Pulsed EPR Studies of Photosystem II Radicals

技术：利用ESEEM和ESE-ENDOR的脉冲电子顺磁共振（EPR）方法以及选择性同位素标记和氨基酸诱变来研究光系统II中各种自由基中心的功能。PSII与紫色无氧细菌反应中心在p650和QA、QB结合位点的同源程度将被研究。时间分辨脉冲EPR光谱将用于探测PSII电子转移辅助因子的相对几何形状以及电子和质子转移耦合的细节。脉冲EPR实验将有助于确定PSII酪氨酸自由基Yz和YD的蛋白质环境差异，以了解这两种酪氨酸中心的功能差异。其他实验将用于更好地描述锰簇和Yz之间的详细磁性和结构相互作用。时间分辨EPR将用于测量Yz的动力学，并通过Yz寻找质子/电子转移耦合的直接证据。快速冷冻淬火实验的目标是捕获自由基YZ。物种在02形成和释放之前。进一步的实验将探讨光氧化叶绿素Chlz在光系统II光保护中的作用。非技术：磁共振实验计划提供新的见解，以了解植物光系统II反应中心的氨基酸和辅因子自由基如何被利用来分裂水，还原醌，并将质子注入被光合膜包围的水相，所有这些都具有高量子产率，并由低能量可见光驱动。光合作用过程是光驱动的优势允许特定的自由基捕获和瞬态动力学实验，这在其他使用自由基中心的生物系统中是困难或不可能的。因此，这些光系统II实验将提供对各种生化自由基机制的见解，这些机制将在更广泛的科学领域产生影响。

Britt9874541Technical: Experiments utilizing the pulsed electron paramagnetic resonance (EPR) methods of ESEEM and ESE-ENDOR along with selective isotopic labelling and mutagenesis of amino acids will be used to investigate the function of various radical centers in Photosystem II. The degree of homology between PSII and purple non-oxygen evolving bacterial reaction centers will be investigated at P6s0 and the QA and QB binding sites. Time resolved pulsed EPR spectroscopy will be utilized to probe the relative geometry of the PSII electron transfer cofactors and details of the coupling of electron and proton transfer. Pulsed EPR experiments will help define the differences between the protein environments of PSII tyrosine radicals Yz and YD, with an aim of understanding the functional differences between these two tyrosine centers. Other experiments will be used to better describe the detailed magnetic and structural interactions between the manganese cluster and Yz. Time resolved EPR will be used to measure the dynamics of Yz, and to search for direct evidence of proton/electron transfer coupling through Yz. Rapid freeze quench experiments will target trapping of the radical YZ. species immediately before 02 formation and release. Further experiments will probe the role of a photoxidizeable chlorophyll, Chlz, in Photosystem II photoprotection.Nontechnical: Magnetic resonance experiments are planned to provide new insights into how amino acid and cofactor radical species of the plant Photosystem II reaction center are utilized to split water, reduce quinones, and inject protons into the aqueous phase enclosed by the photosynthetic membrane, all with high quantum yield, and driven by low energy visible light. The advantage that photosynthetic processes are light driven allows specific radical trapping and transient kinetic experiments that would be difficult or impossible in other biological systems employing radical centers. Therefore these Photosystem II experiments will provide insights into diverse biochemical radical mechanisms that will have impact in the broader scientific arena.