Red-emitting europium-doped carbon dots as allosteric catalase inhibitors: Deciphering mechanisms of structural destabilization, thermodynamic binding, and enzymatic suppression

First demonstration of europium-doped carbon dots as allosteric inhibitors of catalase. • Molecular-level elucidation of thermodynamic and structural interaction mechanisms. • Cellular validation shows concentration-dependent suppression of catalase activity. • Provides a new paradigm for designing targeted enzyme-modulating nanomaterials. • Offers fundamental insights for redox-related therapeutic applications. This study addresses a critical gap in understanding the molecular interactions between rare-earth-doped nanomaterials and antioxidant enzymes, with particular emphasis on their structural and functional implications. Red-emitting europium-doped carbon dots (Eu-CDs) were synthesized via a one-pot hydrothermal method and characterized by strong fluorescence emission at 600 nm, providing excellent optical properties suitable for bioimaging applications. Using a comprehensive multidisciplinary approach that integrates fluorescence spectroscopy, circular dichroism, molecular docking simulations, enzyme kinetics, and cellular assays, we systematically investigated the interaction mechanism between Eu-CDs and bovine liver catalase (BLC). Our results demonstrate that Eu-CDs bind specifically to an allosteric site on BLC with moderate affinity (binding constant ∼10 4 L∙mol -1 ), primarily mediated through hydrogen bonding and van der Waals interactions. Thermodynamic analysis revealed the interaction to be spontaneous and enthalpy-driven, with Δ H  = −31.96 kJ∙mol -1 and Δ G  = −23.30 kJ∙mol -1 . This binding induces significant conformational alterations in BLC, including a 26.9% reduction in α -helical content and a 22.4% increase in random coil structures, accompanied by a 48.7% decrease in thermal stability. Functionally, Eu-CDs were identified as non-competitive inhibitors, reducing the maximum catalytic rate by 76.2% while maintaining substrate affinity. Cellular experiments further confirmed concentration-dependent suppression of intracellular catalase activity in HepG2 cells, with 44.6% inhibition observed at 100 μg∙mL -1 Eu-CDs. These mechanistic insights not only advance fundamental understanding of nanomaterial-enzyme interactions but also establish Eu-CDs as promising candidates for modulating redox-related pathways, with potential applications in biomedical research and therapeutic development. Download: Download high-res image (329KB) Download: Download full-size image