負荷-不均衡対称性の破裂は,強化された親和性および急速な内部複合体電子移転を持つ異体ミオグロビン複合体を生成する
Ethan N Trana1, Judith M Nocek1, Jon Vander Woude1
1Department of Chemistry, Northwestern University , Evanston, Illinois 60208, United States.
Journal of the American Chemical Society
|September 21, 2016
まとめ
マイオグロビン (Mb) ダイマーを設計し 充電不均衡により 結合親和性が著しく向上しました この突破により 速度の定数が 5 度以上増加した タンパク質間電子移転 (ET) が可能になりました
科学分野:
- 生物化学
- タンパク質工学
- 電子伝送ダイナミクス
背景:
- ミオグロビン (Mb) は典型的には単体として機能する.
- 安定した複合体と制御された電子移転を達成する上で,エンジニアリングのMbダイマーには課題があります.
- Mb二重体におけるタンパク質間電子移転に関する以前の研究は,ダイナミックゲートによる制限に直面した.
研究 の 目的:
- マイオグルビン (Mb) ダイマーを設計し,結合親和性が大きく強化され,複合体内電子移転 (ET) が制御される.
- 負荷の不均衡がMbダイマー形成とET運動に及ぼす影響を調査する.
- 機能的なタンパク質複合体を作るための新しいタンパク質工学戦略を開発し,検証する.
主な方法:
- 新しい"ヘムフィルタリング"アプローチを含むインターフェース再設計のためのブラウンのダイナミクス (BD) シミュレーション.
- 補完的な陽性 (Mb(+6) と陰性 (Mb(-6) の表面電荷を導入するためのサイト指向型変異.
- 表現,X線特性,そしてZn-デュテロポルフィリン置換のMbを用いたフォトイニシアチブ電子移転 (ET) のフォトサイクル測定.
主要な成果:
- ~18-19 Å の距離でほぼ平行なヘムを持つ安定したジマーを形成する補完的なMb(+6) とMb(-6) ミュータントが成功しました.
- 負荷の不均衡は,ワイルド型と比較して少なくとも5倍の結合定数を増加させた.
- 電荷分離 (CS) と電荷再結合 (CR) の内複合ET速度の定数は5倍以上増加し,真のタンパク質間ETを示す粘度独立性を示した.
- 初期衰退後に 充電分離中間体Iの 再出現が観測され, 複雑な運動行動を示唆した.
結論:
- 充電不釣り合いの設計されたMbダイマーは,劇的に強化された結合親和性と急速なインタータンパク質ETを示します.
- 開発されたBDシミュレーションと突然変異戦略は,機能的なタンパク質相互作用を設計するのに有効です.
- 観察されたET運動と中間行動は,タンパク質複合体における光誘発電子移転のメカニズムに関する新しい洞察を提供します.
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