Generation of Tailor-made Biocatalysts Using Immune System.
| Project/Area Number |
16350091
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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 |
Chemistry related to living body
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| Research Institution | Osaka Prefecture University |
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Principal Investigator |
FUJII Ikuo Osaka Prefecture University, Graduate School of Science, Professor, 理学系研究科, 教授 (70189984)
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| Co-Investigator(Kenkyū-buntansha) |
TSUMURAYA Takeshi Osaka Prefecture University, Graduate School of Science, Instructor, 理学系研究科, 助手 (00372855)
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| Project Period (FY) |
2004 – 2006
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| Project Status |
Completed (Fiscal Year 2006)
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| Budget Amount *help |
¥15,700,000 (Direct Cost: ¥15,700,000)
Fiscal Year 2006: ¥4,600,000 (Direct Cost: ¥4,600,000)
Fiscal Year 2005: ¥4,600,000 (Direct Cost: ¥4,600,000)
Fiscal Year 2004: ¥6,500,000 (Direct Cost: ¥6,500,000)
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| Keywords | catalytic antibody / transition-state analog / antigen / monoclonal antibody / hybridoma / immunization / co-factor / 抗体触媒 / ハプテン / 補酵素 / ファージライブラリー |
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| Research Abstract |
From the first report of catalytic antibodies in 1986, a variety of the antibodies has been generated and found to catalyze chemical reactions from peri-cyclic reactions to amido-bond hydrolyses. Most of them are elicited by immunization of transition-state analogs. However, unfortunately, the catalytic activities are not sufficient. Therefore, we take advantage of catalytic power of cofactors. In this work, we aim to incorporate synthetic cofactors into antibody-combining sites to generate catalytic antibodies.
A bi-functional hapten was designed and synthesized to elicit both substrate and cofactor binding sites in antibody-combining sites. After immunization to mice with the carrier protein (KLH)-hapten conjugate, we isolated 50 monoclonal antibodies by hybridoma technology. Of the antibodies, 22 were found to be catalytic in acyl-transfer reactions of p-nitrophenol ethyl esters with synthetic co-factors attaching alcohol, amino, and sulfur groups. The kinetic parameters (K_m, VK_<max>,k_<cat> ) for the most active antibodies (27C1 and 25E2) were examined in detail. Furthermore, antibody 27C1 also catalyzed elimination reactions of β-fluoroketone to give the corresponding enone when the synthetic co-factor attaching carboxylate was used. In this work, we have demonstrated generating catalytic antibodies with synthetic cofactors and controlling the catalytic activity by changing the co-factors. Because antibodies can be generated virtually any molecule of interest, combination of antibodies and co-factors should lead to a new class of biocatalysts with tailor-made specificity and reactivity.
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Report
(4 results)
Research Products
(50 results)
