Artificial Hydrogenases Based on Cobaloximes and Heme Oxygenase

Area of Science:

• Biochemistry
• Bioinorganic Chemistry
• Enzyme Engineering

Background:

• Heme oxygenase (HO) is a protein with a heme-binding pocket.
• Cobaloxime catalysts are known for their catalytic properties.
• Artificial enzymes aim to mimic natural enzyme functions.

Purpose of the Study:

• To create novel biohybrid catalysts by inserting cobaloxime into heme oxygenase.
• To investigate the activity of these artificial hydrogenases for H2 evolution.
• To understand the structural basis for the observed catalytic activity.

Main Methods:

• Purification and characterization of biohybrid molecules using UV/visible and EPR spectroscopy.
• Quantum chemical/molecular mechanical docking calculations to model binding pocket conformations.
• Activity assays to determine turnover numbers of the artificial hydrogenases.

Main Results:

• Successful insertion of cobaloxime catalysts into the heme-binding pocket of HO.
• Identification of two distinct binding conformations (hydrophobic and hydrophilic) for the cobaloxime.
• Quantum chemical/molecular mechanical docking revealed open and closed conformations of the HO binding pocket.
• HO-based biohybrids showed up to threefold increased turnover numbers compared to cobaloxime alone or myoglobin adducts.

Conclusions:

• The developed HO-based biohybrids are active artificial hydrogenases for H2 evolution in neutral aqueous solutions.
• The distinct binding conformations and pocket dynamics influence catalytic efficiency.
• Site-directed mutagenesis offers a promising strategy for further optimization of these biohybrid catalysts.