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Site-selective Protonation of the One-electron Reduced Cofactor in [FeFe]-Hydrogenase

preprint
revised on 14.09.2020 and posted on 16.09.2020 by Konstantin Laun, Iuliia Baranova, Jifu Duan, Leonie Kertess, Florian Wittkamp, Ulf-Peter Apfel, Thomas Happe, Moritz Senger, Sven T. Stripp
Hydrogenases are microbial redox enzymes that catalyze H2 oxidation and proton reduction (H2 evolution). While all hydrogenases show high oxidation activities, the majority of [FeFe]-hydrogenases are excellent H2 evolution catalysts as well. Their active site cofactor comprises a [4Fe-4S] cluster covalently linked to a diiron site equipped with carbon monoxide and cyanide ligands that facilitate catalysis at low overpotential. Distinct proton transfer pathways connect the active site niche with the solvent, resulting in a non-trivial dependence of hydrogen turnover and bulk pH. To analyze the catalytic mechanism of [FeFe]-hydrogenase, we employ in situ infrared spectroscopy and infrared spectro-electrochemistry. Titrating the pH under H2 oxidation or H2 evolution conditions reveals the influence of site-selective protonation on the equilibrium of reduced cofactor states. Governed by pKa differences across the active site niche and proton transfer pathways, we find that individual electrons are stabilized either at the [4Fe-4S] cluster (alkaline pH values) or at the diiron site (acidic pH values). This observation is discussed in the context of the natural pH dependence of hydrogen turnover as catalyzed by [FeFe]-hydrogenase.

History

Email Address of Submitting Author

sven.stripp@fu-berlin.de

Institution

Freie Universität Berlin

Country

Germany

ORCID For Submitting Author

0000-0002-8412-0258

Declaration of Conflict of Interest

None

Exports