J. Am. Chem. Soc. https://doi.org/10.1021/jacs.0c04302 (2020)

Hydrogenases are powerful biocatalysts for the production of molecular hydrogen. However, many hydrogenases either are susceptible to oxygen inactivation or, in the case of Escherichia coli Hyd-1, are unable to produce hydrogen because their electron-transferring Fe-S clusters are too oxidizing. To make Hyd-1 a more robust hydrogen producer, Zhang et al. chose two sites near the distal [4Fe–4S] cluster where electrons initially enter Hyd-1 and covalently attached a silver nanocluster (AgNC) to the enzyme surface to make it photoactivatable. The AgNC attached via a cysteine residue installed at either site endowed Hyd-1 with the ability to produce hydrogen at pH 6 under anaerobic or aerobic conditions upon illumination. Covalent, site-specific attachment of the AgNC was necessary for activity, as wild-type Hyd-1 incubated with free AgNC was not active. Electrochemical experiments with AgNC-Hyd-1 complexes indicated that the AgNC is able to capture energized electrons at the protein surface, shuttling them to the internal electron transfer pathway via the distal [4Fe–4S] cluster. This approach provides a potentially versatile method for engineering enzymes with increased catalytic activity in reduction reactions.