光解離性ケージド基質を利用した時間分解構造解析による酵素反応の可視化

書誌事項

タイトル別名
  • Visualization of Enzymatic Reaction by Time-resolved Structural Analysis with Photosensitive Caged Substrate

抄録

<p>Elucidation of molecular mechanism for metalloeznyme-catalyzed reactions is longstanding topics in the bio-inorganic chemistry field, since metalloeznymes play pivotal roles in various biological processes and catalyze reactions with high efficiency under mild conditions. Structural determination of reaction intermediates is key to visualizing the reactions catalyzed by metalloenzymes. Although it is not easy to determine the structure of the transient species by conventional crystallography, newly developed time-resolved X-ray crystallography using X-ray free electron laser (XFEL) has a great potential for the structural characterization of intermediates. However, XFEL-based time-resolved crystallography requires a photo-trigger, hampering its application to non-photosensitive proteins like metalloenzymes. Here, to overcome this issue, we focused on caged substrates, which produce the substrate upon photo-irradiation, to introduce the photo-trigger into the reaction catalyzed by the metalloenzyme. To demonstrate the usefulness of caged substrates for the trigger of XFEL-based time-resolved crystallography, soluble nitric oxide (NO) reductase, which catalyzes the reduction of NO to nitrous oxide (N2O) at a heme active center, and caged NO were used as a model system. Time-resolved spectroscopic analysis showed that the photolysis of caged NO could initiate NO reduction by P450nor in the micro-crystals. Time-resolved crystallography using XFEL enabled us to determine the structures of two intermediates; NO-bound form and subsequent NO-activated form, which provided a unique opportunity to draw the complete picture of the reaction cycle of P450nor. Thus, the combination of caged substrate and XFEL-based time-resolved crystallography is an invaluable method for the visualization of the reactions catalyzed by metalloenzyems.</p>

収録刊行物

  • 薬学雑誌

    薬学雑誌 142 (5), 487-494, 2022-05-01

    公益社団法人 日本薬学会

参考文献 (25)*注記

もっと見る

詳細情報 詳細情報について

問題の指摘

ページトップへ