Probing the Solvent Accessibility of the [4Fe-4S] Cluster of the Hydrogenase Maturation Protein HydF from Thermotoga neapolitana by HYSCORE and 3p-ESEEM
Probing the Solvent Accessibility of the [4Fe-4S] Cluster of the Hydrogenase Maturation Protein HydF from Thermotoga neapolitana by HYSCORE and 3p-ESEEM
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DOI：
10.1021/acs.jpcb.5b03110
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发表时间：
2015-10-29
影响因子：
3.3
通讯作者：
Carbonera, Donatella
中科院分区：
文献类型：
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作者：
Albertini, Marco;Berto, Paola;Carbonera, Donatella
[FeFe]-氢化酶的催化位点，“H-簇”，由一个[4Fe-4S]单元通过一个半胱氨酰残基连接到一个[2Fe]中心，由三个CO，两个CN-和一个桥接二硫代硫酸盐协调组成，在一个复杂的成熟途径中组装，目前尚未完全表征，涉及三个保守蛋白质，HydG，HydE和HydF。HydF是一种复合酶，被认为是H-簇的[2Fe]亚基的支架和载体。这种成熟酶蛋白本身含有一个[4Fe-4S]簇结合位点，具有三个保守的半胱氨酸残基和一个noncysteinyl第四配体。在这项工作中，我们已经利用3 p-ESEEM和HYSCORE光谱，以了解结构和化学环境的[4Fe-4S]簇的HydF从超嗜热生物Thermotoga neapolitana。第四配体的性质和溶剂的可及性的活性位点，包括[4Fe-4S]簇的H/D交换后获得的光谱结果的基础上进行了讨论。我们建议，noncysteinyl连接的铁原子的[4Fe-4S]簇是网站的[2Fe]子簇前体被锚定，并最终处理被交付给氢化酶（HydA）。
The catalytic site of [FeFe]-hydrogenase, the "H-cluster", composed of a [4Fe-4S] unit connected by a cysteinyl residue to a [2Fe] center coordinated by three CO, two CN-, and a bridging dithiolate, is assembled in a complex maturation pathway, at present not fully characterized, involving three conserved proteins, HydG, HydE, and HydF. HydF is a complex enzyme, which is thought to act as a scaffold and carrier for the [2Fe] subunit of the H-cluster. This maturase protein contains itself a [4Fe-4S] cluster binding site, with three conserved cysteine residues and a noncysteinyl fourth ligand. In this work, we have exploited 3p-ESEEM and HYSCORE spectroscopies to get insight into the structure and the chemical environment of the [4Fe-4S] cluster of HydF from the hyperthermophilic organism Thermotoga neapolitana. The nature of the fourth ligand and the solvent accessibility of the active site comprising the [4Fe-4S] cluster are discussed on the basis of the spectroscopic results obtained upon H/D exchange. We propose that the noncysteinyl ligated Fe atom of the [4Fe-4S] cluster is the site where the [2Fe] subcluster precursor is anchored and finally processed to be delivered to the hydrogenase (HydA).