Serving the Society Since 1986
Advanced Search
Volume 25 Issue 3
June 2010
    Citation: ZHAO Lei, REN Xiao-ming, ZHENG Alan C.. Herpes Simplex Virus Type 1 ICP27 Protein: Its Expression, Purification and Specific Antiserum Production .VIROLOGICA SINICA, 2010, 25(3) : 199-205.  http://dx.doi.org/10.1007/s12250-010-3116-2
    shu

    Herpes Simplex Virus Type 1 ICP27 Protein: Its Expression, Purification and Specific Antiserum Production

    • 1.

      State Key Laboratory, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, China

    • 2.

      The Depautment of Animal Science, Beijing University of Agriculture, Beijing 102206, China

    • Corresponding author: ZHENG Alan C., 
    • Received Date: 11 December 2009
      Accepted Date: 01 February 2010
      Available online: 01 June 2010

      Fund Project: Open research fund program of the state key laboratory of virology of China 2009007Hubei province natural science foundation of innovation groups project 2008CDA013Open research fund program of the state key laboratory of virology of China 2007003National natural science foundation of China 30870120Major state basic research development program (973 Program) of China 2010CB 530105The startup fund of the hundred talents program of the Chinese academy of science 20071010-141

    • Herpes simplex virus type 1 (HSV-1) is the causative agent of cold sores and other more serious diseases. HSV-1 infected-cell protein 27 (ICP27) is an immediate-early regulatory phosphoprotein homologous to gene products identified in all classes of herpesviruses so far. To raise the antiserum to ICP27 for further characterization of its biological function, the ICP27 gene was cloned into the pET-28a (+) vector, then ICP27 protein was expressed in E. coli and purified by nickel-nitrilotriacetic acid (Ni2+-NTA) affinity resin column, finally the purified protein was used to raise antiserum. Western blot analysis demonstrated that the antiserum recognized the recombinant protein, and the antiserum was able to probe the ICP27 in HSV-1 infected cells with high specificity by immunofluorescence assay (IFA). Therefore, the specific antiserum will provide a valuable tool for further studies investigating ICP27’s biological function during HSV-1 infection.
    • 加载中

      1. Arnau J, Lauritzen C, Petersen G E, et al. 2006. Current strategies for the use of affinity tags and tag removal for the purification of recombinant proteins. Protein Expr Purif, 48 (1): 1-13.
          doi: 10.1016/j.pep.2005.12.002

      2. Brown C R, Nakamura M S, Mosca J D, et al. 1995. Herpes-Simplex Virus Transregulatory Protein Icp27 Stabilizes and Binds to 3'-Ends of Labile Messenger-Rna. J Virol, 69 (11): 7187-7195.

      3. Fontaine-Rodriguez E C, and Knipe D M. 2008. Herpes simplex virus ICP27 increases translation of a subset of viral late mRNAs. J Virol, 82 (7): 3538-3545.
          doi: 10.1128/JVI.02395-07

      4. Hardy W R, and Sandri-Goldin R M. 1994. Herpes simplex virus inhibits host cell splicing, and regulatory protein ICP27 is required for this effect. J Virol, 68 (12): 7790-7799.

      5. Hunt I. 2005. From gene to protein: a review of new and enabling technologies for multi-parallel protein expression. Protein Expr Purif, 40 (1): 1-22.
          doi: 10.1016/j.pep.2004.10.018

      6. Johnson K E, Song B, and Knipe D M. 2008. Role for herpes simplex virus 1 ICP27 in the inhibition of type Ⅰ interferon signaling. Virology, 374 (2): 487-494.
          doi: 10.1016/j.virol.2008.01.001

      7. Koffa M D, Clements J B, Izaurralde E, et al. 2001. Herpes simplex virus ICP27 protein provides viral mRNAs with access to the cellular mRNA export pathway. EMBO J, 20 (20): 5769-5778.
          doi: 10.1093/emboj/20.20.5769

      8. Markovitz N S. 2007. The herpes simplex virus type 1 UL3 transcript starts within the UL3 open reading frame and encodes a 224-amino-acid protein. J Virol, 81 (19): 10524-10531.
          doi: 10.1128/JVI.00123-07

      9. Mayer M, and Buchner J. 2004. Refolding of inclusion body proteins. Methods Mol Med, 94239-94254.

      10. McGeoch D J, Dalrymple M A, Davison A J, et al. 1988. The complete DNA sequence of the long unique region in the genome of herpes simplex virus type 1. J Gen Virol, 69 (Pt 7): 1531-1574

      11. Mclauchlan J, Phelan A, Loney C, et al. 1992. Herpes-Simplex Virus Ie63 Acts at the Posttranscriptional Level to Stimulate Viral Messenger-Rna 3' Processing. J Virol, 66 (12): 6939-6945.

