Correlation of three-dimensional and electronic structure of [FeFe] hydrogenases

• Applicants: Dr. James Birrell; Dr. Ingrid Span
• Subject Area: Structural Biology
• Term: from 2019 to 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 428175165

The proposed project represents a dedicated effort toward combining structural, spectroscopic, functional and theoretical insight into the active site H-cluster of the [FeFe] hydrogenases, the most active hydrogen-converting enzymes in nature. The H-cluster is constructed from a canonical [4Fe-4S] subcluster coupled to a unique [2Fe] sub-cluster containing a unique bridging azapropane dithiolate as well as carbonyl and cyanide ligands. The H-cluster has been subjected to numerous spectroscopic and functional studies over the years, but as yet there is no clear consensus concerning how the catalytic cycle operates and how spectroscopically identified intermediates can be structurally rationalized. Hence, we will crystallize one of the most active and bidirectional [FeFe] hydrogenases from the sulfate-reducing bacterium Desulfovibrio desulfuricans (DdHydAB) and solve its structure in a wide range of catalytic states. The structural data will be supported by electron paramagnetic resonance, and Fourier-transform infrared spectroscopic studies on the same crystals. Finally, molecular theory calculations of suitable models of the H-cluster will be carried out in order to rationalize how structural differences related to reduction, protonation or ligand binding lead to spectroscopic differences. Ultimately, these studies will provide a more profound understanding of the [FeFe] hydrogenase catalytic cycle, and reveal how active site flexibility influences catalytic performance. Our results will have wide-reaching implications for other iron-sulfur enzymes, as well as for the development of synthetic molecular catalysts composed of non-precious metals.

• DFG Programme: Priority Programmes
• Subproject of: SPP 1927: Iron-Sulfur for Life