电荷不成比例对称性破裂产生异体质肌球蛋白复合体,具有增强的亲和力和快速的内部复合电子转移
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
概括
我们设计了一种通过电荷不成比例显著增强结合亲和度的肌球蛋白 (Mb) 二次体. 这一突破使得蛋白质之间的电子转移 (ET) 速度常数增加了五倍以上.
科学领域:
- 生物化学
- 蛋白质工程
- 电子转移动力学
背景情况:
- 肌球蛋白 (Mb) 通常作为单体起作用.
- 在实现稳定的复合体和受控的电子转移方面,工程Mb二极管提出了挑战.
- 之前对Mb二极体中蛋白质间电子转移的研究因动态关而面临限制.
研究的目的:
- 设计一个具有极大增强结合亲和力和受控的复合内电子转移 (ET) 的肌球蛋白 (Mb) 双质体.
- 调查电荷不成比例对Mb二极体形成和ET动力学的影响.
- 开发和验证用于创建功能性蛋白质复合物的新型蛋白质工程策略.
主要方法:
- 布朗动力学 (BD) 模拟用于接口重新设计,包括一种新的"血过"方法.
- 位点定向的突变发生引入互补的正 (Mb(+6) 和负 (Mb(-6) 表面电荷.
- 使用Zn-deuteroporphyrin替代的X射线表达,X射线特征和光启动电子转移 (ET) 光循环测量.
主要成果:
- 成功创建了互补的Mb(+6) 和Mb(-6) 突变,形成了一个稳定的二元体,其边缘距离约为18-19 Å.
- 与野生类型的Mb相比,充电不成比例增加了至少5个数量级的绑定常数.
- 电荷分离 (CS) 和电荷重组 (CR) 的内复合ET速率常数增加了5个数量级,显示出真正的蛋白间ET的粘度独立性.
- 在初始衰变后观察到电荷分离中间体I的前所未有的重新出现,
结论:
- 具有电荷不成比例的工程Mb二元体显著增强了结合亲和力和快速的蛋白间ET.
- 开发的BD模拟和突变生成策略对于设计功能性蛋白质-蛋白质相互作用是有效的.
- 观察到的ET动力学和中间行为为蛋白质复合体中的光启动电子转移机制提供了新的见解.
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