@article {Vilela-Alves2024.01.10.571421,
	title = {Substrate-dependent oxidative inactivation of a W-dependent formate dehydrogenase involving selenocysteine displacement},
	journal = {bioRxiv},
	year = {2024},
	note = {n/a},
	publisher = {Cold Spring Harbor Laboratory},
	abstract = {<p>Metal-dependent formate dehydrogenases are very promising targets for enzyme optimization and design of bio-inspired catalysts for CO2 reduction, towards novel strategies for climate change mitigation. For effective application of these enzymes, the catalytic mechanism must be fully understood, and the molecular determinants clarified. Despite numerous studies, several doubts persist, namely regarding the role played by the possible dissociation of the SeCys ligand from the Mo/W active site. Additionally, the O2 sensitivity of these enzymes must also be understood as it poses an important obstacle for biotechnological applications. Here we present a combined biochemical, spectroscopic, and structural characterization of Desulfovibrio vulgaris FdhAB (DvFdhAB) when exposed to oxygen in the presence of a substrate (formate or CO2). This study reveals that O2 inactivation is promoted by the presence of either substrate and involves forming a new species in the active site, captured in the crystal structures, where the SeCys ligand is displaced from tungsten coordination and replaced by a dioxygen or peroxide molecule. This new form was reproducibly obtained and supports the conclusion that, although W-DvFdhAB can catalyze the oxidation of formate in the presence of oxygen for some minutes, it gets irreversibly inactivated after prolonged O2 exposure in the presence of either substrate. These results reveal that oxidative inactivation does not require reduction of the metal, as widely assumed, as it can also occur in the oxidized state in the presence of CO2.Competing Interest StatementThe authors have declared no competing interest.AORAldehyde Oxido-reductaseDTTDithiothreitolDvDesulfovibrio vulgarisEPRElectron Paramagnetic ResonanceFdhFormate dehydrogenaseHPHigh PressureMGDMolybdopterin Guanine DinucleotidesNDNew dropROSReactive Oxygen SpeciesSODSuperoxide dismutaseTSAThermal Shift Assay</p>
},
	doi = {10.1101/2024.01.10.571421},
	url = {https://www.biorxiv.org/content/early/2024/01/10/2024.01.10.571421},
	author = {Vilela-Alves, Guilherme and Rita R. Manuel and Viegas, Aldino and Philippe Carpentier and Frederic Biaso and Guigliarelli, Bruno and Pereira, In{\^e}s A. C. and Rom{\~a}o, Maria Jo{\~a}o and Mota, Cristiano}
}
