Stabilization of the Mn cluster by Y_D tyrosine in oxygen evolving complex of photosystem II

Y_D 酪氨酸对光系统 II 放氧复合物中 Mn 簇的稳定性

• 批准号: 13640659
• 负责人: ONO Takaaki
• 金额: $ 1.28万
• 依托单位: The Institute of Physical and Chemical Research
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13640659/
• 关键词: photosynthesis; photosystem II; oxygen evolution; tyrosine D; manganese cluster; Chlamydomonas; D2 protein; green alga; 光化学系; 酸素電極; クラミドモナス; Y_Dチロシン; FTIR; ESR; 部位特異変異; TDチロシン

Function of a redox active Y_D tyrosine residue in photosystem II has been studied by the use of the unicellular green alga Chlamydomonas reinhardtii, in which the tyrosine for Y_D (the 160th tyrosine of the D2 protein) was replaced by phenylalanine by site-directed mutagenesis using His-tagged psbD plasmid. The processes of S state transition in photosynthetic oxygen evolution was monitored by analyzing the flash-induced oxygen evolution using a home-made high sensitive Joliot-type O_2 electrode, which enable us to measure the flash O_2 pattern of cultured algal cells directly without further condensation. In the mutant cells, no fast component was detected in the dark decay from O_2 to S_1 but slow decaying component was normal as compared with the wild-type cells, indicating the direct involvement of Y_D tyrosine in the fast decaying process. The flash O_2 pattern did not change even after prolonged dark incubation, indicating that Y_D does not contribute the redox equilibration between S_1 and S_0. Therefore, higher stability of S_1 than S_0 is predominantly ascribed to their thermodynamic natures.Furthermore, the absence of Y_D tyrosine did not affect the O_2 capability after long dark incubation. The results show that Y_D is not involved in the stability of The Mn cluster in the dark. In fact, PS II cores with high O_2 activity were prepared from the mutants by means of Ni-affinity column chromatography. This preparation is useful for the physicochemical studies, including ESR and FTIR, on photosystem II.

利用单细胞绿藻莱茵衣藻研究了光系统 II 中氧化还原活性 Y_D 酪氨酸残基的功能，其中 Y_D 的酪氨酸（D2 蛋白的第 160 个酪氨酸）通过定点诱变被苯丙氨酸取代His 标记的 psbD 质粒。利用自制的高灵敏Joliot型O_2电极分析闪蒸诱导的析氧过程，监测光合析氧过程中的S态转变过程，无需进一步冷凝即可直接测量培养藻细胞的闪蒸O_2模式。 。在突变细胞中，从O_2到S_1的暗衰变过程中没有检测到快衰变成分，但与野生型细胞相比，慢衰变成分正常，表明Y_D酪氨酸直接参与了快衰变过程。即使在长时间的暗孵育后，闪蒸 O_2 模式也没有改变，表明 Y_D 对 S_1 和 S_0 之间的氧化还原平衡没有贡献。因此，S_1比S_0更高的稳定性主要归因于它们的热力学性质。此外，Y_D酪氨酸的缺失并不影响长时间暗孵育后的O_2能力。结果表明Y_D不参与Mn团簇在暗处的稳定性。事实上，通过Ni亲和柱层析从突变体中制备了具有高O_2活性的PS II核心。该制剂可用于光系统 II 的物理化学研究，包括 ESR 和 FTIR。