Frataxin Accelerates [2Fe-2S] Cluster Formation on the Human Fe鈥揝 Assembly Complex
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- 作者：Nicholas G. Fox ; Deepika Das ; Mrinmoy Chakrabarti ; Paul A. Lindahl ; David P. Barondeau
- 刊名：Biochemistry
- 出版年：2015
- 出版时间：June 30, 2015
- 年：2015
- 卷：54
- 期：25
- 页码：3880-3889
- 全文大小：453K
- ISSN：1520-4995
文摘
Iron鈥搒ulfur (Fe鈥揝) clusters function as protein cofactors for a wide variety of critical cellular reactions. In human mitochondria, a core Fe鈥揝 assembly complex [called SDUF and composed of NFS1, ISD11, ISCU2, and frataxin (FXN) proteins] synthesizes Fe鈥揝 clusters from iron, cysteine sulfur, and reducing equivalents and then transfers these intact clusters to target proteins. In vitro assays have relied on reducing the complexity of this complicated Fe鈥揝 assembly process by using surrogate electron donor molecules and monitoring simplified reactions. Recent studies have concluded that FXN promotes the synthesis of [4Fe-4S] clusters on the mammalian Fe鈥揝 assembly complex. Here the kinetics of Fe鈥揝 synthesis reactions were determined using different electron donation systems and by monitoring the products with circular dichroism and absorbance spectroscopies. We discovered that common surrogate electron donor molecules intercepted Fe鈥揝 cluster intermediates and formed high-molecular weight species (HMWS). The HMWS are associated with iron, sulfide, and thiol-containing proteins and have properties of a heterogeneous solubilized mineral with spectroscopic properties remarkably reminiscent of those of [4Fe-4S] clusters. In contrast, reactions using physiological reagents revealed that FXN accelerates the formation of [2Fe-2S] clusters rather than [4Fe-4S] clusters as previously reported. In the preceding paper [Fox, N. G., et al. (2015) Biochemistry 54, DOI: 10.1021/bi5014485], [2Fe-2S] intermediates on the SDUF complex were shown to readily transfer to uncomplexed ISCU2 or apo acceptor proteins, depending on the reaction conditions. Our results indicate that FXN accelerates a rate-limiting sulfur transfer step in the synthesis of [2Fe-2S] clusters on the human Fe鈥揝 assembly complex.