Turconi S, MacLachlan D J, Bratt P J, Nugent J H, Evans M C
Department of Biology, University College London, U.K.
Biochemistry. 1997 Jan 28;36(4):879-85. doi: 10.1021/bi962010b.
The association of water with the Mn of the water oxidizing complex was investigated using H2(17)O- and 2H2O-reconstituted lyophilized photosystem II particles. The pulsed electron paramagnetic resonance (EPR) technique of electron spin echo envelope modulation (ESEEM) was used to investigate the interaction of the magnetic 2H and 17O nuclei with the paramagnetic S2 state of the Mn complex and other photosystem II components. ESEEM offers a much more specific and sensitive detection of this type of interaction than continuous wave (CW) EPR. Unlike earlier reports using CW EPR, these experiments did not detect any interaction of water with the multiline EPR signal from the S2 state of the Mn complex. No signals indicating specific interaction of either H or O with the multiline signal were detected. Signals due to 2H and 17O were detected only at the Larmour frequency, indicating nonspecific "distant ENDOR" effects. A weak interaction with 17O was detected both in S1, when the Mn is EPR silent, and in S2, but only on the high-field side of g = 2. This interaction may be with the Rieske iron-sulfur center in the cytochrome b6f complex. The results were the same whether the multiline signal was generated by 200 K illumination of dark-frozen samples, or by room temperature illumination in the presence of the inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU). Illumination at room temperature in the presence of an electron acceptor to allow multiple turnovers of the system with cycling of the S states did not result in the appearance of any new interactions. These results appear to exclude close (less than 6 A) binding of water to the Mn center giving rise to the multiline signal, and also to exclude mechanisms in which water oxidation involves the breaking and re-formation of the mu-oxo bridges of the Mn complex. They cannot, however, exclude models in which water binding to the manganese complex and direct oxidation by the manganese complex occur in the higher S states, or are catalyzed by one bis(mu-oxo) Mn dimer while oxidizing equivalents are accumulated in the S2 state by a second bis(mu-oxo) Mn dimer.
利用H₂¹⁷O和²H₂O重构的冻干光系统II颗粒，研究了水与水氧化复合物中锰的关联。采用电子自旋回波包络调制（ESEEM）的脉冲电子顺磁共振（EPR）技术，研究了磁性²H和¹⁷O核与锰复合物顺磁S₂态及其他光系统II组分之间的相互作用。与连续波（CW）EPR相比，ESEEM对这类相互作用的检测更具特异性和敏感性。与早期使用CW EPR的报道不同，这些实验未检测到水与锰复合物S₂态的多线EPR信号有任何相互作用。未检测到表明H或O与多线信号存在特异性相互作用的信号。仅在拉莫尔频率处检测到了由²H和¹⁷O产生的信号，表明存在非特异性的“远程ENDOR”效应。在S₁态（此时锰的EPR信号沉默）和S₂态均检测到了与¹⁷O的弱相互作用，但仅在g = 2的高场侧。这种相互作用可能是与细胞色素b₆f复合物中的 Rieske铁硫中心发生的。无论多线信号是由暗冻样品在200 K光照下产生，还是在存在抑制剂3-(3,4-二氯苯基)-1,1-二甲基脲（DCMU）的情况下室温光照产生，结果都是相同的。在存在电子受体的情况下室温光照，以使系统通过S态循环进行多次周转，并未导致出现任何新的相互作用。这些结果似乎排除了水与产生多线信号的锰中心紧密（小于6 Å）结合的情况，也排除了水氧化涉及锰复合物μ-氧桥断裂和重新形成的机制。然而，它们不能排除在较高S态下水与锰复合物结合并由锰复合物直接氧化的模型，或者在一个双（μ-氧）锰二聚体催化水氧化的同时，另一个双（μ-氧）锰二聚体在S₂态积累氧化当量的模型。
