| Project/Area Number |
09670321
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Virology
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| Research Institution | Kawasaki Medical School |
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Principal Investigator |
OHUCHI Masanobu Faculty of Medicine, Kawasaki Medical School, Assistant Professor, 医学部, 助教授 (80107185)
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| Co-Investigator(Kenkyū-buntansha) |
KAWASAKI Kazunori National Institute of Bioscience and Human-Technology, Researcher, 細胞情報, 主任研究員
MATSUMOTO Akira Faculty of Medicine, Kawasaki Medical School, Professor, 医学部, 教授 (90027318)
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| Project Period (FY) |
1997 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 1999: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1998: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1997: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Keywords | influenza virus / receptor binding affinity / fusion activity / oligosaccharide side chain / fusion pore / Video-FRAP / hemagglutinin / 膜融合反応 / レセプター / 細胞融合能 / 糖側鎖 / 結合力 / HA / 膜融合能 / ノイラミニダーゼ / 赤血球凝集素 |
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| Research Abstract |
Hemagglutinin (HA) of influenza virus mediates the entry of viral genome into the host cell though the membrane fusion process. It was not clear whether the cytoplasmic domain of HA should involve this fusion process or not. We have found that elongation the cytoplasmic domain causes drastic decrease in the fusion activity. This effect depended on the number of amino acid added but was independent of the species of amino acid. Addition of 5 amino acids abolished the cell fusion activity entirely. On the other hand, the cell fusion activity remained even after deletion of the cytoplasmic domain, although the activity was considerably reduced.
To investigate what factor(s) is for the fusion activity, we compared the expression level, oligosaccharide processing, cleavability, acylation, cell surface distribution, and lateral movement of HA in the cellular membrane between the wild type and cytoplasmic domainmodified Has. The lateral movement was assayed with Fab of fluorescein-labeled anti-HA antibody in Video-FRAP (fluorescence recovery after photobleaching). No difference was detected in any factors between both Has. Other parameter must be searched.
We also found that receptor binding affinity of HA is a determinant for the fusion activity, that is, deletion of oligosaccharides near the receptor binding site enhanced the binding affinity and reduced the cell fusion activity proportionally, and the mutation which reduced the binding affinity restored the fusion activity. Thus, the reciprocal relationship was observed between both activities. The HA with high receptor binding affinity formed fusion pores, through which small molecules such as calcein could pass. However, hemoglobin hardly passed through the fusion pore. Enlargement of the fusion pore may be interfered with. Since the viral nucleocapside is larger than hemoglobin, this problem is critical for the viral infection. Accordingly, the control of receptor binding affinity is a vital problem for the virus.
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