Multiple turnover transfer of [2Fe2S] clusters by the iron-sulfur cluster assembly scaffold proteins IscU and IscA

被引:54
作者
Bonomi, F
Iametti, S
Ta, D
Vickery, LE
机构
[1] Univ Milan, DISMA, Biochem Sect, I-20133 Milan, Italy
[2] Univ Calif Irvine, Dept Physiol & Biophys, Irvine, CA 92697 USA
关键词
D O I
10.1074/jbc.M504344200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
IscU/Isu and IscA/Isa (and related NifU and SufA proteins) have been proposed to serve as molecular scaffolds for preassembly of [FeS] clusters to be used in the biogenesis of iron-sulfur proteins. In vitro studies demonstrating transfer of preformed scaffold-[FeS] complexes to apoprotein acceptors have provided experimental support for this hypothesis, but investigations to date have yielded only single-cluster transfer events. We describe an in vitro assay system that allows for real-time monitoring of [ FeS] cluster formation using circular dichroism spectroscopy and use this to investigate de novo [FeS] cluster formation and transfer from Escherichia coli IscU and IscA to apo-ferredoxin. Both IscU and IscA were found to be capable of multiple cycles of [2Fe2S] cluster formation and transfer suggesting that these scaffold proteins are capable of acting "catalytically." Kinetic studies further showed that cluster transfer exhibits Michaelis-Menten behavior indicative of complex formation of holo-IscU and holo-IscA with apoferredoxin and consistent with a direct [FeS] cluster transfer mechanism. Analysis of the dependence of the rate of cluster transfer, however, revealed enhanced efficiency at low ratios of scaffold to acceptor protein suggesting participation of a transient, labile scaffold-[FeS] species in the transfer process.
引用
收藏
页码:29513 / 29518
页数:6
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