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一个双核CuA中心设计在全α螺旋蛋白质支架中
Evan N Mirts1,2, Sergei A Dikanov3, Anex Jose4
1Department of Chemistry, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, United States.
Journal of the American Chemical Society
|July 15, 2020
概括
研究人员在一个非同类蛋白质中设计了一个功能双核铜A (CuA) 中心,即细胞染色体c过氧化酶. 这表明保存的蛋白质折叠对于独特的金属蛋白质特性来说并不必不可少.
科学领域:
- 生物化学和生物物理
- 蛋白质工程
- 生物有机化学
背景情况:
- 金属蛋白功能受到金属结合部位协调球体的严重影响.
- 对于电子转移至关重要的双核铜A (CuA) 中心通常存在于铜素域中.
- 在人工系统中复制原生CuA属性是具有挑战性的,通常需要同类蛋白质支架.
研究的目的:
- 调查功能性CuA中心是否可以被设计成结构非同类蛋白质.
- 为了确定 CuA 独特的特性是否可以在不依赖于保存的铜素折叠的情况下实现.
- 描述细胞染色体c过氧化酶中新型CuA中心的光谱,电子和功能性质.
主要方法:
- 局部定向突变引入细胞染色体c过氧化酶 (Ccp) 的两种突变,从而产生CuA中心 (CuACcp).
- 光谱分析包括紫外线可见吸收,共振拉曼,磁圆二极化和X波段EPR (连续波和HYSCORE).
- 停止流动动学,X射线吸收细结构 (XAFS) 光谱学和残留突变研究以确定结构功能关系.
主要成果:
- 通过两种突变成功将一个完全功能化的CuA中心转化为Ccp (CuACcp).
- 光谱数据证实了价值移位,并显示了与原生CuA相似的EPR参数.
- CuACcp表现出与原生CuA相似的降解潜力,使电子转移到生理伙伴;结构分析确定了CuA结合和稳定的主要残留物.
结论:
- 保存的蛋白质折叠不是赋予金属氧化还原中心特征的先决条件.
- 蛋白质工程可以成功地将复杂的金属辅助因子引入非同类蛋白质支架.
- 这项工作扩大了设计具有定制功能的新型金属蛋白的可能性.
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