| Project/Area Number |
08558072
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 展開研究 |
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| Research Field |
Structural biochemistry
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| Research Institution | Kyushu University |
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Principal Investigator |
UEDA Tadashi Kyushu Univ., Grad.Sch.Pharm.Sci., Assoc.Prof., 大学院・薬学研究科, 助教授 (90184928)
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| Co-Investigator(Kenkyū-buntansha) |
KUROKI Ryota Kirin Brewery Co.Ltd., Central Lab.for Key Technology, Senior Scientist, 基盤技術研究所, 主任研究員
IMOTO Taiji Kyushu Univ., Grad.Sch.Pharm.Sci., Prof., 大学院・薬学研究科, 教授 (90038282)
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| Project Period (FY) |
1996 – 1997
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| Project Status |
Completed (Fiscal Year 1997)
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| Budget Amount *help |
¥8,300,000 (Direct Cost: ¥8,300,000)
Fiscal Year 1997: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1996: ¥6,800,000 (Direct Cost: ¥6,800,000)
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| Keywords | protein / folding / dialysis / immunoglobulin G / lysozyme / リボヌクレアーゼA / タカアミラーゼA / エノラーゼ / 単鎖Fv / インクルージョンボディ / クエン酸シンターゼ |
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| Research Abstract |
Recently, we have developed the effective renaturation method of denatured and reduced proteins using slow dialysis based on the function of a molecular chaperone. The method was applicable to renature both hen egg-white lysozyme and monoclonal antibody (IgG1/kappa) effectively from their denatured and reduced form. The aim of this project is to investigate whether or not the renaturation method is applicable to renature the denatured and reduced the other proteins. First, since a denatured and reduced monoclonal antibody (IgG1/kappa) effectively renatured to intact by use of the slow dialysis method, I analyzed a renaturation mechanism of the monoclonal antibody from denatured and reduced form in vitro. As a result, intact form was found to form by the favorable interaction between folded heavy chain and light chain. Next, the slow dialysis method was shown to have an advantage in the effective renaturation of denatured and reduced lysozyme, which had been expressed from E.coli or in which methionine residues had been labeled with ^<13>Cnuclei. Moreover, the method was applicable to the effective renaturation of denatured and reduced monomeric protein such as ribonuclease A and Take-amylase A and oligomeric protein such as enolase. For the renaturation of unstable proteins, the addition of glycerol and ammoniumsulfate to the ranturation buffer was also shown to be effective.
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