Electron Transfer in the Cytochrome c - Peroxidases Complex

细胞色素c-细胞色素c过氧化物酶（CcP）系统提供了一种 非常丰富的机会来调查的亲密细节 蛋白质相互作用和长程电子转移 生理上相关的血红素蛋白之间的联系。 的目标 这项研究建议包括两个方面。 第一个领域是 描述马和酵母iso-1之间的相互作用 细胞色素c与酵母CcP。 实验旨在阐明 细胞色素c和CcP之间结合的化学计量将是 执行以确定是否以及在何种条件下， 细胞色素c可以与CcP结合。高亲和力细胞色素c CcP上的结合位点将通过定点 诱变 单个氨基酸在结合中的作用 现场将通过热力学测量进行评估 结合亲和力 任何次级相互作用点的位置 也将通过定点诱变来确定。 的 第二个感兴趣的领域是表征电子转移 在细胞色素c和CcP的血红素基团之间。 瞬态 动力学技术将被用来研究电子的速率 从亚铁细胞色素c转移到CcP的氧化中间体 以及血红素铁和色氨酸之间的氧化还原平衡速率， 191在CcP化合物II中。 反应动力学的特异性 电子传递途径将通过位点- CcP的定向诱变。 %%% 这项工作将有助于更好地理解蛋白质-蛋白质 识别一般，和血红素蛋白，其中包括 血红蛋白和能量代谢中的蛋白质，特别是

The cytochrome c-cytochrome c peroxidase (CcP) system provides an exceedingly rich opportunity to investigate the intimate details of protein-protein interactions and long-range electron transfer between physiologically related heme proteins. The objectives of this research proposal lie in two areas. This first area is to characterize the interaction between both horse and yeast iso-1 cytochrome c with yeast CcP. Experiments designed to clarify the stoichiometry of binding between cytochrome c and CcP will be performed to determine if, and under what conditions, more than one cytochrome c may bind to CcP. The high-affinity cytochrome c binding site on CcP will be mapped through site-directed mutagenesis. The role of individual amino acids at the binding site will be assessed through thermodynamic measurements of the binding affinity. The location of any secondary interaction sites will also be determined through site-directed mutagenesis. The second area of interest is to characterize electron transfer between the heme groups of cytochrome c and CcP. Transient-state kinetic techniques will be used to investigate the rate of electron transfer from ferrocytochrome c to the oxidized intermediate of CcP and the rate of redox equilibration between the heme iron and Trp- 191 in CcP compound II. The specificity of the reaction dynamics and the electron transfer pathway will be explored through site- directed mutagenesis of CcP. %%% This work should lead to a better understanding of protein-protein recognition in general, and of heme proteins, which include hemoglobin and the proteins in energy metabolism, in particular