DNA-directed enzyme immobilization on Fe₃O₄ modified with nitrogen-doped graphene quantum dots as a highly efficient and stable multi-catalyst system

The exploration of new tactics for manufacturing artificial immobilized multi-enzyme systems based on enzyme cascades has recently attracted considerable interest because of the urgent need for multi-enzyme catalysts and the high cost of free enzymes. Because of the inevitable limitations of native enzymes such as instability and storage issues, the development of nanozyme–enzyme cascades is needed. A versatile strategy was developed for fabricating an efficient multi-catalyst system by immobilizing glucose oxidase (GOx) on ferriferous oxide nanocomposites functionalized with nitrogen-doped graphene quantum dots (Fe₃O₄@N-GQDs) through DNA-directed immobilization. The Fe₃O₄@N-GQDs acted as a carrier for the natural enzyme and showed high peroxidase activity which enabled an enzyme cascade that included GOx to be set up. This multi-catalyst system showed great catalytic activity, reversibility and operational stability. The surfaces of GOx-targeted magnetic nanoparticles were regenerated by mild dehybridization of DNA. The Michaelis constant (K_m) and maximum initial velocity (V_max) of the multi-catalyst system were 1.069 mM and 11.2 × 10⁻⁸ M s⁻¹, respectively, which are considerably better than the corresponding values for adsorbed and free bienzyme combinations. The increased bioactivity of the multi-catalyst system is ascribed to the satisfactory peroxidase-like activity of Fe₃O₄@N-GQDs, the enzyme–promoting effect of the QDs and enhancement by DNA-directed immobilization. Because of the diverse range of possible nanozyme–enzyme combinations and high efficiency of this approach, this work provides a novel pathway for the manufacturing of synthetic enzyme catalyst systems, which have great potential in the field of biotechnology.