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Analytical Techniques for Assaying Nitric Oxide Bioactivity
Published on: June 18, 2012
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酸化窒素は[Fe4S4]クラスター・ニトロシル化時に800mVのマイナスシフトでエンドヌクレアースIIIリドックス活性を調節する
Journal of the American Chemical Society
|August 28, 2018
まとめ
酸化窒素への曝露は,エンドヌクレアゼIII (EndoIII) の鉄硫黄群をダイニトロシル鉄複合体とルーシンに変換する.
科学分野:
- 生化学と分子生物学
- バイオ有機化学
- 電気化学
背景:
- エンドヌクレアースIII (EndoIII) は[Fe4S4]クラスタを含むDNA修復酵素である.
- タンパク質中の鉄硫黄のクラスターは,酸化窒素 (NO) によって変化しやすい.
- DNA修復酵素における[Fe4S4]クラスター・ニトロシル化による機能的影響は十分に理解されていません.
研究 の 目的:
- エンドオIIIの[Fe4S4]クラスタのニトロシル化産物を特徴付ける.
- EndoIIIの酸化還元活性とDNA修復機能に対するニトロシル化の影響を調査する.
- 窒素化がDNA媒介の電荷輸送に影響を与えるメカニズムを解明する.
主な方法:
- DNA改変黄金の電気化学と タンパク質フィルムの電圧測定
- エレクトロフォレティック・モビリティ・シフト・アッセイ (EMSA)
- トリプシンで消化されたタンパク質の質量スペクトロメトリー (MS).
- 超精細サブレベル相関 (HYSCORE) パルス電子パラマグネティック共振 (EPR) スペクトロスコーピー.
主要な成果:
- EndoIIIのニトロシル化により,ディニトロシル鉄複合体と報告されていないルッシン赤エステルが得られます.
- ニトロシル化EndoIIIは,減少ポテンシャル (~800mV) の有意な負のシフトを示している.
- 同様のDNA結合親和性にもかかわらず,EndoIIIのDNA結合再酸化活動は,ニトロシル化によって廃止される.
結論:
- エンドヌクレアースIII [Fe4S4] クラスター・ニトロシル化により,異なる鉄・ニトロシル種が生成される.
- クラスター・ニトロシル化により,EndoIIIの酸化還元特性が大幅に変化する.
- [Fe4S4]クラスタのニトロシレーションは,DNAに結合した酸化還元活性とDNA媒介の電荷輸送を調節する.
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