      12. Mears W E, Lam V, and Rice S A. 1995. Identification of nuclear and nucleolar localization signals in the herpes simplex virus regulatory protein ICP27. J Virol, 69 (2): 935-947.

      13. Phelan A, and Clements J B. 1997. Herpes simplex virus type 1 immediate early protein IE63 shuttles between nuclear compartments and the cytoplasm. J Gen Virol, 78: 3327-3331.
          doi: 10.1099/0022-1317-78-12-3327

      14. Ryou C, Prusiner S B, and Legname G. 2003. Cooperative binding of dominant-negative prion protein to kringle domains. J Mol Biol, 329 (2): 323-333.
          doi: 10.1016/S0022-2836(03)00342-5

      15. Sandri-Goldin R M. 1998. ICP27 mediates HSV RNA export by shuttling through a leucine-rich nuclear export signal and binding viral intronless RNAs through an RGG motif. Genes Dev, 12 (6): 868-879.
          doi: 10.1101/gad.12.6.868

      16. Sandri-Goldin R M. 2008. The many roles of the regulatory protein ICP27 during herpes simplex virus infection. Front Biosci, 13: 5241-5256.

      17. Sedlackova L, and Rice S A. 2008. Herpes simplex virus type 1 immediate-early protein ICP27 is required for efficient incorporation of ICP0 and ICP4 into virions. J Virol, 82 (1): 268-277.
          doi: 10.1128/JVI.01588-07

      18. Tanaka M, Kagawa H, Yamanashi Y, et al. 2003. Construction of an excisable bacterial artificial chromosome containing a full-length infectious clone of herpes simplex virus type 1: Viruses reconstituted from the clone exhibit wild-type properties in vitro and in vivo. J Virol, 77:1382-1391.
          doi: 10.1128/JVI.77.2.1382-1391.2003

    • 加载中

    Figures(5)

    PDF

    Article Metrics

    Article views(4193) PDF downloads(11) Cited by()

    Related
    Proportional views

      Herpes Simplex Virus Type 1 ICP27 Protein: Its Expression, Purification and Specific Antiserum Production

        Corresponding author: ZHENG Alan C.,
      • 1. State Key Laboratory, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, China
      • 2. The Depautment of Animal Science, Beijing University of Agriculture, Beijing 102206, China
      • Received Date: 11 December 2009
      • Accepted Date: 01 February 2010
      Fund Project:  Open research fund program of the state key laboratory of virology of China 2009007Hubei province natural science foundation of innovation groups project 2008CDA013Open research fund program of the state key laboratory of virology of China 2007003National natural science foundation of China 30870120Major state basic research development program (973 Program) of China 2010CB 530105The startup fund of the hundred talents program of the Chinese academy of science 20071010-141

      Abstract: Herpes simplex virus type 1 (HSV-1) is the causative agent of cold sores and other more serious diseases. HSV-1 infected-cell protein 27 (ICP27) is an immediate-early regulatory phosphoprotein homologous to gene products identified in all classes of herpesviruses so far. To raise the antiserum to ICP27 for further characterization of its biological function, the ICP27 gene was cloned into the pET-28a (+) vector, then ICP27 protein was expressed in E. coli and purified by nickel-nitrilotriacetic acid (Ni2+-NTA) affinity resin column, finally the purified protein was used to raise antiserum. Western blot analysis demonstrated that the antiserum recognized the recombinant protein, and the antiserum was able to probe the ICP27 in HSV-1 infected cells with high specificity by immunofluorescence assay (IFA). Therefore, the specific antiserum will provide a valuable tool for further studies investigating ICP27’s biological function during HSV-1 infection.

        HTML