Engineering Cu-Ce-a Nanozymes: Revolutionary Alloy Nanomaterials Mimicking Cytochrome c Oxidase for Ultra-Sensitive Cytochrome c Detection

The design of synthetic analogs of cytochrome c oxidase (CcO) is a formidable task due to its intricate structure encompassing multiple metal prosthetic sites and protein subunits. In recent years, artificial enzymes based on alloy nanomaterials have garnered significant attention due to the alloy design approach holds promise for the effective tuning of the properties of metal catalysts. In this study, we present copper-cerium alloy nanozymes (Cu-Ce-a NEs), where Cu mimics the active site of CcO, while Ce endows the alloy phase and enhances the capacity to catalyze the oxidation to cytochrome c (Cyt c). Cu-Ce-a NEs functionally mimics CcO, the terminal enzyme in the respiratory electron transport chain (ETC), by catalyzing the four-electron reduction of dioxygen to water. Utilizing the CcO-like properties of Cu-Ce-a NEs, we successfully implemented the electrochemical detection of Cyt c. The Cu-Ce-a NEs based electrochemical sensor revealed a favorable linear range spanning from 2 to 20 μM Cyt c, with a detection limit (LOD) of 2 μM. This method demonstrates high accuracy in Cyt c quantitation in pharmaceuticals, with results closely aligning with the actual concentrations. This finding not only offers new perspectives in the design of enzyme analogs, but also underscores the potential of this method for clinical Cyt c detection, highlighting its significance in biomedical research and diagnostics.