Elucidation of the Molecular Mechanisms of Photosystem II Complex Based on Its Crystal Structure Analysis

光系统II（PSII）是由14-17个跨膜亚基和3个膜周（外源）亚基组成的超分子膜蛋白复合物，总分子量为350 kDa。本研究旨在分析PSII的晶体结构，并在此基础上阐明PSII中发生的电子转移、水裂解和放氧反应的分子机理。为此，我们从嗜热蓝细菌Thermosynechococcus vulcanus中结晶出PSII复合物，并在3.7 nm分辨率下分析了其晶体结构，其中，我们根据在当前分辨率下可见的一些残基的大侧链的电子密度图，指定了PSII大亚基CP 47，CP 43，D1，D2的70-80%残基。我们构建了3种参与氧释放的外源蛋白的结构，其中12 kDa蛋白的结构为首次报道。整个结构还含有14个跨膜螺旋，其中一些被分配给一些低分子量亚基，包括细胞色素b559的α和β亚基，其他螺旋未被鉴定。在反应中心4叶绿素中，我们发现“特殊二聚体”PD 1-PD 2的距离比它们与两个“辅助叶绿素”的距离要短，这表明PSII反应中心不是一个同质的“四聚体”。在D_2和细胞色素b559之间，我们确定了两个β-胡萝卜素，从而暗示了PSII的二次电子传递途径是从细胞色素b559或ChlZD_2依次经过两个β-胡萝卜素，再到ChlD_2，PD_2，PD_1。我们还得到了Mn团簇的电子密度，它是一个Y形或“3+1”模型，如以前所报道的。我们进一步提高了PSII晶体的分辨率到3.5 μ m，修改了我们原来的PSII结构模型，并在修改后的结构的基础上更详细地分析了PSII的功能。

Photosystem II (PSII) is a supra-molecular membrane-protein complex consisting of 14-17 membrane-spanning subunits and 3 membrane-peripheral (extrinsic) subunits with a total molecular mass of 350 kDa. This research aimed to analyze the crystal structure of PSII and, on the basis of this, to elucidate the molecular mechanisms of electron transfer, water-splitting and oxygen-evolving reactions taken place in PSII. For this purpose, we crystallized the PSII complex from a thermophilic cyanobacterium Thermosynechococcus vulcanus and analyzed its crystal structure at 3.7Å resolution in which, we assigned 70-80% residues of PSII large subunits CP47,CP43,D1,D2 based on the electron density maps of some residue's large side chains which were visible at the current resolution. We built the structures of all the 3 extrinsic proteins involved in oxygen evolution, of which, the structure of 12 kDa protein was reported for the first time. The whole structure contained in addition 14 trans-membrane helices, some of which were assigned to some low-molecular mass subunits including the α and β-subunits of cytochrome b559,and other helices were not identified. In the reaction center 4 chlorophylls, we identified that the "special dimer" PD1-PD2 has a shorter distance than those between them and the two "accessory chlorophylls", suggesting that the PSII reaction center is not a homogenous "tetramer". We assigned two β-carotenes between the region of D2 and cytochrome b559, thus implied that the secondary electron transfer pathway in PSII is from cytochrome b559 or ChlZD_2 via the two β-carotenes in series and then to ChlD_2,PD_2,PD_1. We also obtained the electron density for the Mn-cluster which is a Y-shaped or "3+1" model as reported previously. We further improved the PSII crystal resolution to 3.5Å, modified our original PSII structure model, and analyzed the PSII functions in more details based on the modified structure.