Biocatalytic systems based on
enzyme cascade reactions have attracted growing interest in the field of biocatalytic medicine. However, it is a major challenge to reasonably construct
enzyme cascade reactions with high stability, selectivity, and catalytic efficiency for the in vivo biocatalytic application. Herein, two-in-one engineered
glucose oxidase (GOx-Fe0 ) is fabricated by a biomineralization strategy, through which a nanozyme (Fe0 NP) is anchored within the inner cavity of GOx. Then, GOx-Fe0 is immobilized in a pH-sensitive
metal-organic framework (MOF) zeolitic imidazolate framework-8 (ZIF-8) to establish a stable and effective MOF-immobilized two-in-one engineered
enzyme, GOx-Fe0 @ZIF-8. In vitro studies show that GOx-Fe0 @ZIF-8 exhibits excellent stability and high pH/
glucose selectivity, and the shorter spacing between cascade
enzymes can increase the cascade throughput and effectively improve the reaction efficiency of the
enzyme cascade. In vivo experiments exhibit that GOx-Fe0 @ZIF-8 solves the instability and systemic toxicity of free
enzymes, and achieves deep
tumor penetration and significant chemodynamic therapeutic efficacy through a pH/
glucose-selective
enzyme cascade reaction in
tumor site. Taken together, such an orchestrated
enzyme engineering strategy can effectively improve enzyme stability, selectivity, and
enzyme cascade reaction efficiency via chemical transformations, and also provide a promising strategy for the application of biocatalytic cascade reactions in vivo.