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Open AccessArticle

Infrared Characterization of the Bidirectional Oxygen-Sensitive [NiFe]-Hydrogenase from E. coli

Department of Physics, Experimental Molecular Biophysics, Freie Universität Berlin, 14195 Berlin, Germany
Department of Physics, Genetic Biophysics, Freie Universität Berlin, 14195 Berlin, Germany
Author to whom correspondence should be addressed.
These authors contributed equally.
Catalysts 2018, 8(11), 530;
Received: 16 October 2018 / Revised: 31 October 2018 / Accepted: 6 November 2018 / Published: 8 November 2018
[NiFe]-hydrogenases are gas-processing metalloenzymes that catalyze the conversion of dihydrogen (H2) to protons and electrons in a broad range of microorganisms. Within the framework of green chemistry, the molecular proceedings of biological hydrogen turnover inspired the design of novel catalytic compounds for H2 generation. The bidirectional “O2-sensitive” [NiFe]-hydrogenase from Escherichia coli HYD-2 has recently been crystallized; however, a systematic infrared characterization in the presence of natural reactants is not available yet. In this study, we analyze HYD-2 from E. coli by in situ attenuated total reflection Fourier-transform infrared spectroscopy (ATR FTIR) under quantitative gas control. We provide an experimental assignment of all catalytically relevant redox intermediates alongside the O2- and CO-inhibited cofactor species. Furthermore, the reactivity and mutual competition between H2, O2, and CO was probed in real time, which lays the foundation for a comparison with other enzymes, e.g., “O2-tolerant” [NiFe]-hydrogenases. Surprisingly, only Ni-B was observed in the presence of O2 with no indications for the “unready” Ni-A state. The presented work proves the capabilities of in situ ATR FTIR spectroscopy as an efficient and powerful technique for the analysis of biological macromolecules and enzymatic small molecule catalysis. View Full-Text
Keywords: redox enzymes; FTIR spectroscopy; small molecules redox enzymes; FTIR spectroscopy; small molecules
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MDPI and ACS Style

Senger, M.; Laun, K.; Soboh, B.; Stripp, S.T. Infrared Characterization of the Bidirectional Oxygen-Sensitive [NiFe]-Hydrogenase from E. coli. Catalysts 2018, 8, 530.

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