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Nanomaterials with inherent enzyme-mimetic properties have been extensively studied in biosensing field, but it is still challenging to overcome their natively weak enzyme-mimetic activity and further promote their stabilities. In this work, a three-in-one artificial enzyme with synergistically enhanced peroxidase-like activity and outstanding stability is constructed by restricting Au-loaded CeO2 nanorods (NR) (1% Au:CeO2 mass ratio) inside Cu-based metal–organic frameworks (Cu(PABA)), termed Au1/CeO2 NR@Cu(PABA). It has been demonstrated that the artificial enzyme possesses significantly improved performance in catalytic decomposition of H2O2, with ∼2.41 and ∼1.49 times higher activity than its individual constituents (Au1/CeO2 NR and Cu(PABA)), benefiting from the synergistic effect and the unique restricted structure; owns strong affinities to substrates, with separately ∼8.21- and ∼3.13-fold lower Michaelis constants toward H2O2 and o-phenylenediamine, respectively, than horseradish peroxidase; and exhibits desirable pH (4–12), thermal (30–80 °C), organic solvent (N,N-dimethylformamide, acetone, methanol, etc.), and long-term storage (30 days) stabilities, advantaging practical applications. Taking aforementioned superior properties of Au1/CeO2 NR@Cu(PABA), a universal colorimetric/fluorometric dual-mode sensing platform is built and utilized for detection of biomarkers (e.g., glucose, galactose, and cholesterol) in biological fluids with satisfactory recoveries (90.2–108%). This work offers new horizons in designing high-efficiency, stable, and credible biomimetic catalysts to accelerate future advanced engineering of nanozymes.

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