Changes in the electronic structure around Ni in oxidized and reduced selenium-containing hydrogenases from...

Area of Science:

• Biochemistry
• Biophysics
• Enzymology

Background:

• Methanococcus voltae harbors selenium-containing hydrogenases crucial for energy metabolism.
• Understanding the structural and electronic properties of these enzymes is key to elucidating their catalytic mechanisms.
• Previous studies highlighted unique structural features in F420-non-reducing hydrogenase.

Purpose of the Study:

• To characterize the active site structure of the selenium-containing F420-reducing hydrogenase from M. voltae using EPR spectroscopy.
• To compare the active site structures of F420-reducing and F420-non-reducing hydrogenases.
• To investigate the role of selenium in the enzyme's electronic properties.

Main Methods:

• Anaerobic purification of F420-reducing hydrogenase from M. voltae.
• Electron Paramagnetic Resonance (EPR) spectroscopy of purified enzyme under various conditions (reduced, oxidized, illuminated).
• Isotopic enrichment with 77Se to probe nickel-selenium interactions.

Main Results:

• EPR spectra of the purified enzyme are consistent with the active [NiFe]hydrogenase state.
• Hyperfine interaction between nickel and 77Se was observed in the reduced state, suggesting a specific orientation.
• Oxidized states of both hydrogenases showed EPR spectra typical of the inactive 'ready' state, with no Ni-Se hyperfine splitting.
• The active site structures of the two hydrogenases were found to be highly similar.

Conclusions:

• The selenium atom in F420-reducing hydrogenase is positioned to interact with the nickel ion, similar to F420-non-reducing hydrogenase.
• The structural similarity of the active sites suggests that the unusual subunit structure of F420-non-reducing hydrogenase does not significantly alter the active site.
• EPR data provides insights into the electronic configuration and orientation of the nickel-selenium interaction in different redox states.