Surface-Engineered Nanoparticles Enhance the Peroxidase Activity of Heme-Containing Proteins

Enzymes interfaced with nanomaterials often lose more than 90% of their activity; yet, some nanomaterials have been shown to enhance enzyme activity. However, these findings are largely observational and lack clear and actionable design principles. Systematic studies are needed to develop nanomaterials that can control and tune enzyme activity. Given that enzyme-nanomaterial interactions are mediated by their surface functional groups, we hypothesized that engineering nanoparticle surfaces could allow for controlled tuning of the enzyme activity. In this study, we used peptide-functionalized gold nanoparticles (PGNPs) as a programmable platform to investigate how surface functionalization affects enzyme activity. By varying the peptide sequences, we examined the effects of charge, hydrophobicity, peptide length, and structure on the peroxidase activity of cytochrome C (Cyt C). Our results showed that carefully designed ligands can significantly enhance enzyme activity, exceeding 10-fold compared with the free enzyme. Molecular dynamics simulations provided insights into the molecular basis of these findings, revealing the preferred orientation of Cyt C upon adsorption and key interaction patterns between the enzyme and peptide ligands, thus bridging experimental results with a mechanistic understanding. Furthermore, PGNPs proved to be a versatile platform for boosting peroxidase activity of other heme-containing proteins such as lactoperoxidase, hemoglobin, and catalase by 13.4-, 3.9-, and 4.2-fold, respectively. This study highlights the potential of nanoparticle surface engineering to activate enzymes at interfaces in a tunable manner, offering a promising alternative to protein engineering for developing biocatalysts.