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

Area of Science:

• Biochemistry
• Enzymology
• Spectroscopy

Background:

• [FeFe] hydrogenases are crucial enzymes for H2 production and consumption.
• Understanding their catalytic mechanism is key to developing clean energy technologies.
• The CO-inhibited H_ox-CO state presents a barrier to studying the active enzyme.

Purpose of the Study:

• To investigate the light-triggered activation mechanism of [FeFe] hydrogenases.
• To study the dynamics of the catalytic site following CO photolysis.
• To establish a method for real-time observation of the enzyme's catalytic cycle.

Main Methods:

• Multiscale UV-pump IR-probe spectroscopy was employed.
• The study focused on the reversible photochemical activation of the CO-inhibited H_ox-CO state.
• Time scales from picoseconds to milliseconds were analyzed.

Main Results:

• Photolysis of the H_ox-CO state leads to rapid CO dissociation within picoseconds.
• The unbound CO remains dissociated for up to milliseconds, creating an active enzyme state.
• This transient active state allows for H2 binding and subsequent catalytic turnover.

Conclusions:

• Photochemical CO release provides a powerful tool to initiate and study [FeFe] hydrogenase activity.
• The observed millisecond time window enables real-time investigation of the complete catalytic cycle.
• This methodology overcomes diffusion limitations and offers high temporal resolution for enzymatic studies.