Location of Protective Epitopes on the E Protein of Japanese Encephalitis Virus and its Expression
| Project/Area Number |
61570233
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Virology
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| Research Institution | Tokyo Metropolitan Institute for Neurosciences |
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Principal Investigator |
YASUI Kotaro Tokyo Metropolotan Institue for Neurosciences.Senior Research Scientist, 微生物, 副参事 (90073080)
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| Co-Investigator(Kenkyū-buntansha) |
MIYAMOTO Michiko Tokyo Metropolitan Institute for Neurosciences,Research Scientist, 微生物, 主事 (40190821)
OGIMOTO Mami Tokyo Metropolotan Institute for Neurosciences.Research Scientist, 微生物, 主事 (80158609)
KIMURA-KURODA Junko Tokyo Metropolitan Institute for Neurosciences.Research Scientist, 微生物, 主事 (20142151)
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| Project Period (FY) |
1986 – 1987
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| Project Status |
Completed (Fiscal Year 1987)
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| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1987: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1986: ¥1,200,000 (Direct Cost: ¥1,200,000)
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| Keywords | Japanese encephalitis virus / GlycoproteinE / Protective epitope / monoclonal antibody / monoclonal antibody escaped mutant / Expression of E protein / recombinant baculovirus / コンポーネントワクチン / 日本脳炎ウイルス / フラビウイルス / 感染防御抗原 / エピトープモノクローナル抗体 / 発現ベクター / アミノ酸配列 |
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| Research Abstract |
cDNA of the genomic RNA of Japanese encephalitis(JE) virus were molecularly cloned and sequenced. Amino acid sequences of the E protein were deduced from the base sequnce data.The protein was composed 3 domain structures and epitopes were located mainly on these domains.The E proteins of flaviviruses might have the similar domain structure because amino acid sequence homology of the E proteins was very high between JE virus and other flaviviruses and all cystein residues on the E proteins were completly conserved among the flaviviruses sequenced.
The protective epitopes were located restricted sites on the folded E protein.Monoclonal antibody escaped mutants of the protective epitopes were isolated and their base sequences of the E protein gene were analysed.Protective epitopes might be located on the N terminal domain of the E protein because the amino acid replacement of these monoclonal antibody escaped mutants were observed on the N terminal domain.
The E proteins from the constructs which contained the E protein gene and other parts of the structural and nonstructural protein genes were expressed by using recombinant baculovirus,SV40 plasmid and in vitro translation system.Among them similar structural E protein to the JE virion , that is natively folded E protein,was expressed from the construct which contained pre M,E and SSl sequences,however unfolded denature form E proteins were expressed from the constructs which containd only E sequence.To express natively folded E protein preM sequence might be important at upstream of the E protein sequence.It became possible to develop the compornent E protein vaccine and diagnostic reagents using the recombinant baculovirus because the animals immunized with the E protein which was expressed by the recombinant baculovirus contained JE virus preM,E and NSl protein genes could produced high titred neutralizing antibodies.
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Report
(2 results)
Research Products
(12 results)
