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Analysis of mode of gene expression of human parvovirus and application of the recombinant viral protein in clinical diagnosis

Research Project

Project/Area Number 63570212
Research Category

Grant-in-Aid for General Scientific Research (C)

Allocation TypeSingle-year Grants
Research Field Virology
Research InstitutionKyushu University

Principal Investigator

SATO Hiroyuki  Kyushu University, Faculty of Medicine, Lecturer, 医学部, 講師 (70136456)

Project Period (FY) 1988 – 1989
Project Status Completed (Fiscal Year 1989)
Budget Amount *help
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1989: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1988: ¥1,300,000 (Direct Cost: ¥1,300,000)
KeywordsHuman Parvovirus / Monoclonal Antibody / Neutralizing Antibody / Gene Expression / モノクローナル抗体 / ウイルスゲノムのクローニング
Research Abstract

Human parvovirus B19 can replicate only in erythroid precursors in bone marrow and fetal liver in vitro. This extremely dependence on host cell differentiation causes the limited materials of the virus and makes difficult the early diagnosis of the infection and vaccination for high risk group patient such as recipient of bone marrow transplantation, patients suffering from hemolytic anemias and pregnant women. In this study, we identified the site of viral capsid protein which involved the neutralization of the virus, using monoclonal antibody and site specific synthetic polypeptides, and also successfully observed the expression of VP-2 in mammalian cells and E.coli which could be used for the antigen for the serological diagnosis and vaccination as follow.
1.Identification of neutralization responsive site in VP-2 We obtained a monoclonal antibody BE11 which protected decrease of CFU-e in cultured human bone marrow cells challenged by the virus in vitro. The site of epitope of this monoclonal antibody was determined using 10 synthetic polypeptides deduced by the amino acid sequence of VP-2. BE11 only reacted a peptide which represented from 328th to 344th amino acid from the N-terminal of VP-2.
2.Gene expression of VP-2 in mammalian cells Accumulation of 58 kd of viral protein in the nucleus was observed in COS monkey cells transfected a plasmid containing SV40 late promoter and coding region of VP-2. This suggests the existence of nuclear localization signal in VP-2.
3.Gene expression of VP-2 in E.coli. A plasmid was constructed by cloning the coding region of VP-2 into the downstream of the coding region of signal peptide of outer membrane protein of E.coli. A 58 kd of protein having the antigenicity of VP-2 was successfully expressed in the periplasmic space of E.coli. We are now setting up the assay system for clinical diagnosis using this recombinant protein.

Report

(3 results)
  • 1989 Annual Research Report   Final Research Report Summary
  • 1988 Annual Research Report
  • Research Products

    (6 results)

All Other

All Publications (6 results)

  • [Publications] Hiroyuki Sato: "Identificatin of the neutralization responsive site in the major structural protein of human parvovirus B19."

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Hiroyuki Sato: "Expression of the major structural protein of human parvovirus B19 in E.coli."

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Hiroyuki Sato: "Identification of the neutralization responsive site in the major structural protein of human parvovirus B19."

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Hiroyuki Sato: "Expression of the major structural protein of human parvovirus B19 in E.coli"

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1989 Final Research Report Summary
  • [Publications] Hiroyuki Sato: "Identification of the neutralization responsive site in the major structural protein of human parvovirus B19."

    • Related Report
      1989 Annual Research Report
  • [Publications] Hiroyuki Sato: "Expression of the major structural protein of human parvovirus B19 in E.coli."

    • Related Report
      1989 Annual Research Report

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Published: 1988-04-01   Modified: 2016-04-21  

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