Electrochemistry, AFM, and PM-IRRA Spectroscopy of Immobilized Hydrogenase: Role of a Hydrophobic Helix in Enzyme Orientation for Efficient H2 Oxidation

摘要

Nickel-iron hydrogenase ([NiFe] Hase) catalyzes hydrogen splitting into protons and electrons, and is a potential biocatalyst in fuel cells.Three FeS clusters aligned as a conductive wire drive electrons from the [NiFe] active site to the surface of the enzyme, where the redox partner (including the electrode) binds. Direct enzyme connection gave access to thermodynamic and kinetic data of enzymatic reactions through direct electron transfer (DET). Mediated electron transfer (MET) allowed recreation of the physiological electron-transfer chain, and/or connection of unfavorably oriented enzymes. Previous work demonstrated that DET or MET processes for H2 oxidation by a soluble, O2-sensitive [NiFe] Hase from Desulfovibrio species could be controlled by electrostatic interaction. The presence of an acidic patch of amino acids, coupled to a dipole moment pointing towards the distal FeS cluster (positioned at the surface of the enzyme), allowed orientation of the enzyme, which turned upside down as a function of the charge on the electrochemical interface.