REACTION CENTER DYNAMICS

反应中心动力学

• 批准号: 2185754
• 负责人: CRAIG C SCHENCK
• 金额: $ 17.29万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-04-01 至 1997-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2185754
• 关键词: Raman spectrometry; Rhodospirillales; X ray crystallography; bacterial pigment; bacterial proteins; bioenergetics; biophysics; chemical kinetics; chlorophyll; conformation; cryostat; electron spin resonance spectroscopy; electron transport; electronic spectra; fluorescence; fluorescence spectrometry; intermolecular interaction; laser spectrometry; mathematical model; mutant; photochemistry; photosynthetic bacteria; photosynthetic reaction centers; protein sequence; protein structure function; quinones; site directed mutagenesis; thermodynamics; time resolved data

The bacterial reaction center protein (RC) is an integral membrane protein that initiates the light-driven electron transfer reactions of photosynthesis. With the advent of a high-resolution crystal structure, the RC has emerged as a central biophysical paradigm for understanding structure/function relationships in proteins. A broad range of biochemical, spectroscopic, structural and theoretical techniques has been trained on the RC system, in an effort to answer a question of fundamental biological relevance: How does the RC separate electrical charges across a membrane bilayer with unitary quantum yield? The complete specification of the mechanism requires the study of a host of energy transfer, electron transfer, proton transfer and conformational equilibrium reactions. A multidisciplinary approach may yield the most meaningful insights. In this proposal, attention is focused on the earliest electron transfer reactions that occur on the femtosecond and picosecond time scales. The P.I. proposes to approach the following specific mechanistic questions: 1) What accounts for the unidirectionality of electron transfer in the context of a quasisymmetric protein structure? 2) How is the sequence of electron transfer events to be described? In particular, what is the role of Beta(L), the monomeric bacteriochlorophyll? What kinetic description of these events is appropriate: superexchange, two step, inhomogeneous distributions of RCs? Is the initially-prepared excited state vibrationally equilibrated prior to electron transfer? What other types of relaxation processes are important to the mechanism? 3) What are the molecular determinants of the ground state and excited state spectroscopic and redox properties? Does the protein play an active or passive role in the mechanism? 4) What theoretical description of these processes is appropriate? Is the nonadiabatic theory of electron transfer adequate or is some other formalism required? The principles that are to be elucidated by these studies may be generally applicable to other energy-transducing proteins. The P.I. proposes to use time-resolved and steady-state optical spectroscopic techniques to observe the formation and decay of intermediates in wild-type and selected mutant RCs. Time-resolved fluorescence and absorption experiment will be conducted over a wide- range of experimental conditions. Global kinetic analysis will be applied to the data in order to attempt an unbiased, possibly unique mechanistic interpretation. The P.I. will also make modifications to the mutagenesis system and select more RC mutants for physicochemical analysis. In collaboration with others, mutant structures will be determined by x-ray crystallography, ENDOR spectra will be obtained, and resonance Raman spectra will be measured.

细菌反应中心蛋白（RC）是一种积分膜 启动发光电子转移反应的蛋白质 光合作用。 随着高分辨率晶体结构的出现 RC已成为一种理解的中央生物物理范式 蛋白质中的结构/功能关系。 广泛的 生化，光谱，结构和理论技术具有 在RC系统上接受了培训，以回答 基本生物学相关性：RC如何分开电气 带有统一量子产率的膜双层的电荷？ 这 该机制的完整规范需要研究大量 能量传递，电子传递，质子转移和构象 平衡反应。 多学科的方法可能会产生最多的 有意义的见解。 在此提案中，注意力集中在最早的电子传输上 在飞秒和皮秒时间尺度上发生的反应。 这 P.I.建议解决以下特定机械问题： 1）哪些内容说明了电子传输的单向性 准对称蛋白质结构的背景？ 2）序列如何 要描述的电子传输事件？ 特别是什么是 Beta（L）的作用，单体细菌氯粘菌的作用？ 动力学 这些事件的描述适当：superexchange，两个步骤， RCS的不均匀分布？ 是最初准备的兴奋 电子传输之前的状态振动平衡？ 还有什么 放松过程的类型对机制很重要？ 3）什么 是基态和激发态的分子决定因素 光谱和氧化还原特性？ 蛋白质会发挥活性还是 在机制中被动角色？ 4）这些理论描述 过程合适吗？ 是电子的非绝热理论 转移适当的或需要其他形式主义？ 原则 这些研究通常适用于这些研究 到其他能量传递的蛋白质。 P.I.建议使用时间分辨和稳态光学 光谱技术观察的形成和衰变 野生型和选定突变体RC中的中间体。 时间分辨 荧光和吸收实验将在广泛的 实验条件范围。 全球动力学分析将是 应用于数据，以尝试无偏见的，可能是唯一的 机械解释。 P.I.还将对 诱变系统并选择更多的RC突变体进行物理化学 分析。 与他人合作，突变结构将是 通过X射线晶体学确定，将获得Endor光谱，并且 共振拉曼光谱将被测量。