In Vitro Assembly as a Tool to Investigate Catalytic Intermediates of [NiFe]-Hydrogenase.

ABSTRACT: [NiFe]-hydrogenases catalyze the reversible reaction H₂ ⇄ 2H⁺ + 2e^-. Their basic module consists of a large subunit, coordinating the NiFe(CO)(CN)₂ center, and a small subunit that carries electron-transferring iron-sulfur clusters. Here, we report the in vitro assembly of fully functional [NiFe]-hydrogenase starting from the isolated large and small subunits. Activity assays complemented by spectroscopic measurements revealed a native-like hydrogenase. This approach was used to label exclusively the NiFe(CO)(CN)₂ center with ⁵⁷Fe, enabling a clear view of the catalytic site by means of nuclear resonance vibrational spectroscopy. This strategy paves the way for in-depth studies of [NiFe]-hydrogenase catalytic intermed

SUBMITTER: Caserta G 

PROVIDER: S-EPMC7932190 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature