既存のタンパク質・スキャフォールドによる新しい触媒活動の設計と進化
Hee-Sung Park1, Sung-Hun Nam, Jin Kak Lee
1Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 373-1, Kusung-Dong, Yusung-Gu, Daejon 305-701, Korea.
まとめ
タンパク質工学は,新しい酵素構造にベータ-ラクタマース活性を成功裏に導入しました. この進化した酵素は,セフォタキシムを効率的に水分解し,細菌の耐性を高めます.
科学分野:
- プロテイン工学は,タンパク質の
- 酵素設計 酵素デザイン
- バイオカタリシス バイオカタリシス
背景:
- 新しい機能を持つ酵素を設計することは,タンパク質工学の重要な目標です.
- 既存のタンパク質の構造を改変することは,新しい生物触媒を作り出すための有望な戦略です.
研究 の 目的:
- 活性サイトループを改変することによって,酵素の触媒活動を変化させる方法を開発する.
- ベータ-ラクタマース活性をグリオキサラゼII基架に導入する.
主な方法:
- アクティブサイトループの同時挿入,削除,および置換.
- ポイント変異によって酵素の活性を微調整する.
- グリオキサラーゼIIのアルファベタ/ベタアルファメタルヒドロラーゼ基板の設計.
主要な成果:
- 新しい酵素evMBL8 (進化したメタロベータ-ラクタマース8) が,ベータ-ラクタマース活性で作成されました.
- エンジニアリングされた酵素は,元のグリオキサラゼIIの活性を失いました.
- EvMBL8は,1.8 x 10 (((2) (モル/リットル) ((-1) 秒 ((-1)) の (kcat/Km) appでセフォタキシム水解を効率的に触媒化しました.
結論:
- アクティブサイトループを改変する戦略は,新しい機能を持つ酵素を作るのに有効です.
- エンジニアリングされたevMBL8は,重要なβ-ラクタマース活性を示し,cefotaxime.toにEscherichia coliの成長に対する実質的な耐性を授与します.
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