| Project/Area Number |
05454076
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
食品科学・栄養科学
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| Research Institution | KYUSHU UNIVERSITY |
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Principal Investigator |
MURAKAMI Hiroki Kyushu Univ.DIVISION OF AGRICULTURE, 大学院・農学研究科, 教授 (60038271)
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| Co-Investigator(Kenkyū-buntansha) |
ASO Youichi Kyushu Univ.DIVISION OF AGRICULTURE ASSOCIATE PROFESSOR, 大学院・農学研究科, 助教授 (10117054)
OGATA Seiya Kyushu Univ.DIVISION OF AGRICULTURE PROFESSOR, 大学院・農学研究科, 教授 (20038277)
MUKAI Junichiro Kyushu Univ.DIVISION OF AGRICULTURE PROFESSOR, 大学院・農学研究科, 教授 (70038199)
SHIRAHATA Sanetake Kyushu Univ.DIVISION OF AGRICULTURE ASSOCIATE PROFESSOR, 大学院・農学研究科, 助教授 (90154377)
TACHIBANA Hirofumi Kyushu Univ.DIVISION OF AGRICULTURE ASSOCIATE PROFESSOR, 大学院・農学研究科, 講師 (70236545)
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| Project Period (FY) |
1993 – 1994
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| Project Status |
Completed (Fiscal Year 1994)
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| Budget Amount *help |
¥6,400,000 (Direct Cost: ¥6,400,000)
Fiscal Year 1994: ¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 1993: ¥4,100,000 (Direct Cost: ¥4,100,000)
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| Keywords | Monoclonal antibody / Glycosylation / Foods / Animal cell / Light chain / Culture engineering / Medium / Monosaccharide / 培養工学 / モノクローナル坑体 / 糖鎖 / コンカナバリンA / 有用物質 |
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| Research Abstract |
Although immunoglobulin light chains usually lack carbohydrates, some light chains contain carbohydrate chains in their variable region. We have found a N-glycosylated carbohydrate chain on the light chain-hypervariable region of a human monoclonal antibody which is reactive to lung adenocarcinoma and is produced by a human hybridoma. A carbohydrate chain linked to one of the light chain glycoforms is characterized as hybird-type, which is rare for any immunoglobulin isotype. To clarify the role of carbohydrates in the light chain variable region, we attempted to modify the glycosylation on this paticular light chain. Carbohydrate moiety changes on this light chain produced in concanavalin A-resistant hybridoma clones and the following treatment of these variant light chains with various glycosidases lead to an alteration in the antigen binding activity. It has become increasingly clear that the glycosylation of glycoproteins in mammalian cells is dependent on the culture environments.
… More
Thus we have characterized the effects of different monosaccharides availability in the culture medium on light chain glycosylation and the resulting biological properties of the antibodies. Defferent glucose concentrations in the culture medium lead to an altered antigen-binding ability, which is shown to be a result of a glycosylation alteration on the light chain. When the cells were cultured in the presence of various kinds of monosaccharides substituting glucose, the produced antibodies again altered their antigen-binding activities. Analysis of the antibody light chains produced under these conditions revealed substantial changes in light chain-glycosylation. Glycosylation authenticity and consistency for the production of therapeutic glycoproteins are of particular interest in animal cell cultures. Glycosylation of glycoproteins is dependent on the culture environments of the host cell. If the cell itself is lacking sensitivity to environmental changes, the expression pattern of a certain glycoform is expected to be reproducible, and glycosylation variations caused by a changing culture environment should be reduced. Therefore we attemped to screen cell clones tolerance to glucose availability variations from glycosylation mutants. Cell clones lacking sensitivity to a glucose level change for light chain glycosylation were screened from the lectin-resistant variants to obtain clones which produce glycoforms reproducibly in various culture environments. Less
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