| 研究課題/領域番号 |
23K04928
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| 研究種目 |
基盤研究(C)
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| 配分区分 | 基金 |
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| 応募区分 | 一般 |
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| 審査区分 |
小区分37010:生体関連化学
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| 研究機関 | 東京工業大学 |
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研究代表者 |
Maity Basudev 東京工業大学, 生命理工学院, 特任助教 (60815421)
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| 研究期間 (年度) |
2023-04-01 – 2026-03-31
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| 研究課題ステータス |
交付 (2023年度)
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| 配分額 *注記 |
4,680千円 (直接経費: 3,600千円、間接経費: 1,080千円)
2025年度: 1,430千円 (直接経費: 1,100千円、間接経費: 330千円)
2024年度: 1,430千円 (直接経費: 1,100千円、間接経費: 330千円)
2023年度: 1,820千円 (直接経費: 1,400千円、間接経費: 420千円)
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| キーワード | RNaseA crystal / Mn-carbonyl comples / CO release reaction / X-ray crystal structure / Protein crystal / Multiple binding sites / Chemical reaction |
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| 研究開始時の研究の概要 |
In this research, we aim to construct protein crystal template with multiple metal binding sites to study chemical reaction using serial crystallography. By determining the structures at various time intervals, we can explore the reaction mechanism of various metal catalyzed reaction.
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| 研究実績の概要 |
We used RNaseA as a template to study a reaction in multiple protein environments. We immobilized a Mn-carbonyl complex in RNaseA crystal by soaking. The X-ray crystal structure revealed four different metal binding sites which are located at N-terminal, Asp53, His105 and His119. In all the cases, the density of three CO ligands were observed at the Mn center which showed the characteristic CO stretching frequency at 1931 cm-1. Then, we performed the CO release reaction by exposing the single crystal in UV light. The reaction at Asp53 and His105 was very fast and after 10 min of irradiation, all the CO were released whereas other sites required longer time. This showed the direct effect of protein environment in a reaction.
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| 現在までの達成度 (区分) |
現在までの達成度 (区分)
1: 当初の計画以上に進展している
理由
The reaction is progressing smoothly as we planed. This is because our proposed work was based on some preliminary result (X-ray structure) and then we optimize the metal immobilization and determination of high resolution X-ray structure. Since we obtained the high-resolution X-ray structure, it became easy to understand each reaction center and the local protein environment. Currently, we are focusing on the next step to study the details of CO reaction within the protein crystal.
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| 今後の研究の推進方策 |
As a next plan, we are aiming to study the detail CO release reaction within the protein crystal. We already obtained the preliminary data after the light irradiation. Therefore, we first focus on optimizing the light exposure time to capture the intermediates during the CO release reaction. In addition, we also plan to add coordinating ligand such as imidazole in the buffer solution which are expected to bind to Mn after the release of CO. This will give additional information about the trapped intermediate. We also plan to perform QM calculation to validate the experimental results.
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