An Artificial Metal-Free Peroxidase Designed Using a Ferritin Cage for Bioinspired Catalysis.

Area of Science:

• Biomimetic chemistry
• Protein engineering
• Catalysis

Background:

• Designing artificial enzymes requires precise active site engineering.
• Histidine-rich peptides show peroxidase-mimetic activity but depend on supramolecular structure.
• Ferritin cages offer a potential scaffold for organizing catalytic residues.

Purpose of the Study:

• To design and characterize a ferritin-based artificial enzyme with a histidine-rich active center.
• To investigate the peroxidase-mimetic activity of engineered ferritin mutants.
• To explore the role of the ferritin cage environment in enhancing catalytic efficiency.

Main Methods:

• Engineering ferritin mutants with specific histidine residue arrays on the internal surface.
• Determining crystal structures of ferritin mutants to identify histidine clusters.
• Assessing peroxidase-mimetic activity using 3,3',5,5'-tetramethylbenzidine (TMB) and hydrogen peroxide.
• Employing molecular dynamics simulations to study substrate-enzyme interactions.

Main Results:

• Ferritin mutants formed well-defined histidine clusters (His-clusters) due to histidine-histidine interactions.
• Engineered ferritin exhibited significant peroxidase-mimetic activity, oxidizing TMB in the presence of hydrogen peroxide.
• Molecular dynamics simulations confirmed the co-localization of TMB and hydrogen peroxide within the histidine-rich clusters.
• The confined ferritin cage environment was shown to enhance substrate interactions at the active site.

Conclusions:

• A simple and effective strategy for designing metal-free artificial enzymes using ferritin cages was demonstrated.
• The engineered ferritin acts as a robust platform for creating bioinspired catalysts.
• This approach opens new avenues for innovations in biocatalysis and enzyme mimetics.