| Project/Area Number |
15580305
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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 |
Applied molecular and cellular biology
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| Research Institution | Iwate University |
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Principal Investigator |
TAIRA Hideharu Iwate University, Faculty of Agriculture, Professor, 農学部, 教授 (70045756)
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| Co-Investigator(Kenkyū-buntansha) |
YAMASHITA Tetsuro Iwate University, Faculty of Agriculture, Associate Professor, 農学部, 助教授 (20202377)
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| Project Period (FY) |
2003 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2004: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 2003: ¥2,700,000 (Direct Cost: ¥2,700,000)
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| Keywords | Sendai virus / membrane protein / molecular chaperon / HN protein / N-glycosylation / hemagglutinin / neuraminidase / cellular transport |
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| Research Abstract |
The roles of N-linked glycosylation in the intracellular transport and biological activities of the Sendai virus hemagglutinin-neuraminidase (HN) proteins were studied. The protein contained four potential N-glycosylation sites ; g1, g2, g3 and g4. By site-directed mutagenesis of these positions, the mature protein contained three N-linked oligosaccharides attached to g1, g3 and g4. The role of each added oligosaccharide in the structure and functions of the protein was identified by characterization of surface expression, hemadsorption, and neuraminidase activities of the corresponding mutant proteins. The elimination of oligosaccharide-attachment sites g3 or g4 located in the globular head decreased the transport of the HN protein to the cell surface and the immunological reactivity to the globular domain specific monoclonal antibody. The elimination of sites of g1 or g2 did not suppress the intracellular transport and the biological activities.
A rapid and sensitive biosensor method for the interaction of the glycosylation mutant protein with antibody and with molecular chaperon was developed. Monoclonal antibody to HN protein or peptide antibody to BiP was immobilized on a sensor chip. Both monoclonal and peptide antibodies showed good binding with the cell lysates transfected with the expression plasmid of the HN glycosylation mutant. Glycosylation-defective HN mutants should be useful for elucidating the roles of individual oligosaccharide chains in the folding of HN proteins.
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