Development of an artificial peroxidase based on a human carbonic anhydrase protein

Artificial metalloenzymes are constructed through the incorporation of a metal catalyst into a protein scaffold. The protein scaffold provides second coordination sphere around the catalytic moiety, endowing it with various attributes to the reaction that the free catalyst does not possess. In this PhD thesis, we will present the development of an artificial peroxidase (ArPase) by combining an Fe-TAML (TAML = Tetra Amide Macrocyclic Ligand) catalyst with a human carbonic anhydrase scaffold. We will show that the protein scaffold enhances the peroxidase activity of the catalyst. Through directed evolution facilitated by screening on cell lysates, the activity of ArPase was improved both in terms of total turnover number and enantiomeric excess. Additionally, a library of chimeric hCAII scaffolds was explored for further improving the performance of the ArPase.
Overall, this thesis highlights the advantage of “genetic” optimization of a Fe-TAML catalyst through the assembly of an ArPase.