Quantifying the ion selectivity of the Ca2+ site in photosystem II: evidence for direct involvement of Ca2+ in O2 formation.
Quantifying the ion selectivity of the Ca2+ site in photosystem II: evidence for direct involvement of Ca2+ in O2 formation.
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DOI：
10.1021/bi010679z
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发表时间：
2001-06
期刊：
影响因子：
2.9
通讯作者：
J. Vrettos;Daniel A. Stone;G. Brudvig
中科院分区：
文献类型：
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作者：
J. Vrettos;Daniel A. Stone;G. Brudvig
钙是光系统 II (PSII) 的释氧复合物 (OEC) 中的重要辅助因子。去除 Ca2+ 或用除 Sr2+ 之外的任何金属离子取代它都会抑制氧气的析出。我们使用稳态酶动力学来测量经一系列单价、二价和三价金属离子处理的 PSII 样品中的 O2 释放速率，以确定金属离子对 Ca2+ 结合位点的亲和力的基础。我们的结果表明，PSII 中的 Ca2+ 结合位点的行为与其他蛋白质中的 Ca2+ 结合位点非常相似，我们讨论了这对 PSII 中该位点结构的影响。作为时间函数的活性测量表明，对于所有研究的 PSII 样品，结合位点在 4 小时内达到平衡。金属离子的结合亲和力由 17 和 23 kDa 外源多肽调节；它们的去除使金属离子的结合自由能降低了 2.5 kcal/mol，但不会显着改变达到平衡所需的时间。单价离子被有效地排除在 Ca2+ 结合位点之外，不会抑制 O2 的释放。离子半径与 Ca2+ (0.99 A) 相似的二价和三价金属离子与 Ca2+ 竞争性结合，并具有最高的结合亲和力，而较小的金属离子结合较弱，较大的金属离子不竞争性结合。这与具有刚性配体阵列的尺寸选择性 Ca2+ 结合位点一致。尽管有大量金属离子竞争性地取代 OEC 中的 Ca2+，但只有 Sr2+ 能够部分恢复活性。通过比较所研究的金属离子的物理特性，我们确定水离子的 pK(a) 是决定金属离子功能能力的因素。这表明 Ca2+ 直接参与水氧化的化学过程，而不仅仅是 OEC 中的结构辅助因子。我们认为 Ca2+ 的作用是充当路易斯酸，结合底物水分子并调节其反应性。
Calcium is an essential cofactor in the oxygen-evolving complex (OEC) of photosystem II (PSII). The removal of Ca2+ or its substitution by any metal ion except Sr2+ inhibits oxygen evolution. We used steady-state enzyme kinetics to measure the rate of O2 evolution in PSII samples treated with an extensive series of mono-, di-, and trivalent metal ions in order to determine the basis for the affinity of metal ions for the Ca2+-binding site. Our results show that the Ca2+-binding site in PSII behaves very similarly to the Ca2+-binding sites in other proteins, and we discuss the implications this has for the structure of the site in PSII. Activity measurements as a function of time show that the binding site achieves equilibrium in 4 h for all of the PSII samples investigated. The binding affinities of the metal ions are modulated by the 17 and 23 kDa extrinsic polypeptides; their removal decreases the free energy of binding of the metal ions by 2.5 kcal/mol, but does not significantly change the time required to reach equilibrium. Monovalent ions are effectively excluded from the Ca2+-binding site, exhibiting no inhibition of O2 evolution. Di- and trivalent metal ions with ionic radii similar to that of Ca2+ (0.99 A) bind competitively with Ca2+ and have the highest binding affinity, while smaller metal ions bind more weakly and much larger ones do not bind competitively. This is consistent with a size-selective Ca2+-binding site that has a rigid array of coordinating ligands. Despite the large number of metal ions that competitively replace Ca2+ in the OEC, only Sr2+ is capable of partially restoring activity. Comparing the physical characteristics of the metal ions studied, we identify the pK(a) of the aqua ion as the factor that determines the functional competence of the metal ion. This suggests that Ca2+ is directly involved in the chemistry of water oxidation and is not only a structural cofactor in the OEC. We propose that the role of Ca2+ is to act as a Lewis acid, binding a substrate water molecule and tuning its reactivity.