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Temperature Dependence of Structural Dynamics at the Catalytic Cofactor of [FeFe]-hydrogenase

Cite this: Inorg. Chem. 2020, 59, 22, 16474–16488
Publication Date (Web):November 4, 2020
Copyright © 2020 American Chemical Society

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    Abstract Image

    [FeFe]-hydrogenases are nature’s blueprint for efficient hydrogen turnover. Understanding their enzymatic mechanism may improve technological H2 fuel generation. The active-site cofactor (H-cluster) consists of a [4Fe-4S] cluster ([4Fe]H), cysteine-linked to a diiron site ([2Fe]H) carrying an azadithiolate (adt) group, terminal cyanide and carbon monoxide ligands, and a bridging carbon monoxide (μCO) in the oxidized protein (Hox). Recently, the debate on the structure of reduced H-cluster states was intensified by the assignment of new species under cryogenic conditions. We investigated temperature effects (4–280 K) in infrared (IR) and X-ray absorption spectroscopy (XAS) data of [FeFe]-hydrogenases using fit analyses and quantum-chemical calculations. IR data from our laboratory and literature sources were evaluated. At ambient temperatures, reduced H-cluster states with a bridging hydride (μH, in Hred and Hsred) or with an additional proton at [4Fe]H (Hred′) or at the distal iron of [2Fe]H (Hhyd) prevail. At cryogenic temperatures, these species are largely replaced by states that hold a μCO, lack [4Fe]H protonation, and bind an additional proton at the adt nitrogen (HredH+ and HsredH+). XAS revealed the atomic coordinate dispersion (i.e., the Debye–Waller parameter, 2σ2) of the iron–ligand bonds and Fe–Fe distances in the oxidized and reduced H-cluster. 2σ2 showed a temperature dependence typical for the so-called protein–glass transition, with small changes below ∼200 K and a pronounced increase above this “breakpoint”. This behavior is attributed to the freezing-out of larger-scale anharmonic motions of amino acid side chains and water species. We propose that protonation at [4Fe]H as well as ligand rearrangement and μH binding at [2Fe]H are impaired because of restricted molecular mobility at cryogenic temperatures so that protonation can be biased toward adt. We conclude that a H-cluster with a μCO, selective [4Fe]H or [2Fe]H protonation, and catalytic proton transfer via adt facilitates efficient H2 conversion in [FeFe]-hydrogenase.

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    Supporting Information

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    • Experimental and computational FTIR procedures (text), evaluation of IR spectra (Figures S1–S8, Tables S1–S5, and separate spectral file), further XAS data (Figure S9), computational data (Figures S10–S14, Tables S6–S8, and separate movie file), and calculated H-cluster structures (Figure S15 and separate coordinates file) (PDF)

    • Coordinates of Hox structures (PDF)

    • Digitized experimental spectral data (XLSX)

    • Animated structure (MP4)

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    Cited By

    This article is cited by 16 publications.

