Experimental analysis of the electrostatic repulsion between enzyme and substrates to elucidate the reaction mechanism, and development anti-malaria agent
| Project/Area Number |
16H04751
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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 |
Structural biochemistry
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| Research Institution | Kyoto University |
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Principal Investigator |
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| Project Period (FY) |
2016-04-01 – 2019-03-31
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| Project Status |
Completed (Fiscal Year 2019)
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| Budget Amount *help |
¥17,680,000 (Direct Cost: ¥13,600,000、Indirect Cost: ¥4,080,000)
Fiscal Year 2018: ¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000)
Fiscal Year 2017: ¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000)
Fiscal Year 2016: ¥9,360,000 (Direct Cost: ¥7,200,000、Indirect Cost: ¥2,160,000)
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| Keywords | 酵素 / X線結晶構造解析 / 結晶構造 / プロトン化状態 / 酵素反応 / 構造生物学 / タンパク質 / 結晶構造解析 / 基質の歪み / オロチジン一リン酸脱炭酸酵素 |
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| Outline of Final Research Achievements |
In order to elucidate the details of the destabilization of the enzyme-substrate complex, we established a base for high-resolution X-ray crystallographic analysis while avoiding the influence of crystal damage by X-rays. First, we enabled large-scale sample preparation comparing to several times larger than conventional ones. We then identified the crystal form which is the most suitable for analysis and established the protocol for its preparation. Also, we analyzed the relationship between the absorbed X-ray dose of the crystal and the damage of the internal structure of the enzyme. The analysis enabled us to collect damage-less diffraction dataset. The best resolution of the obtained dataset was 0.99 angstrom.
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| Academic Significance and Societal Importance of the Research Achievements |
酵素基質複合体の不安定化は、理論的には全ての酵素が利用可能な機構であるにもかかわらず、実験的な解析例はほとんど無い。また、酵素と基質の間には引力が働くと考えるのが常識であり、静電的な反発が活性に関わるとの報告は存在しない。本研究はこのように、理論的に可能でありながら、これまでに報告例のない機構の解明を目指して、これに必要な試料の調製方法の確立、データ取得手法の検討、ならびにデータ取得などを行った。
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Report
(4 results)
Research Products
(14 results)
