Magnetic Resonance Center CERM, University of Florence, Via Luigi Sacconi 6, 50019 Sesto Fiorentino, Florence, Italy; Department of Chemistry, University of Florence, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy.
Magnetic Resonance Center CERM, University of Florence, Via Luigi Sacconi 6, 50019 Sesto Fiorentino, Florence, Italy; Department of Chemistry, University of Florence, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy.
J Mol Biol. 2021 May 14;433(10):166924. doi: 10.1016/j.jmb.2021.166924. Epub 2021 Mar 10.
The late-acting steps of the pathway responsible for the maturation of mitochondrial [4Fe-4S] proteins are still elusive. Three proteins ISCA1, ISCA2 and NFU1 were shown to be implicated in the assembly of [4Fe-4S] clusters and their transfer into mitochondrial apo proteins. We present here a NMR-based study showing a detailed molecular model of the succession of events performed in a coordinated manner by ISCA1, ISCA2 and NFU1 to make [4Fe-4S] clusters available to mitochondrial apo proteins. We show that ISCA1 is the key player of the [4Fe-4S] protein maturation process because of its ability to interact with both NFU1 and ISCA2, which, instead do not interact each other. ISCA1 works as the promoter of the interaction between ISCA2 and NFU1 being able to determine the formation of a transient ISCA1-ISCA2-NFU1 ternary complex. We also show that ISCA1, thanks to its specific interaction with the C-terminal cluster-binding domain of NFU1, drives [4Fe-4S] cluster transfer from the site where the cluster is assembled on the ISCA1-ISCA2 complex to a cluster binding site formed by ISCA1 and NFU1 in the ternary ISCA1-ISCA2-NFU1 complex. Such mechanism guarantees that the [4Fe-4S] cluster can be safely moved from where it is assembled on the ISCA1-ISCA2 complex to NFU1, thereby resulting the [4Fe-4S] cluster available for the mitochondrial apo proteins specifically requiring NFU1 for their maturation.
负责线粒体 [4Fe-4S] 蛋白成熟的途径的后期步骤仍然难以捉摸。已经表明，三种蛋白质 ISCA1、ISCA2 和 NFU1 参与了 [4Fe-4S] 簇的组装及其转移到线粒体脱辅基蛋白中。我们在此提出了一项基于 NMR 的研究，该研究展示了一个详细的分子模型，说明了 ISCA1、ISCA2 和 NFU1 以协调的方式依次进行的一系列事件，以使 [4Fe-4S] 簇能够与线粒体脱辅基蛋白结合。我们表明，ISCA1 是 [4Fe-4S] 蛋白成熟过程的关键参与者，因为它能够与 NFU1 和 ISCA2 相互作用，而 NFU1 和 ISCA2 彼此之间不能相互作用。ISCA1 作为 ISCA2 和 NFU1 之间相互作用的促进剂，能够确定 ISCA1-ISCA2-NFU1 三元复合物的形成。我们还表明，ISCA1 通过其与 NFU1 的 C 端簇结合结构域的特异性相互作用，将 [4Fe-4S] 簇从在 ISCA1-ISCA2 复合物上组装的位置转移到由 ISCA1 和 NFU1 形成的三元 ISCA1-ISCA2-NFU1 复合物中的簇结合位点。这种机制保证了 [4Fe-4S] 簇可以安全地从 ISCA1-ISCA2 复合物上组装的位置转移到 NFU1，从而使 [4Fe-4S] 簇可用于特别需要 NFU1 进行成熟的线粒体脱辅基蛋白。
