Formation of [4Fe-4S] Clusters in the Mitochondrial Iron鈥揝ulfur Cluster Assembly Machinery

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• 作者：Diego Brancaccio ; Angelo Gallo ; Maciej Mikolajczyk ; Kairit Zovo ; Peep Palumaa ; Ettore Novellino ; Mario Piccioli ; Simone Ciofi-Baffoni ; Lucia Banci
• 刊名：Journal of the American Chemical Society
• 出版年：2014
• 出版时间：November 19, 2014
• 年：2014
• 卷：136
• 期：46
• 页码：16240-16250
• 全文大小：620K
• ISSN：1520-5126

文摘

The generation of [4Fe-4S] clusters in mitochondria critically depends, in both yeast and human cells, on two A-type ISC proteins (in mammals named ISCA1 and ISCA2), which perform a nonredundant functional role forming in vivo a heterocomplex. The molecular function of ISCA1 and ISCA2 proteins, i.e., how these proteins help in generating [4Fe-4S] clusters, is still unknown. In this work we have structurally characterized the Fe/S cluster binding properties of human ISCA2 and investigated in vitro whether and how a [4Fe-4S] cluster is assembled when human ISCA1 and ISCA2 interact with the physiological [2Fe-2S]²⁺ cluster-donor human GRX5. We found that (i) ISCA2 binds either [2Fe-2S] or [4Fe-4S] cluster in a dimeric state, and (ii) two molecules of [2Fe-2S]²⁺ GRX5 donate their cluster to a heterodimeric ISCA1/ISCA2 complex. This complex acts as an 鈥渁ssembler鈥?of [4Fe-4S] clusters; i.e., the two GRX5-donated [2Fe-2S]²⁺ clusters generate a [4Fe-4S]²⁺ cluster. The formation of the same [4Fe-4S]²⁺ cluster-bound heterodimeric species is also observed by having first one [2Fe-2S]²⁺ cluster transferred from GRX5 to each individual ISCA1 and ISCA2 proteins to form [2Fe-2S]²⁺ ISCA2 and [2Fe-2S]²⁺ ISCA1, and then mixing them together. These findings imply that such heterodimeric complex is the functional unit in mitochondria receiving [2Fe-2S] clusters from hGRX5 and assembling [4Fe-4S] clusters before their transfer to the final target apo proteins.