Bacterial [FeFe]-hydrogenases represent promising biocatalysts for high-efficiency production or consumption of hydrogen in bio-electrolysers or bio-fuel cells; thus enabling the use of hydrogen as Energy Vector. The main factor that hinders their application is high sensitivity to oxygen, which is an irreversible inhibitor that destroys the active site of the enzyme.
So far, the [FeFe] hydrogenase from Clostridium beijerinckii, named CbA5H, seems to be the only one with a unique self-protection mechanism against irreversible damage caused by oxygen and other oxidants1. Integrating X-ray crystallography, rational protein design, direct electrochemistry and Fourier transform infrared spectroscopy, the protein mechanism that regulates the reversible transition between the catalytic Hox state and the inactive but oxygen-resistant Hinact state in the CbA5H was studied2.
The open question is what makes this enzyme so unique, and how can we apply the same mechanism to better-characterized enzymes?
The discovery and isolation of CbA5H from a pilot plant revolutionized the understanding of oxygen interactions in [FeFe] hydrogenases, highlighting the importance of enhancing similar protective mechanisms against oxidative damage in these enzymes. Firstly, this research expands the hydrogenase library with two new hydrogenases uncharacterized before by spectroscopy activity and FTIR spectra.
Secondly the study aids in identifying essential residues involved in protection or activity through domain shuffling and mutagenesis, revealing previously unrecognized functional elements.
Insights into oxygen resistance in [FeFe] hydrogenases may inform the design of artificial biocatalysts for biotechnological applications, especially in cascade reactions to boost efficiency and versatility.