Electron Transfer within Protein Complexes

描述（由申请人提供）：蛋白质-蛋白质电子转移（ET）在生物学和化学中占据中心地位。除此之外，蛋白质-蛋白质识别和对接的潜在现象，几乎是所有生物过程的核心，特别适合通过测量蛋白质间ET来研究，其对距离/途径的高度依赖，在存在更多非反应性结构的情况下，作为探测反应性结构的独特“过滤器”。我们的研究结果表明：(a)蛋白质-蛋白质界面上的ET通常由具有不同界面结构和表面水化程度的状态集合之间和内部的构象转换调节，(b)构象移动系统中的ET光周期提供了可以测量这种动态过程的“时钟”，以及(c)光周期时钟可用于表征从皮秒到秒的时间尺度上构象耦合的功能。了解蛋白质-蛋白质相互作用和蛋白质间ET的目标使我们最近非常成功地通过重新设计对接界面的电荷反转策略来控制它们，其中一个结果是在ps-ns时间尺度上发现了蛋白质间单线态ET光循环。我们建议：(i)改进接口重新设计策略，将其应用于新的合作伙伴，并使用它来发现具有单线态ET光循环的新系统；（ii）通过一个紧密集成的程序来检查构象调制的ET，该程序由ET随溶剂变化的动力学测量、动态配合物的结构表征（NMR）和计算（布朗动力学/MD/斯摩鲁乔斯基方程）方法组成。要研究的三个系统涉及不同的界面：肌红蛋白（Mb），蛋白质重新设计的载体，主要通过静电相互作用与电子受体伴侣蛋白结合；结构表征的细胞色素c过氧化物酶（CcP）和细胞色素c （Cc）之间的复合物主要通过疏水相互作用结合；[锌;Fe]血红蛋白（Hb）杂化体在ET-伙伴链之间的界面上表现出混合的相互作用。我们将验证这样一个假设，即每个体系都表现出三种类型的ET构象控制的不同组合：动态过程、不同表面水化程度的构象之间的转换和/或体积变化。我们还建议将从这些研究中得到的理解“翻译”到核苷酸调节的问题，构象连接的ET在氮酶的催化作用中。

DESCRIPTION (provided by applicant): Protein-protein electron transfer (ET) occupies a central place in biology and chemistry. Beyond this, the underlying phenomena of protein-protein recognition and docking, which are central to almost all biological processes, are particularly amenable to study through measurements of inter-protein ET, whose steep dependence on distance/pathways acts as a unique 'filter' for probing reactive configurations in the presence of more numerous non-reactive ones. Our results indicate that (a) ET across a protein-protein interface typically is modulated by conformational conversion between and within ensembles of states with different interface structures and degrees of surface hydration, (b) ET photocycles in conformationally mobile systems provide 'clocks' against which such dynamic processes can be measured, and (c) the photocycle clocks can be used to characterize the coupling of conformation to function over timescales from picoseconds to seconds. The goal of understanding protein-protein interactions and interprotein ET has led us to recent, highly successful efforts at their control through a charge-reversal strategy for redesign of the docking interface, one result being the discovery of an interprotein singlet ET photocycle on ps-ns timescales. We propose: (i) to refine the interface redesign strategy, to apply it to new partners, and to use it to discover new systems with singlet ET photocycles; (ii) to examine conformationally-modulated ET through a tightly integrated program comprised of kinetic measurements of the variations of ET with solvent, structural characterization of dynamic complexes (NMR), and computational (Brownian Dynamics/MD/Smoluchowski equation) approaches. The three systems to be studied involve distinct interfaces: myoglobin (Mb), the vehicle for protein redesign efforts, binds to electron acceptor partner proteins primarily through electrostatic interactions; the structurally characterized complex between cytochrome c peroxidase (CcP) and cytochrome c (Cc) is primarily bound by hydrophobic interactions; [Zn; Fe] hemoglobin (Hb) hybrids exhibit a mixture of interactions at the interface between the ET- partner chains. We will test the hypothesis that, as a result, each system exhibits a distinct combination of three types of conformational control of ET: dynamic processes, conversion among conformations with different degrees of surface hydration, and/or changes in volume. We also propose to 'translate' the understanding derived from these studies to the problem of nucleotide-regulated, conformationally-linked ET in catalysis by nitrogenase. PUBLIC HEALTH RELEVANCE: Electron transfer between proteins is vital to the physiological processes of respiration and metabolism. Beyond this, the underlying phenomena of protein-protein recognition and docking are central to almost all biological processes that underlie human physiology and health. Our research program addresses fundamental questions of interprotein electron transfer, how it is modulated by atomic motions at the protein-protein interface, and how it might be controlled by protein redesign. It translates these findings into the realm of catalysis by the enzyme, nitrogenase.