Mechanism and Macromolecular Organization in Photosynthetic Reaction Centers

为了了解细菌光合作用反应中心的初始电荷分离步骤的机理，提出了几条研究路线。(I)测量特殊对最低Qy电子态的共振拉曼强度和激发轮廓，结合光化学烧孔、吸收和电吸收(斯塔克效应)光谱，以获得关于激发态动力学的信息；(Ii)在特殊对附近的特定位点突变体的制备和表征，其扰乱其氧化还原特性，以获得关于可能影响单向电子转移的局部介电特性的信息；以及(Iii)研究施主、受体和中间态氧化还原性质与野生型不同的RC突变体的量子产率和动力学，以确定快电荷分离和慢电荷复合反应的机理。光合作用反应中心是能够维持启动光合作用的光诱导电荷分离反应的最小的叶绿素-蛋白质复合体。第一个反应是已知的任何系统中最快的反应之一。除了这些反应作为地球上储存的化学能的主要来源的明显重要性外，这些反应还具有更广泛的生物物理和化学意义。这是因为电子转移反应在自然界中无处不在，而且由于这类反应相对简单，因此有希望能够在基础水平上了解反应机理。细菌反应中心是第一个用X射线结晶学表征的膜蛋白，因此我们的所有工作都归入结构-功能关系的总标题下。这项研究的重点是通过改变内部或外部电场来系统地改变反应中心内电荷转移反应的速率和机理。内部磁场可以通过使用重组DNA方法改变特定的氨基酸或氨基酸的延伸来改变。外场既可以用电极施加到大块样品上，也可以作为跨膜电位施加。后者在几乎所有的生物组装中都非常重要，我们试图了解将这种潜力与生物功能相结合的机制。

In order to understand the mechanism of the initial charge separation steps in bacterial photosynthetic reaction centers, several lines of investigation are proposed. (i) Measurements of the resonance Raman intensities and excitation profile for the lowest Qy electronic state of the special pair, combined with photochemical holeburning, absorption and electroabsorption (Stark effect) spectroscopy, to obtain information on excited state dynamics; (ii) preparation and characterization of site-specific mutants in the vicinity of the special pair which perturb its redox properties to obtain information on the local dielectric properties which may affect unidirectional electron transfer; and (iii) studies of the quantum yields and kinetics of RC mutants whose donor, acceptor and mediating state redox properties are different from wild-type in applied electric fields to determine the mechanism of the fast charge separation and slower charge recombination reactions. %%% Photosynthetic reaction centers are the smallest chlorophyll- protein complex capable of sustaining the light-induced charge separation reactions which initiate photosynthesis. The first reactions are among the fastest reactions known in any system. In addition to the obvious importance of these reactions as the primary source of stored chemical energy on earth, these reactions have much broader biophysical and chemical significance. This is because electron transfer reactions are ubiquitous in nature, and because of the relative simplicity of this type of reaction, there is some prospect of being able to understand the reaction mechanism at a fundamental level. Bacterial reaction centers were the first membrane protein to be characterized by x-ray crystallography, and consequently all of our work falls under the general heading of structure-function relationships. The proposed research focuses on systematically altering the rates and mechanisms of charge-transfer reactions in the reaction center by modifying either internal or external electric fields. Internal fields can be altered by changing specific amino acids or stretches of amino acids using recombinant DNA methods. External fields can be applied either with electrodes to a bulk sample or as transmembrane potentials. The latter are of very great importance in nearly all biological assemblies and we seek to understand the mechanisms which couple such potentials to biological function.