Aydin A Orkun, de Lichtenberg Casper, Liang Feiyan, Forsman Jack, Graça André T, Chernev Petko, Zhu Shaochun, Mateus André, Magnuson Ann, Cheah Mun Hon, Schröder Wolfgang P, Ho Felix, Lindblad Peter, Debus Richard J, Mamedov Fikret, Messinger Johannes
Molecular Biomimetics, Department of Chemistry- Ångström, Uppsala University, Uppsala, 751 20, Sweden.
Department of Plant and Environmental Sciences, University of Copenhagen, Frederiksberg C, 1871, Denmark.
Photosynth Res. 2025 Apr 22;163(3):28. doi: 10.1007/s11120-025-01147-4.
Light-driven water oxidation by photosystem II sustains life on Earth by providing the electrons and protons for the reduction of CO to carbohydrates and the molecular oxygen we breathe. The inorganic core of the oxygen evolving complex is made of the earth-abundant elements manganese, calcium and oxygen (MnCaO cluster), and is situated in a binding pocket that is connected to the aqueous surrounding via water-filled channels that allow water intake and proton egress. Recent serial crystallography and infrared spectroscopy studies performed with PSII isolated from Thermosynechococcus vestitus (T. vestitus) support that one of these channels, the O1 channel, facilitates water access to the MnCaO cluster during its S→S and S→S→S state transitions, while a subsequent CryoEM study concluded that this channel is blocked in the cyanobacterium Synechocystis sp. PCC 6803, questioning the role of the O1 channel in water delivery. Employing site-directed mutagenesis we modified the two O1 channel bottleneck residues D1-E329 and CP43-V410 (T. vestitus numbering) and probed water access and substrate exchange via time resolved membrane inlet mass spectrometry. Our data demonstrates that water reaches the MnCaO cluster via the O1 channel in both wildtype and mutant PSII. In addition, the detailed analysis provides functional insight into the intricate protein-water-cofactor network near the MnCaO cluster that includes the pentameric, near planar 'water wheel' of the O1 channel.
光系统II驱动的水氧化为地球上的生命提供了电子和质子，用于将二氧化碳还原为碳水化合物以及我们呼吸所需的分子氧。放氧复合体的无机核心由地球上储量丰富的元素锰、钙和氧（MnCaO簇）组成，位于一个结合口袋中，该口袋通过充满水的通道与周围的水环境相连，这些通道允许水进入并排出质子。最近对从嗜热栖热菌（T. vestitus）分离出的光系统II进行的系列晶体学和红外光谱研究表明，这些通道之一，即O1通道，在其S→S和S→S→S状态转变过程中促进水进入MnCaO簇，而随后的冷冻电镜研究得出结论，在集胞藻PCC 6803中该通道是堵塞的，这对O1通道在水输送中的作用提出了质疑。我们利用定点诱变技术对两个O1通道瓶颈残基D1-E329和CP43-V410（按T. vestitus编号）进行了修饰，并通过时间分辨膜进样质谱法探测了水的进入和底物交换。我们的数据表明，在野生型和突变型光系统II中，水都通过O1通道到达MnCaO簇。此外，详细分析为MnCaO簇附近复杂的蛋白质-水-辅因子网络提供了功能见解，该网络包括O1通道的五聚体近平面“水轮”。
