Structural and Spectroscopic Insights into Catalytic Intermediates of a [NiFe]-hydrogenase from Group 3.

Hydrogenases catalyze reversible H₂ production and are potential models for renewable energy catalysts. Here, the full redox landscape of a group 3 [NiFe]-hydrogenase from methanothermococcus thermolithotrophicus is elucidated, resembling group 1 enzymes. Structural and spectroscopic analyses reveal a catalytic-ready state with nickel seesaw coordination, enabling intermediate trapping and advancing mechanistic understanding of oxygen-sensitive [NiFe] enzymes.