    1. Alexander Katsyv, Anuj Kumar, Patricia Saura, Maximilian C. Pöverlein, Sven A. Freibert, Sven T. Stripp, Surbhi Jain, Ana P. Gamiz-Hernandez, Ville R. I. Kaila, Volker Müller, Jan M. Schuller. Molecular Basis of the Electron Bifurcation Mechanism in the [FeFe]-Hydrogenase Complex HydABC. Journal of the American Chemical Society 2023, 145 (10) , 5696-5709.
    2. Lili Sun, Carole Duboc, Kaiji Shen. Bioinspired Molecular Electrocatalysts for H2 Production: Chemical Strategies. ACS Catalysis 2022, 12 (15) , 9159-9170.
    3. Sven T. Stripp, Benjamin R. Duffus, Vincent Fourmond, Christophe Léger, Silke Leimkühler, Shun Hirota, Yilin Hu, Andrew Jasniewski, Hideaki Ogata, Markus W. Ribbe. Second and Outer Coordination Sphere Effects in Nitrogenase, Hydrogenase, Formate Dehydrogenase, and CO Dehydrogenase. Chemical Reviews 2022, 122 (14) , 11900-11973.
    4. Moritz Senger, Jifu Duan, Mariia V. Pavliuk, Ulf-Peter Apfel, Michael Haumann, Sven T. Stripp. Trapping an Oxidized and Protonated Intermediate of the [FeFe]-Hydrogenase Cofactor under Mildly Reducing Conditions. Inorganic Chemistry 2022, 61 (26) , 10036-10042.
    5. Christina Felbek, Federica Arrigoni, David de Sancho, Aurore Jacq-Bailly, Robert B. Best, Vincent Fourmond, Luca Bertini, Christophe Léger. Mechanism of Hydrogen Sulfide-Dependent Inhibition of FeFe Hydrogenase. ACS Catalysis 2021, 11 (24) , 15162-15176.
    6. Sven T. Stripp. In Situ Infrared Spectroscopy for the Analysis of Gas-processing Metalloenzymes. ACS Catalysis 2021, 11 (13) , 7845-7862.
    7. Hulin Tai, Shun Hirota, Sven T. Stripp. Proton Transfer Mechanisms in Bimetallic Hydrogenases. Accounts of Chemical Research 2021, 54 (1) , 232-241.
    8. Jason W. Sidabras, Sven T. Stripp. A personal account on 25 years of scientific literature on [FeFe]-hydrogenase. JBIC Journal of Biological Inorganic Chemistry 2023, 28 (4) , 355-378.
    9. Marco Lorenzi, Joe Gellett, Afridi Zamader, Moritz Senger, Zehui Duan, Patricia Rodríguez-Maciá, Gustav Berggren. Investigating the role of the strong field ligands in [FeFe] hydrogenase: spectroscopic and functional characterization of a semi-synthetic mono-cyanide active site. Chemical Science 2022, 13 (37) , 11058-11064.
    10. Federica Arrigoni, Fabio Rizza, Luca Bertini, Luca De Gioia, Giuseppe Zampella. Toward Diiron Dithiolato Biomimetics with Rotated Conformation of the [FeFe]‐Hydrogenase Active Site: A DFT Case Study on Electron‐Rich, Isocyanide‐Based Scaffolds. European Journal of Inorganic Chemistry 2022, 2022 (17)
    11. James A. Birrell, Patricia Rodríguez-Maciá, Edward J. Reijerse, Maria Alessandra Martini, Wolfgang Lubitz. The catalytic cycle of [FeFe] hydrogenase: A tale of two sites. Coordination Chemistry Reviews 2021, 449 , 214191.
    12. Simone Morra, Jifu Duan, Martin Winkler, Philip A. Ash, Thomas Happe, Kylie A. Vincent. Electrochemical control of [FeFe]-hydrogenase single crystals reveals complex redox populations at the catalytic site. Dalton Transactions 2021, 50 (36) , 12655-12663.
    13. Md Estak Ahmed, Dibyajyoti Saha, Lianke Wang, Marcello Gennari, Somdatta Ghosh Dey, Vincent Artero, Abhishek Dey, Carole Duboc. An [FeFe]‐Hydrogenase Mimic Immobilized through Simple Physiadsorption and Active for Aqueous H 2 Production. ChemElectroChem 2021, 8 (9) , 1674-1677.
    14. Konstantin Laun, Iuliia Baranova, Jifu Duan, Leonie Kertess, Florian Wittkamp, Ulf-Peter Apfel, Thomas Happe, Moritz Senger, Sven T. Stripp. Site-selective protonation of the one-electron reduced cofactor in [FeFe]-hydrogenase. Dalton Transactions 2021, 50 (10) , 3641-3650.
    15. Sven T. Stripp. Bonds from bands. Nature Reviews Chemistry 2021, 5 (3) , 146-147.
    16. Marco Lorenzi, Gustav Berggren. [FeFe] Hydrogenases and Their Functional Models. 2021, 731-756.

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