Rational design of a mononuclear metal site into the archaeal Rieske-type protein scaffold
Rational design of a mononuclear metal site into the archaeal Rieske-type protein scaffold
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DOI：
10.1074/jbc.m414051200
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发表时间：
2005-03-11
影响因子：
4.8
通讯作者：
Scott, RA
中科院分区：
文献类型：
--
作者：
Iwasaki, T;Kounosu, A;Scott, RA
含有 Rieske 型 [2Fe-2S] 簇的蛋白质在生命的所有三个领域中发挥着重要作用。我们将两个组氨酸配体改造为来自硫磺硫化叶菌的超嗜热古菌 Rieske 型铁氧还蛋白中的 Rieske 型 [2Fe2S] 簇，以修饰配体的类型和间距，并成功地将氧化还原活性位点的金属和簇类型以最小的结构变化转化为天然 Rieske 型蛋白支架。光谱分析明确地在工程化的局部金属结合位点处建立了红氧还蛋白型单核 Fe3+/2+ 中心（Zn2+ 根据表达条件占据铁位点）。这些结果表明配体的类型和间距在生物金属硫蛋白支架的体内簇识别/插入/组装中的重要性。我们认为，局部金属结合位点的早期配体取代和置换事件可能主要允许祖先氧化还原蛋白模块中的金属和簇类型转换，这极大地增强了它们使用有限数量的碱性蛋白支架在生物电子转移导管中进行广泛的独特氧化还原化学的能力。
Proteins containing Rieske-type [2Fe-2S] clusters play essential functions in all three domains of life. We engineered the two histidine ligands to the Rieske-type [2Fe2S] cluster in the hyperthermophilic archaeal Rieske-type ferredoxin from Sulfolobus solfataricus to modify types and spacing of ligands and successfully converted the metal and cluster type at the redox-active site with a minimal structural change to a native Rieske-type protein scaffold. Spectroscopic analyses unambiguously established a rubredoxin-type mononuclear Fe3+/2+ center at the engineered local metal-binding site ( Zn2+ occupies the iron site depending on the expression conditions). These results show the importance of types and spacing of ligands in the in vivo cluster recognition/ insertion/ assembly in biological metallosulfur protein scaffolds. We suggest that early ligand substitution and displacement events at the local metal-binding site(s) might have primarily allowed the metal and cluster type conversion in ancestral redox protein modules, which greatly enhanced their capabilities of conducting a wide range of unique redox chemistry in biological electron transfer conduits, using a limited number of basic protein scaffolds.