Iron-sulfur cluster biosynthesis -: Characterization of Escherichia coli CyaY as an iron donor for the assembly of [2Fe-2S] clusters in the scaffold IscU
Iron-sulfur cluster biosynthesis -: Characterization of Escherichia coli CyaY as an iron donor for the assembly of [2Fe-2S] clusters in the scaffold IscU
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DOI：
10.1074/jbc.m513569200
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发表时间：
2006-06-16
影响因子：
4.8
通讯作者：
Fontecave, Marc
中科院分区：
文献类型：
--
作者：
Layer, Gunhild;Ollagnier-de Choudens, Sandrine;Fontecave, Marc
铁-硫[ Fe-S]簇的生物成因需要铁和硫化物的协调递送。硫化物由使用L-半胱氨酸作为硫源的半胱氨酸脱硫酶提供。到目前为止，生理性铁供体还没有明确确定。CyaY是frataxin的细菌直系同源物，frataxin是一种被认为参与真核生物中铁硫簇形成的铁结合蛋白，是一个很好的候选者，因为它被证明可以结合铁。然而，没有功能性的体外研究表明参与[ Fe-S]簇生物合成的CyaY已报道到目前为止。在本文中，我们首次证明了CyaY和IscS，半胱氨酸脱硫酶参与铁硫簇组装之间的特定相互作用。铁负载的CyaY蛋白的分析表明，铁3+的强结合和铁2+的弱结合CyaY。生化分析表明，CyaY-Fe 3+蛋白对应于单体，中间体形式（二聚体-五聚体）和低聚物的混合物，中间体对应于CyaY的唯一稳定和可溶性含铁形式。使用光谱方法，进一步证明了这种形式在IscS和半胱氨酸存在下在支架蛋白IscU上的[ Fe-S]簇组装期间在体外作为铁供体是功能性的。所有这些结果都指向CyaY和[ Fe-S]簇生物合成之间的联系，并讨论了该过程的可能机制。
The biogenesis of iron-sulfur [ Fe-S] clusters requires the coordinated delivery of both iron and sulfide. Sulfide is provided by cysteine desulfurases that use L-cysteine as sulfur source. So far, the physiological iron donor has not been clearly identified. CyaY, the bacterial ortholog of frataxin, an iron binding protein thought to be involved in iron-sulfur cluster formation in eukaryotes, is a good candidate because it was shown to bind iron. Nevertheless, no functional in vitro studies showing an involvement of CyaY in [ Fe-S] cluster biosynthesis have been reported so far. In this paper we demonstrate for the first time a specific interaction between CyaY and IscS, a cysteine desulfurase participating in iron-sulfur cluster assembly. Analysis of the iron-loaded CyaY protein demonstrated a strong binding of Fe3+ and a weak binding of Fe2+ by CyaY. Biochemical analysis showed that the CyaY-Fe3+ protein corresponds to a mixture of monomer, intermediate forms ( dimer-pentamers), and oligomers with the intermediate one corresponding to the only stable and soluble iron-containing form of CyaY. Using spectroscopic methods, this form was further demonstrated to be functional in vitro as an iron donor during [ Fe-S] cluster assembly on the scaffold protein IscU in the presence of IscS and cysteine. All of these results point toward a link between CyaY and [ Fe-S] cluster biosynthesis, and a possible mechanism for the process is discussed.