The oxygen-resistant [FeFe]-hydrogenase CbA5H harbors an unknown radical signal

[FeFe]-hydrogenases catalyze the reversible conversion of molecular hydrogen into protons and electrons with remarkable efficiency. However, their industrial applications are limited by their oxygen sensitivity. Recently, it was shown that the [FeFe]-hydrogenase from Clostridium beijerinckii (CbA5H)...

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Bibliographic Details
Published in:Chemical science (Cambridge) 2022-06, Vol.13 (24), p.7289-7294
Main Authors: Heghmanns, Melanie, Rutz, Andreas, Kutin, Yury, Engelbrecht, Vera, Winkler, Martin, Happe, Thomas, Kasanmascheff, Müge
Format: Article
Language:English
Subjects:
Chemistry
Clusters
Continuous radiation
Density distribution
Electron paramagnetic resonance
Hydrogenase
Industrial applications
Iron 57
Oxidation
Oxygen
Protons
Signal generation
Spectroscopy
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Summary:[FeFe]-hydrogenases catalyze the reversible conversion of molecular hydrogen into protons and electrons with remarkable efficiency. However, their industrial applications are limited by their oxygen sensitivity. Recently, it was shown that the [FeFe]-hydrogenase from Clostridium beijerinckii (CbA5H) is oxygen-resistant and can be reactivated after oxygen exposure. In this work, we used multifrequency continuous wave and pulsed electron paramagnetic resonance (EPR) spectroscopy to characterize the active center of CbA5H, the H-cluster. Under oxidizing conditions, the spectra were dominated by an additional and unprecedented radical species. The generation of this radical signal depends on the presence of an intact H-cluster and a complete proton transfer pathway including the bridging azadithiolate ligand. Selective 57 Fe enrichment combined with isotope-sensitive electron-nuclear double resonance (ENDOR) spectroscopy revealed a spin density distribution that resembles an H-cluster state. Overall, we uncovered a radical species in CbA5H that is potentially involved in the redox sensing of CbA5H. Electron paramagnetic resonance spectroscopy revealed an unprecedented radical species in the oxygen-resistant [FeFe]-hydrogenase CbA5H. Analysis of the isotope-sensitive data suggests that it is related to the active site, the H-cluster.
ISSN:2041-6520
2041-6539
DOI:10.1039/d2sc00385f