Photoactivation of [FeFe] Hydrogenase Studied by Multiscale Time-Resolved Infrared Spectroscopy

High Resolution Image Download MS PowerPoint Slide [FeFe] hydrogenases catalyze the reversible cleavage of H 2, a clean fuel, at exceptional rates. Therefore, understanding their catalytic mechanism is of high importance. Here, we employ multiscale UV pump -IR probe spectroscopy to study the reversible photochemical activation of the CO-inhibited H ox -CO state over picosecond to millisecond time scales. Since this process transforms the catalytic site into the active, H 2 -binding H ox state, photolysis of H ox -CO represents a unique strategy for studying light-triggered activation and the catalytic cycle of the enzyme with high time resolution. We show that the extrinsic CO of H ox -CO dissociates in picoseconds and remains unbound for up to milliseconds. During this time, the enzyme is available for H 2 binding and further catalytic transformations. This time window is sufficiently large to study catalytic processes from the earliest steps to completion of the catalytic cycle. Our approach provides a basis for investigating the catalytic cycle of [FeFe] hydrogenases in real time and without diffusion limitation.