Electron Inventory of the Iron-Sulfur Scaffold Complex HypCD Essential in [NiFe]-Hydrogenase Cofactor Assembly

09 February 2021, Version 1
This content is a preprint and has not undergone peer review at the time of posting.


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 developed an efficient protocol for the production and isolation of the functional HypCD complex that facilitated detailed spectroscopic investigations. The results obtained by UV/Vis-, electron paramagnetic Resonance (EPR)-, Resonance Raman-, Fourier-transform infrared (FTIR), and Mössbauer spectroscopy provide comprehensive evidence for an electron inventory fit to drive multi-electron redox reactions. We demonstrate the redox activity of the HypCD complex reporting the interconversion of the [4Fe-4S]2+/+ couple. Additionally, we observed a reversible redox conversion between the [4Fe-4S]2+ and a [3Fe-4S]+ cluster. MicroScale thermophoresis indicated preferable binding between the HypCD complex and its interaction partner HypEF under reducing conditions. Together, these results suggest a redox cascade involving the [4Fe-4S] cluster and a conserved disulfide bond of HypD that may facilitate the synthesis of the [Fe](CN)2CO cofactor precursor on the HypCD scaffold complex.


iron-sulfur clusters
redox chemistry
bioinorganic chemistry

Supplementary materials



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