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2-His-1-Glu非ヘム鉄中心をミオグロビンに導入すると,酸化窒素還元酵素の活性が認められる

Ying-Wu Lin¹, Natasha Yeung, Yi-Gui Gao

¹Department of Chemistry, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.

Journal of the American Chemical Society

|July 1, 2010

まとめ

研究者は,精子クジラのミオグロビンに新しい2-His-1-Glu金属センターを設計し,新しい酸化窒素還元酵素 (NOR) モデルを作成しました. このエンジニアリングされたタンパク質は,銅または鉄との有意なNOR活性を示し,酵素機能の洞察を提供しました.

科学分野:

バイオケミストリーバイオケミストリー
酵素工学とは
構造生物学構造生物学とは

背景:

非ヘム鉄酵素は,保存された2−ヒス−1−グル金属の中心を利用する.
精子クジラのミオグルビン (swMb) は,よく特徴づけられたタンパク質であり,エンジニアリングに適しています.

研究の目的:

2-His-1-Gluメタルセンターを swMb.に組み込むために.
異なる金属イオンで設計されたタンパク質の酸化窒素還元酵素 (NOR) 活性を調べる.
構造的および光譜学的研究を通じて,NOR活動のメカニズムを解明する.

主な方法:

サイト・ディレクテッド・ミュータゲネシスは,swMb (Fe(B) Mb(-His)) の2−His-1-Glu金属センターを設計するために使用されました.
高解像度のX線結晶学により,Cu (II) -CN (II) -Fe (B) -Mb (His) の構造が決定されました.
電子パラマグネティック共振 (EPR) スペクトロスコピーは,酸化窒素還元中の反応中間物質を分析した.

主要な成果:

機能的な2-His-1-Glu金属センターが swMb.に成功裏に設計されました.
Cu (I) -Fe (B) -Mb (His) と Fe (II) -Fe (B) -Mb (His) の両方が酸化窒素還元酵素 (NOR) 活性を示し,銅はより高い活性を示した.
構造データとEPRデータは,CuとFe結合酵素の異なる中間物質を明らかにし,金属リガンドと結合金属の役割を強調しました.

結論:

エンジニアリングされたFe(B) Mb(-His) は,NOR.の新しいタンパク質モデルとして機能します.
エンジニアリングされた金属リガンド (Glu29) と結合金属イオン (Cuまたは Fe) の両方が,NORの活性にとって重要である.
この研究は,新たに特定されたgNOR酵素ファミリーについての洞察を提供します.

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