Studies of the Primary Reactions of Bacterial Photosynthesis

伍德伯里博士建议研究细菌光合反应中心初始电子转移的机制。特别感兴趣的是进一步了解蛋白质基质在调节光合电子转移的热力学和动力学参数方面所起的作用。为此，伍德伯里博士将利用定向诱变的组合来改变反应中心辅助因子附近的氨基酸和瞬态光谱来分析氧化还原活性辅助因子之间电子转移的动力学和热力学，并确定参与该反应的机械上重要的中间体。提出了几个具体项目。其中一个涉及到使用位点定向诱变来研究氢键对初始电子供体氧化还原性能的影响，然后确定改变这一重要热力学参数对电子转移机制的影响。Woodbury博士将在这个项目中执行大部分的快速光谱；突变体是由艾伦和威廉姆斯设计和制造的。另一个项目是通过在不同生物体的反应中心基因之间交换DNA序列区域来比较不同生物体的反应中心的电子转移机制。在蛋白主链水平上，反应中心具有接近两折旋转的对称性。然而，存在于两边的氨基酸有很大的不同。结果是电子转移是非常不对称的，几乎只发生在两种可能的途径中的一种，即使两种途径存在化学上相同的辅因子。这将通过交换双方之间的区域，并进行方向，动力学和初始电子转移的中间状态的光谱分析来研究。这个项目有两个更大的目标。首先，了解光合作用过程中的太阳能转换本身就是本实验室的主要兴趣所在。从本质上讲，生物体使用的所有能量最终都来自这个来源，对这个过程的机械理解对于开发需要特定能量输入的分子装置（这就是反应中心的真正含义）非常有用。其次，反应中心为辅助因子-蛋白质相互作用的研究提供了一个很好的实验室。很少有蛋白质-发色团系统可以在短至飞秒（包括分子运动的时间尺度）的时间尺度上对反应机制进行如此详细的探索。在这里，从这些研究中获得的信息可以用于合理设计许多类型的分子装置，并用于理解非常广泛的蛋白质介导的化学反应。

Dr. Woodbury proposes to investigate the mechanism of initial electron transfer in bacterial photosynthetic reaction centers. Particular interests lie in furthering the understanding of the role that the protein matrix plays in modulating the thermodynamic and kinetic parameters of photosynthetic electron transfer. To this end, Dr. Woodbury will utilize a combination of directed mutagenesis to alter the amino acids near the reaction center cofactors and transient spectroscopy to assay the kinetics and thermodynamics of electron transfer between redox active cofactors as well as to identify the mechanistically important intermediates involved in this reaction. Several specific projects are proposed. One involves the use of site-directed mutagenesis to investigate the effect of hydrogen bonding on the redox properties of the initial electron donor and then to determine the effect of changing this important thermodynamic parameter on the electron transfer mechanism. Dr. Woodbury will perform most of the fast spectroscopy in this project; the mutants were designed and constructed by Allen and Williams. Another project involves comparing the electron transfer mechanisms of reaction centers from different organisms by exchanging regions of DNA sequence between the reaction center genes of these organisms. At the level of the protein backbone, the reaction center has a near two fold-rotational symmetry. However, the amino acids present on the two sides are largely different. The result is that electron transfer is very asymmetric, occurring almost exclusively down one of the two possible pathways, even though two pathways exist with chemically identical cofactors. This will be studied by exchanging regions between the two sides and performing spectroscopic analysis of the direction, kinetics and intermediate states involved in initial electron transfer. %%% There are two broader goals of the project. First, an understanding of solar energy conversion during photosynthesis is itself a major interest of this laboratory. Essentially all of the energy used by living organisms ultimately comes from this source, and a mechanistic understanding of this process could be very useful in the development of molecular devices that require a specific energy input (which is what a reaction center really is). Second, the reaction center provides an excellent laboratory for the study of cofactor-protein interactions in general. There are few protein-chromophore systems where reaction mechanism can be probed in such detail on time scales as short as femtoseconds (which encompasses the time scale of molecular motion). Here again, the information obtained from these studies can be used in the rational design of many types of molecular devices and in the understanding of a very wide range of protein-mediated chemical reactions.