| Project/Area Number |
11694094
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| Research Category |
Grant-in-Aid for Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Functional biochemistry
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| Research Institution | Himeji Institute of Technology |
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Principal Investigator |
KOIDE Takehiko Himeji Institute of Technology, Faculty of Science, Department of Life Science, Professor, 理学部, 教授 (60018695)
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| Co-Investigator(Kenkyū-buntansha) |
TOKUNAGA Fuminori Himeji Institute of Technology, Faculty of Science, Department of Life Science, Research Associate, 理学部, 助手 (00212069)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥5,000,000 (Direct Cost: ¥5,000,000)
Fiscal Year 2000: ¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 1999: ¥2,600,000 (Direct Cost: ¥2,600,000)
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| Keywords | Quality Control / ER-Associated Degradation / Molecular Chaperones / Antithrombin / Mannosidase I / Edem / Mannose Trimming / Thyroglobulin / 新生タンパク質 / 小胞体品質管理機構 / アンチトロンビン変異体 / 分泌異常 / タンパク分解 / プロテインC / 変異タンパク質 |
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| Research Abstract |
We investigated the quality control mechanism of newly synthesized proteins in the endoplasmic reticulum (ER) by using under-γ-carboxylated protein C (uc-PC), antithrombin (AT) ΔGlu313 and Pro429stop mutants and cogTg (thyroglobulin Leu2266Pro mutant) as models of misfolded proteins. [Results] 1. Analyses of the effects of high level co-expression of molecular chaperones (BiP, ERp72, ER94 or PDI) in CHO cells on the secretion and degradation of misfolded proteins. 1) In cells expressing a high level of BiP, PDI or ERp72, even wild-type PC was degraded without secretion. 2) Degradation of uc-PC was enhanced with co-expression of ERp72. In contrast, it was rather suppressed in cases of BiP and PDI.3) Wild-type AT was also partially degraded in CHO cells co-expressing BiP or ERp72. 4) Degradation of ΔGlu313 AT was enhanced with co-expression of BiP or ERp72, while that of Pro429stop AT was enhanced only with ERp72. 5) CogTg co-expressed with BiP or ER94 was mostly retained and accumulated in the ER.Thus, the effects of over-expression of molecular chaperones were quite variable among proteins.2. Mannose trimming of N-linked oligosaccharide(s) is proposed to be important for quality control of newly synthesized proteins and formation of Man8B form is critical for targeting misfolded glycoproteins to the degradation pathway. We examined the effects of a high level of co-expression of mannosidase I (whose action generates a Man8B form) or Edem (mannosidase I homologue) on the degradations of ΔGlu313 AT and Pro429stop AT.The result showed that degradations of both AT mutants were enhanced in CHO cells co-expressing a high level of Edem. This result strongly suggests that Edem is a key factor which introduces misfolded glycoproteins into the degradation pathway.
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