Psb27, a transiently associated protein, binds to the chlorophyll binding protein CP43 in photosystem II assembly intermediates
Psb27, a transiently associated protein, binds to the chlorophyll binding protein CP43 in photosystem II assembly intermediates
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DOI：
10.1073/pnas.1111597108
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发表时间：
2011-11-08
影响因子：
11.1
通讯作者：
Pakrasi, Himadri B.
中科院分区：
文献类型：
--
作者：
Liu, Haijun;Huang, Richard Y. -C.;Pakrasi, Himadri B.
光系统II（PSII）是位于蓝藻和叶绿体类囊体膜上的一个大型多亚基色素蛋白复合物，介导光驱动的氧气从水中的演化。最近，一个高分辨率的X射线结构的成熟PSII复杂的已成为可用。两个PSII多肽，D1和CP 43，提供了许多的配体的无机Mn 4Ca中心，这是必不可少的水氧化。由于其不寻常的氧化还原化学，PSII经常经历降解，然后逐步组装。Psb 27是一种小的腔多肽，在这个复杂的组装途径中充当重要的辅助因子。然而，PSII组装中间体中Psb 27的结构位置仍然难以捉摸。在这里，我们报告说，Psb 27结合CP 43在这样的组装中间体。我们处理纯化的基因标记的PSII组装中间复合物从蓝细菌集胞藻6803与化学交联剂研究Psb 27和各种PSII蛋白之间的分子间相互作用。首先，使用水溶性1-乙基-3-（3-二甲基氨基丙基）碳二亚胺（EDC）交联具有彼此紧密缔合的互补带电基团的蛋白质。在His 27 Delta ctpAPS II制备物中，鉴定了含有Psb 27和CP 43的58 kDa交联物质。该物质在不存在Psb 27的HT 3 Delta ctpA Delta psb 27 PSII复合物中不形成。第二，同型双官能硫醇可裂解交联剂3，3 '-二硫代双（磺基琥珀酰亚胺基丙酸酯）（DTSSP）用于在His 27 Δ ctpAPS II制剂中将Psb 27可逆地交联至CP 43，其允许使用液相色谱/串联MS将交联位点定位为Psb 27 K（63）CP 43 D（321）（胰蛋白酶）和CP 43 K（215）Psb 27 D（58）AGGLK（63）CP 43 D（321）（胰凝乳蛋白酶）。我们的数据表明，Psb 27作为一个重要的调节蛋白在PSII组装通过特定的相互作用与CP 43的管腔域。
Photosystem II (PSII), a large multisubunit pigment-protein complex localized in the thylakoid membrane of cyanobacteria and chloroplasts, mediates light-driven evolution of oxygen from water. Recently, a high-resolution X-ray structure of the mature PSII complex has become available. Two PSII polypeptides, D1 and CP43, provide many of the ligands to an inorganic Mn4Ca center that is essential for water oxidation. Because of its unusual redox chemistry, PSII often undergoes degradation followed by stepwise assembly. Psb27, a small luminal polypeptide, functions as an important accessory factor in this elaborate assembly pathway. However, the structural location of Psb27 within PSII assembly intermediates has remained elusive. Here we report that Psb27 binds to CP43 in such assembly intermediates. We treated purified genetically tagged PSII assembly intermediate complexes from the cyanobacterium Synechocystis 6803 with chemical cross-linkers to examine intermolecular interactions between Psb27 and various PSII proteins. First, the water-soluble 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) was used to cross-link proteins with complementary charged groups in close association to one another. In the His27 Delta ctpAPSII preparation, a 58kDa cross-linked species containing Psb27 and CP43 was identified. This species was not formed in the HT3 Delta ctpA Delta psb27PSII complex in which Psb27 was absent. Second, the homobifunctional thiol-cleavable cross-linker 3,3'-dithiobis(sulfosuccinimidylpropionate) (DTSSP) was used to reversibly cross-link Psb27 to CP43 in His27 Delta ctpAPSII preparations, which allowed the use of liquid chromatography/tandem MS to map the cross-linking sites as Psb27K(63) CP43D(321) (trypsin) and CP43K(215) Psb27D(58)AGGLK(63) CP43D(321) (chymotrypsin), respectively. Our data suggest that Psb27 acts as an important regulatory protein during PSII assembly through specific interactions with the luminal domain of CP43.