FeFe hydrogenases with modified azadithiolate bridgeheads serve as unidirectional hydrogen oxidation catalysts

Semi-synthetic FeFe hydrogenases hold exciting potential as tuneable artificial metalloenzymes for efficient hydrogen conversion (H2 2 H+ + 2e−). This work investigates the role of the bridgehead ligand in the FeFe hydrogenase active site through the study of two bridgehead-modified active-site variants (2-oxapropane 1, 3-dithiolate, ODT, and propane 1, 3-dithiolate, PDT). Replacing the native 2-azapropane 1, 3-dithiolate (ADT) bridgehead with ODT or PDT significantly modifies the active site’s electronic structure and disrupts proton-transfer; yet, some catalytic activity remains, suggesting that alternative catalytic pathways may exist. Notably, when adsorbed onto an electrode, the ODT enzymes operate as proficient catalysts for hydrogen oxidation at higher oxidative overpotentials, achieving current densities of up to 1 mA cm− 2, comparable to those observed for the respective ADT enzyme under the same conditions. Electrochemical analysis reveals similar catalytic behaviour between the ODT and PDT enzymes, suggesting that a proton transfer pathway between the active site and the protein environment still operates, albeit disrupted, in both bridgehead variants under these conditions. Insights into the electronic and catalytic interplay of these modifications could advance the optimisation of semi-synthetic FeFe hydrogenases and design of new bioinspired catalysts.