Electrochemical control of [FeFe]-hydrogenase single crystals reveals complex redox populations at the catalytic site

Morra, Simone; Duan, Jifu; Winkler, Martin; Ash, Philip A.; Happe, Thomas; Vincent, Kylie A.

Abstract

Elucidating the distribution of intermediates at the active site of redox metalloenzymes is vital to understanding their highly efficient catalysis. Here we demonstrate that it is possible to generate, and detect, the key catalytic redox states of an [FeFe]-hydrogenase in a protein crystal. Individual crystals of the prototypical [FeFe]-hydrogenase I from Clostridium pasteurianum (CpI) are maintained under electrochemical control, allowing for precise tuning of the redox potential, while the crystal is simultanaously probed via Fourier Transform Infrared (FTIR) microspectroscopy. The high signal/noise spectra reveal potential-dependent variation in the distribution of redox states at the active site (H-cluster) according to state-specific vibrational bands from the endogeneous CO and CN- ligands. CpI crystals are shown to populate the same H-cluster states as those detected in solution, including the oxidised species Hox, the reduced species Hred/HredH+, the super-reduced HsredH+ and the hydride species Hhyd. The high sensitivity and precise redox control offered by this approach also facilitates the detection and characterisation of low abundance species that only accumulate within a narrow window of conditions, revealing new redox intermediates.

Citation

Morra, S., Duan, J., Winkler, M., Ash, P. A., Happe, T., & Vincent, K. A. (2021). Electrochemical control of [FeFe]-hydrogenase single crystals reveals complex redox populations at the catalytic site. Dalton Transactions, 50(36), 12655-12663. https://doi.org/10.1039/d1dt02219a

• Journal Article Type: Article
• Acceptance Date: Jul 12, 2021
• Online Publication Date: Jul 13, 2021
• Publication Date: Sep 28, 2021
• Deposit Date: Aug 19, 2021
• Publicly Available Date: Aug 23, 2021
• Journal: Dalton Transactions
• Print ISSN: 1477-9226
• Electronic ISSN: 1477-9234
• Publisher: Royal Society of Chemistry
• Peer Reviewed: Peer Reviewed
• Volume: 50
• Issue: 36
• Pages: 12655-12663
• DOI: https://doi.org/10.1039/d1dt02219a
• Keywords: Hydrogenase, bioinorganic chemitry
• Public URL: https://nottingham-repository.worktribe.com/output/6057908
• Publisher URL: https://pubs.rsc.org/en/content/articlelanding/2021/dt/d1dt02219a