Loiseau Laurent, Ollagnier-de Choudens Sandrine, Lascoux David, Forest Eric, Fontecave Marc, Barras Frédéric
Laboratoire de Chimie Bactérienne, UPR-CNRS 9043, Institut de Biologie Structurale et Microbiologie, 31 Chemin Joseph Aiguier, 13402 Marseille Cedex 20, France.
J Biol Chem. 2005 Jul 22;280(29):26760-9. doi: 10.1074/jbc.M504067200. Epub 2005 May 18.
Biogenesis of iron-sulfur (Fe-S) cluster-containing proteins relies on assistance of complex machineries. To date three systems, NIF, ISC, and SUF, were reported to allow maturation of Fe-S proteins. Here we report that the csdA-csdE (formally ygdK) genes of Escherichia coli constitute a sulfur-generating system referred to as CSD which also contributes to Fe-S biogenesis in vivo. This conclusion was reached by applying a thorough combination of both in vivo and in vitro strategies and techniques. Yeast two-hybrid analysis allowed us to show that CsdA and CsdE interact. Enzymology analysis showed that CsdA cysteine desulfurase activity is increased 2-fold in the presence of CsdE. Mass spectrometry analysis and site-directed mutagenesis showed that residue Cys-61 from CsdE acted as an acceptor site for sulfur provided by cysteine desulfurase activity of CsdA. Genetic investigations revealed that the csdA-csdE genes could act as multicopy suppressors of iscS mutation. Moreover, both in vitro and in vivo investigations pointed to a specific connection between the CSD system and quinolinate synthetase NadA.
含铁硫（Fe-S）簇蛋白的生物合成依赖于复杂机制的协助。迄今为止，已报道有三种系统，即NIF、ISC和SUF，可使Fe-S蛋白成熟。在此我们报道，大肠杆菌的csdA-csdE（原称ygdK）基因构成了一个称为CSD的硫生成系统，该系统在体内也有助于Fe-S的生物合成。这一结论是通过全面结合体内和体外的策略与技术得出的。酵母双杂交分析表明CsdA和CsdE相互作用。酶学分析表明，在CsdE存在的情况下，CsdA半胱氨酸脱硫酶活性增加了2倍。质谱分析和定点诱变表明，CsdE的Cys-61残基作为CsdA半胱氨酸脱硫酶活性提供的硫的受体位点。遗传学研究表明，csdA-csdE基因可作为iscS突变的多拷贝抑制子。此外，体外和体内研究均表明CSD系统与喹啉酸合成酶NadA之间存在特定联系。
