Ligand-Based Redox Chemistry at the Mo-Containing Active Site of Cupriavidus necator Formate Dehydrogenase.

Formate dehydrogenases catalyze the reversible two-electron interconversion of formate and carbon dioxide and generally are assumed to operate through metal-centered redox chemistry at their molybdenum or tungsten active sites. The enzymes also are thought to possess a terminal sulfido ligand which is expected to play a central role in the catalytic mechanism. Here, we investigate the molybdenum-containing formate dehydrogenase FdsDABG from Cupriavidus necator using a combination of Mo K-edge X-ray absorption spectroscopy (XAS), high-energy-resolution fluorescence-detected (HERFD) XAS, extended X-ray absorption fine structure analysis (EXAFS), and density functional theory (DFT) calculations. We observe only subtle spectroscopic changes upon reduction of the oxidized enzyme by formate, suggesting that redox chemistry does not involve the metal; moreover, EXAFS analysis shows no evidence for a terminal sulfido ligand in the oxidized enzyme. DFT calculations support these findings and suggest that the oxidized enzyme possesses a cysteine persulfido structure, which is reduced by cleavage of the S-S bond to form an Mo-SH species, leaving the formal oxidation state of molybdenum unchanged. Collectively, these results suggest that the catalytic reaction of formate dehydrogenases involves ligand-based redox chemistry at a metal center formally reduced to the Mo(IV) state in the oxidized enzyme.