGA with the same patterns as those of the intact viral capsids.Further structural studies with P domain are needed in order to better understand the HBGA binding mechanisms and virus-receptor interaction.

中图分类号:

胡波，范志宇，王芳，魏后军，宋艳华，仇汝龙，徐为中，薛家宾. 兔出血症病毒衣壳蛋白P区二聚体的表达及其与受体结合能力分析[J]. 畜牧兽医学报, 2016, 47(2): 325-330.

HU Bo，FAN Zhi-yu，WANG Fang，WEI Hou-jun，SONG Yan-hua，QIU Ru-long，XU Wei-zhong，XUE Jia-bin. Expression of the P Dimer of Rabbit Hemorrhagic Disease Virus Capsid Protein and Analysis of Its Binding Ability to Receptor[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2016, 47(2): 325-330.

0
/ / 推荐

导出引用管理器 EndNote|Reference Manager|ProCite|BibTeX|RefWorks

链接本文: https://www.xmsyxb.com/CN/10.11843/j.issn.0366-6964.2016.02.015

https://www.xmsyxb.com/CN/Y2016/V47/I2/325

参考文献

［1］MITRO S，KRAUSS H.Rabbit hemorrhagic disease：a review with special reference to its epizootiology［J］.Eur J Epidemiol，1993，9(1)：70-78.
［2］ABRANTES J，VAN DER LOO W，LE PENDU J，et al.Rabbit haemorrhagic disease (RHD) and rabbit haemorrhagic disease virus (RHDV)：a review［J］.Vet Res，2012，43：12.
［3］WANG X，XU F，LIU J，et al.Atomic model of rabbit hemorrhagic disease virus by cryo-electron microscopy and crystallography［J］.PLoS Pathog，2013，9(1)：e1003132.
［4］LAURENT S，VAUTHEROT J F，LE GALL G，et al.Structural，antigenic and immunogenic relationships between European brown hare syndrome virus and rabbit haemorrhagic disease virus［J］.J Gen Virol，1997，78（Pt 11)：2803-2811.
［5］ESTEVES P J，ABRANTES J，CARNEIRO M，et al.Detection of positive selection in the major capsid protein VP60 of the rabbit haemorrhagic disease virus (RHDV)［J］.Virus Res，2008，137(2)：253-256.
［6］KINNEAR M，LINDE C C.Capsid gene divergence in rabbit hemorrhagic disease virus［J］.J Gen Virol，2010，91(Pt 1)：174-181.
［7］ROHAYEM J，BERGMANN M，GEBHARDT J，et al.Antiviral strategies to control calicivirus infections［J］.Antiviral Res，2010，87(2)：162-178.
［8］NYSTRÖM K，LE GALL-RECULÉ G，GRASSI P，et al.Histo-blood group antigens act as attachment factors of rabbit hemorrhagic disease virus infection in a virus strain-dependent manner［J］.PLoS Pathog，2011，7(8)：e1002188.
［9］RUVOËN-CLOUET N，GANIÈRE J P，ANDRÉ-FONTAINE G，et al.Binding of rabbit hemorrhagic disease virus to antigens of the ABH histo-blood group family［J］.J Virol，2000，74(24)：11950-11954.
［10］HUANG P，FARKAS T，MARIONNEAU S，et al.Noroviruses bind to human ABO，Lewis，and secretor histo-blood group antigens：identification of 4 distinct strain-specific patterns［J］.J Infect Dis，2003，188(1)：19-31.
［11］CAO S，LOU Z，TAN M，et al.Structural basis for the recognition of blood group trisaccharides by norovirus［J］.J Virol，2007，81(11)：5949-5957.
［12］TAN M，MELLER J，JIANG X.C-terminal arginine cluster is essential for receptor binding of norovirus capsid protein［J］.J Virol，2006，80(15)：7322-7331.
［13］CHEN Y，TAN M，XIA M，et al.Crystallography of a Lewis-binding norovirus，elucidation of strain-specificity to the polymorphic human histo-blood group antigens［J］.PLoS Pathog，2011，7(7)：e1002152.
［14］TAN M，HUANG P，MELLER J，et al.Mutations within the P2 domain of norovirus capsid affect binding to human histo-blood group antigens：evidence for a binding pocket［J］.J Virol，2003，77(23)：12562-12571.
［15］CHEN M，SONG Y，FAN Z，et al.Immunogenicity of different recombinant rabbit hemorrhagic disease virus-like particles carrying CD8⁺ T cell epitope from chicken ovalbumin (OVA)［J］.Virus Res，2014，183：15-22.
［16］TAN M，HEGDE R S，JIANG X.The P domain of norovirus capsid protein forms dimer and binds to histo-blood group antigen receptors［J］.J Virol，2004，78(12)：6233-6242.
［17］HUKOWSKA-SZEMATOWICZ B，TOKARZ-DEPTULA B，DEPTULA W.Genetic variation and phylogenetic analysis of rabbit haemorrhagic disease virus (RHDV) strains［J］.Acta Biochim Pol，2012，59(4)：459-465.
［18］PUGGIONI G，CAVADINI P，MAESTRALE C，et al.The new French 2010 rabbit hemorrhagic disease virus causes an RHD-like disease in the Sardinian Cape hare (Lepus capensis mediterraneus)［J］.Vet Res，2013，44：96.
［19］LE GALL-RECULÉ G，LAVAZZA A，MARCHANDEAU S，et al.Emergence of a new lagovirus related to rabbit hemorrhagic disease virus［J］.Vet Res，2013，44：81.
［20］WANG X，HAO H，QIU L，et al.Phylogenetic analysis of rabbit hemorrhagic disease virus in China and the antigenic variation of new strains［J］.Arch Virol，2012，157(8)：1523-1530.
［21］RADEMACHER C，KRISHNA N R，PALCIC M，et al.NMR experiments reveal the molecular basis of receptor recognition by a calicivirus［J］.J Am Chem Soc，2008，130(11)：3669-3675.
［22］TAN M，JIANG X.The P domain of norovirus capsid protein forms a subviral particle that binds to histo-blood group antigen receptors［J］.J Virol，2005，79(22)：14017-14030.
［23］LEUTHOLD M M，DALTON K P，HANSMAN G S.Structural analysis of a rabbit hemorrhagic disease virus binding to histo-blood group antigens［J］.J Virol，2015，89(4)：2378-2387.

兔出血症病毒衣壳蛋白P区二聚体的表达及其与受体结合能力分析

Expression of the P Dimer of Rabbit Hemorrhagic Disease Virus Capsid Protein and Analysis of Its Binding Ability to Receptor

RichHTML

PDF

可视化

被引次数

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 0

编辑推荐

Metrics

本文评价