Role of histidine-86 in the catalytic mechanism of ferredoxin:thioredoxin reductase.
Role of histidine-86 in the catalytic mechanism of ferredoxin:thioredoxin reductase.
复制标题
组氨酸 86 在铁氧还蛋白：硫氧还蛋白还原酶催化机制中的作用。
DOI：
10.1021/bi802074p
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发表时间：
2009
期刊：
影响因子：
2.9
通讯作者：
Johnson,MichaelK
中科院分区：
文献类型：
--
作者：
Walters,ElizabethM;Garcia-Serres,Ricardo;Naik,SunilG;Bourquin,Florence;Glauser,DominiqueA;Schürmann,Peter;Huynh,BoiHanh;Johnson,MichaelK
铁氧还蛋白：硫氧还蛋白还原酶利用[Fe2S2]铁氧还蛋白作为单电子供体催化植物叶绿体中硫氧还蛋白的还原，因此在光合作用的光调控中起着核心作用。活性中心包含一个[Fe4S4]团簇，紧邻氧化还原活性二硫键，以顺序的单电子步骤裂解，光谱和结晶学研究相结合揭示了在氧化、单电子和双电子还原状态下涉及新的中心特定的团簇化学的催化机理。组氨酸-86已经成为催化机制中潜在的质子供体/受体，这是因为在氧化还原循环过程中，咪唑环的位置发生了与氧化还原相关的变化，并且H86Y突变体的活性大大降低。在这里，我们报道了聚球藻中[Fe4S4]中心的光谱和氧化还原表征。PCC6803H86Y铁氧还蛋白：索氧化还蛋白还原酶在纯化态和N-乙基马来酰亚胺修饰态的可及氧化还原状态下，使用UV−可见光吸收和变温磁圆二色谱、电子顺磁共振、共振拉曼光谱和Mo-̈穆斯堡尔谱相结合。结果表明，His86是形成部分价局域化的[Fe4S4]2+团簇的必要条件，[Fe4S4]2+团簇是两电子还原中间体的标志。结合现有的结构数据，光谱结果表明，His86在团簇相互作用的硫醇的质子化/去质子化以及在两电子还原的中间体中将团簇与硫醇相互作用的团簇锚定在靠近团簇的位置上具有功能作用。
Ferredoxin:thioredoxin reductase catalyzes the reduction of thioredoxins in plant chloroplasts using the [Fe2S2] ferredoxin as a one-electron donor and as such plays a central role in light regulation of oxygenic photosynthesis. The active-site comprises a [Fe4S4] cluster next to a redox-active disulfide that is cleaved in sequential one-electron steps and the combination of spectroscopic and crystallographic studies have revealed a catalytic mechanism involving novel site specific cluster chemistry in the oxidized, one-electron- and two-electron-reduced redox states. Histidine-86 has emerged as a potential proton donor/acceptor in the catalytic mechanism based on redox-related changes in the positioning of the imidazole ring during redox cycling and greatly decreased activity for the H86Y variant. Here we report on spectroscopic and redox characterization of the [Fe4S4] center inSynechocystissp. PCC 6803 H86Y ferredoxin:thoredoxin reductase in the accessible redox states of both the as purified andN-ethylmaleimide-modified forms, using the combination of UV−visible absorption and variable-temperature magnetic circular dichroism, EPR, resonance Raman and Mössbauer spectroscopies. The results demonstrate that His86 is required for formation of the partially valence-localized [Fe4S4]2+cluster that is the hallmark of two-electron-reduced intermediate. Taken together with the available structural data, the spectroscopic results indicate a functional role for His86 in protonation/deprotonation of the cluster-interacting thiol and anchoring the cluster interacting thiol in close proximity to the cluster in the two-electron-reduced intermediate.