Laboratory of Molecular Medicine, Wenzhou Medical College, Wenzhou, 325035, Zhejiang, People's Republic of China.
Biometals. 2012 Dec;25(6):1177-84. doi: 10.1007/s10534-012-9580-4. Epub 2012 Sep 4.
Human mitochondrial protein mitoNEET is a novel target of type II diabetes drug pioglitazone, and contains a redox active [2Fe-2S] cluster that is hosted by a unique ligand arrangement of three cysteine and one histidine residues. Here we report that zinc ion can compete for the [2Fe-2S] cluster binding site in human mitoNEET and potentially modulate the physiological function of mitoNEET. When recombinant mitoNEET is expressed in Escherichia coli cells grown in M9 minimal media, purified mitoNEET contains very little or no iron-sulfur clusters. Addition of exogenous iron or zinc ion in the media produces mitoNEET bound with a [2Fe-2S] cluster or zinc, respectively. Mutations of the amino acid residues that hosting the [2Fe-2S] cluster in mitoNEET diminish the zinc binding activity, indicating that zinc ion and the [2Fe-2S] cluster may share the same binding site in mitoNEET. Finally, excess zinc ion effectively inhibits the [2Fe-2S] cluster assembly in mitoNEET in E. coli cells, suggesting that zinc ion may impede the function of mitoNEET by blocking the [2Fe-2S] cluster assembly in the protein.
人线粒体蛋白 mitoNEET 是 II 型糖尿病药物吡格列酮的一个新靶点，其含有一个氧化还原活性的[2Fe-2S]簇，由三个半胱氨酸和一个组氨酸残基的独特配体排列所组成。在这里，我们报告锌离子可以与人线粒体蛋白 mitoNEET 中的[2Fe-2S]簇结合位点竞争，并可能调节 mitoNEET 的生理功能。当重组 mitoNEET 在生长于 M9 最低培养基的大肠杆菌细胞中表达时，纯化的 mitoNEET 几乎不含或不含铁硫簇。在培养基中添加外源性铁或锌离子，分别产生与[2Fe-2S]簇或锌结合的 mitoNEET。mitoNEET 中含有[2Fe-2S]簇的氨基酸残基的突变降低了锌结合活性，表明锌离子和[2Fe-2S]簇可能在 mitoNEET 中共享相同的结合位点。最后，过量的锌离子有效地抑制了大肠杆菌细胞中 mitoNEET 中的[2Fe-2S]簇组装，表明锌离子可能通过阻止蛋白质中[2Fe-2S]簇的组装来阻碍 mitoNEET 的功能。
