期刊
BIOCHEMICAL JOURNAL
卷 478, 期 17, 页码 3281-3295出版社
PORTLAND PRESS LTD
DOI: 10.1042/BCJ20210224
关键词
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资金
- German Research Foundation (DFG) via the SPP 1927 priority program 'Iron-Sulfur for Life' [SO1325/5-2, STR1554/5-1, AD178/7-1, SCHU1251/17-2, PI610/2-1]
- DFG [EXC 2008/1-390540038]
The study demonstrates that the [Fe](CN)2CO cofactor and the [4Fe-4S] cluster of the HypCD complex have redox activities, with potential-dependent interconversion processes. Furthermore, potential-dependent disulfide formation between the metal centers may suggest an electron confurcation step.
The [4Fe-4S] cluster containing scaffold complex HypCD is the central construction site for the assembly of the [Fe](CN)2CO cofactor precursor of [NiFe]-hydrogenase. While the importance of the HypCD complex is well established, not much is known about the mechanism by which the CN- and CO ligands are transferred and attached to the iron ion. We report an efficient expression and purification system producing the HypCD complex from E. coli with complete metal content. This enabled in-depth spectroscopic characterizations. The results obtained by EPR and Mossbauer spectroscopy demonstrate that the [Fe](CN)2CO cofactor and the [4Fe-4S] cluster of the HypCD complex are redox active. The data indicate a potential-dependent interconversion of the [Fe]2+/3+ and [4Fe-4S]2+/+ couple, respectively. Moreover, ATR FTIR spectroscopy reveals potentialdependent disulfide formation, which hints at an electron confurcation step between the metal centers. MicroScale thermophoresis indicates preferable binding between the HypCD complex and its in vivo interaction partner HypE under reducing conditions. Together, these results provide comprehensive evidence for an electron inventory fit to drive multi-electron redox reactions required for the assembly of the CN- and CO ligands on the scaffold complex HypCD.
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