van de Ven M, Preston C, Seibert M, Gratton E
Department of Physiology and Biophysics, University of Illinois, Urbana.
Biochim Biophys Acta. 1990 Feb 2;1015(2):173-9. doi: 10.1016/0005-2728(90)90017-x.
We have measured the decay of chlorophyll a fluorescence at 4 degrees C under anaerobic conditions in stabilized photosystem II reaction center complex isolated from spinach, using multifrequency (2-400 MHz) cross-correlation phase fluorometry. Examination of our data shows that although the fluorescence decay of open reaction centers (i.e., when both the electron donor P-680 and the electron acceptor pheophytin are capable of engaging in charge separation) can be analyzed as a multiexponential decay, another representation of the data is obtained when the decay is analyzed using a continuous distribution of lifetimes. Our results on the open reaction center differ from the two lifetime components of 25 ps and 35 ns published by Mimuro et al. (Biochim. Biophys. Acta 933 (1988) 478-486) for the D1-D2-cytochrome b-559 complex, obtained for F682 at 4 degrees C by a time-resolved photon-counting spectrofluorometer. When the reaction centers are closed by pretreatment with sodium dithionite and methyl viologen followed by exposure to laser excitation, conditions known to result in accumulation of reduced pheophytin, a dramatic decrease in the contribution of the slow lifetime component(s) is observed. These results suggest that the slow distribution lifetime component(s) in the 5-20 ns range originate(s) in the back reaction of the charge separated state. On the other hand, the fast lifetime component(s) in the picosecond range may be only partially related to the charge separation, since no dramatic change is observed upon closure of the reaction center. Perhaps, this component is related, in part, to the excitation energy migration among the various chromophores in the reaction center preparations.
我们使用多频（2 - 400 MHz）互相关相位荧光测定法，测量了在厌氧条件下，从菠菜中分离得到的稳定化光系统II反应中心复合物在4摄氏度时叶绿素a荧光的衰减情况。对我们数据的研究表明，虽然开放反应中心（即电子供体P - 680和电子受体脱镁叶绿素均能够参与电荷分离时）的荧光衰减可以被分析为多指数衰减，但当使用连续寿命分布来分析衰减时，会得到数据的另一种表示形式。我们关于开放反应中心的结果与Mimuro等人（《生物化学与生物物理学报》933 (1988) 478 - 486）发表的关于D1 - D2 - 细胞色素b - 559复合物在4摄氏度下通过时间分辨光子计数分光荧光计测得的F682的两个寿命成分（25皮秒和35纳秒）不同。当反应中心通过连二亚硫酸钠和甲基紫精预处理然后暴露于激光激发而关闭时（已知这些条件会导致还原态脱镁叶绿素的积累），观察到慢寿命成分的贡献显著降低。这些结果表明，5 - 20纳秒范围内的慢分布寿命成分源于电荷分离态的反向反应。另一方面，皮秒范围内的快寿命成分可能仅部分与电荷分离有关，因为在反应中心关闭时未观察到显著变化。也许，该成分部分与反应中心制剂中各种发色团之间的激发能量迁移有关。
