| 研究課題/領域番号 |
22K15050
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| 研究種目 |
若手研究
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| 配分区分 | 基金 |
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| 審査区分 |
小区分43020:構造生物化学関連
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| 研究機関 | 沖縄科学技術大学院大学 |
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研究代表者 |
Matthews Melissa 沖縄科学技術大学院大学, 生体分子電子顕微鏡解析ユニット, ポストドクトラルスカラー (20866298)
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| 研究期間 (年度) |
2022-04-01 – 2025-03-31
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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 |
2,210千円 (直接経費: 1,700千円、間接経費: 510千円)
2023年度: 1,300千円 (直接経費: 1,000千円、間接経費: 300千円)
2022年度: 910千円 (直接経費: 700千円、間接経費: 210千円)
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| キーワード | cryoEM / RNA / ADAR / RNA editing / structural biology / cryo-electron microscopy / adenosine deaminase / dsRNA binding domain / autoimmune disease |
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| 研究開始時の研究の概要 |
ADARs are enzymes responsible for conversion of adenosine to inosine in double-stranded RNA (dsRNA) in humans. Double-stranded RNA-binding domains (dsRBDs) are present in all ADARs and play a key role in determining which RNAs get edited.
An unhealthy level of editing by ADARs leads to autoimmune and neurological diseases. This project will determine 3D structures of ADAR:dsRNA complexes using cryo-electron microscopy. The structures from this work will provide molecular frameworks for RNA editing technologies which include dsRBDs, enabling the creation of new, versatile therapeutic techniques.
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| 研究実績の概要 |
The protein of interest (adenosine deaminase acting on dsRNA 2, ADAR2) has been successfully characterized in complex with two target dsRNA sequences by cryoEM. To accomplish this, roughly 300,000 particles extracted from 9300 micrographs containing ADAR2:GLI-61bp complex or 225,000 particles extracted from 15,000 micrographs were averaged together. Deaminase and double-stranded RNA binding domains were elucidated to 4-3.5-angstrom resolution, while RNA resolution varied from 6-3.5 angstroms. At the current resolution, cryoEM reconstructions are sufficient to elucidate the locations of deaminase domains and dsRBDs. Although density is not observed for all dsRBDs, novel interactions between protein and RNA can be hypothesized.
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| 現在までの達成度 (区分) |
現在までの達成度 (区分)
3: やや遅れている
理由
Several months were spent troubleshooting unforeseen problems with protein purification, but the issue has now been resolved. Quality sample was prepared and analyzed by cryoEM.
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| 今後の研究の推進方策 |
Work is ongoing to prepare accurate molecular models and perform biochemical experiments to validate these models. After that, work will begin on a manuscript.
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