MS-based cross-linking analysis reveals the location of the PsbQ protein in cyanobacterial photosystem II
MS-based cross-linking analysis reveals the location of the PsbQ protein in cyanobacterial photosystem II
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DOI：
10.1073/pnas.1323063111
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发表时间：
2014-03-25
影响因子：
11.1
通讯作者：
Pakrasi, Himadri B.
中科院分区：
文献类型：
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作者：
Liu, Haijun;Zhang, Hao;Pakrasi, Himadri B.
PsbQ是植物、藻类和蓝藻中调节光系统II(PSII)水分解活性的外源蛋白质组分。然而，在目前嗜热蓝藻PSII的晶体结构中没有观察到PsbQ。因此，PsbQ在PSII复合体中的结构位置仍然未知。在这里，我们报道了从模式蓝藻聚球藻中分离出的二聚体PSII复合体的化学交联、免疫检测和液相色谱/串联MS分析。PCC 6803，以确定PsbQ在PSII中的结合部位。PsbQ的K-120与PsbO的K-180和K-59，以及PsbQ的K-102和CP47的D-440之间的交联表明，PsbQ与PsbO和CP47蛋白密切相关。我们进一步表明，psbO基因的遗传缺失导致PSII复合体中完全不存在PsbQ，以及PSII的二聚体形式的丧失。总体而言，我们的数据提供了蓝藻PSII中PsbQ神秘结合部位的分子水平描述。这些结果也有助于我们理解PsbQ蛋白在PSII组装过程中的顺序掺入，以及它对PSII放氧活性的稳定作用。
PsbQ is a luminal extrinsic protein component that regulates the water splitting activity of photosystem II (PSII) in plants, algae, and cyanobacteria. However, PsbQ is not observed in the currently available crystal structures of PSII from thermophilic cyanobacteria. The structural location of PsbQ within the PSII complex has therefore remained unknown. Here, we report chemical cross-linking followed by immunodetection and liquid chromatography/tandem MS analysis of a dimeric PSII complex isolated from the model cyanobacterium, Synechocystis sp. PCC 6803, to determine the binding site of PsbQ within PSII. Our results demonstrate that PsbQ is closely associated with the PsbO and CP47 proteins, as revealed by cross-links detected between K-120 of PsbQ and K-180 and K-59 of PsbO, and between K-102 of PsbQ and D-440 of CP47. We further show that genetic deletion of the psbO gene results in the complete absence of PsbQ in PSII complexes as well as the loss of the dimeric form of PSII. Overall, our data provide a molecular-level description of the enigmatic binding site of PsbQ in PSII in a cyanobacterium. These results also help us understand the sequential incorporation of the PsbQ protein during the PSII assembly process, as well as its stabilizing effect on the oxygen evolution activity of PSII.