Nitrosylation of human glutathione transferase P1-1 with dinitrosyl diglutathionyl iron complex in vitro and in vivo
Nitrosylation of human glutathione transferase P1-1 with dinitrosyl diglutathionyl iron complex in vitro and in vivo
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DOI：
10.1074/jbc.m507916200
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发表时间：
2005-12-23
影响因子：
4.8
通讯作者：
Lo Bello, M
中科院分区：
文献类型：
--
作者：
Cesareo, E;Parker, LJ;Lo Bello, M
“我们最近已经表明，二亚硝酰二谷氨酸亚硫酰铁复合物，一种可能的体内一氧化氮（NO）供体，以非凡的亲和力与人谷胱甘肽转移酶（GST）P1-1的活性位点之一结合，并在二聚体的相邻亚基中引发负协同作用。这种强烈的相互作用也已在人类Mu、α和Theta GST类中观察到，表明GST可作为细胞内NO载体或清除剂的共同机制。我们在这里提出的GST P1-1的晶体结构，在复杂的二亚硝酰二glutathionyl铁配体在高分辨率。在该复合物中，活性位点Tyr-7通过置换GSH配体之一来通过其酚盐基团与铁原子配位。通过用His、Cys或Phe残基对该残基进行定点诱变，证明了该催化残基在结合一氧化氮供体中的至关重要性。复合物的相对结合亲和力在所有三种突变体中相对于野生型强烈降低约3个数量级。表达重组GST P1-1酶的完整大肠杆菌细胞的电子顺磁共振光谱研究表明，细菌细胞，在响应NO处理，能够形成的二亚硝酰二谷氨酸铁复合物使用细胞内的铁和GSH。我们假设该复合物通过与GST P1-1结合而在体内稳定。
`We have recently shown that dinitrosyl diglutathionyl iron complex, a possible in vivo nitric oxide ( NO) donor, binds with extraordinary affinity to one of the active sites of human glutathione transferase ( GST) P1-1 and triggers negative cooperativity in the neighboring subunit of the dimer. This strong interaction has also been observed in the human Mu, Alpha, and Theta GST classes, suggesting a common mechanism by which GSTs may act as intracellular NO carriers or scavengers. We present here the crystal structure of GST P1-1 in complex with the dinitrosyl diglutathionyl iron ligand at high resolution. In this complex the active site Tyr-7 coordinates to the iron atom through its phenolate group by displacing one of the GSH ligands. The crucial importance of this catalytic residue in binding the nitric oxide donor is demonstrated by site-directed mutagenesis of this residue with His, Cys, or Phe residues. The relative binding affinity for the complex is strongly reduced in all three mutants by about 3 orders of magnitude with respect to the wild type. Electron paramagnetic resonance spectroscopy studies on intact Escherichia coli cells expressing the recombinant GST P1-1 enzyme indicate that bacterial cells, in response to NO treatment, are able to form the dinitrosyl diglutathionyl iron complex using intracellular iron and GSH. We hypothesize the complex is stabilized in vivo through binding to GST P1-1.