Because of the high catalytic activities and substrate specificity, natural enzymes have been widely used in industrial, medical, and biological fields, etc. Although promising, they often suffer from intrinsic shortcomings such as high cost, low operational stability, and difficulties of recycling. To overcome these shortcomings, researchers have been devoted to the exploration of artificial enzyme mimics for a long time. Since the discovery of ferromagnetic nanoparticles with intrinsic horseradish peroxidase-like activity in 2007, a large amount of studies on nanozymes have been constantly emerging in the next decade. Nanozymes are one kind of nanomaterials with enzymatic catalytic properties. Compared with natural enzymes, nanozymes have the advantages such as low cost, high stability and durability, which have been widely used in industrial, medical, and biological fields. A thorough understanding of the possible catalytic mechanisms will contribute to the development of novel and high-efficient nanozymes, and the rational regulations of the activities of nanozymes are of great significance. In this review, we systematically introduce the classification, catalytic mechanism, activity regulation as well as recent research progress of nanozymes in the field of biosensing, environmental protection, and disease treatments, etc. in the past years. We also propose the current challenges of nanozymes as well as their future research focus. We anticipate this review may be of significance for the field to understand the properties of nanozymes and the development of novel nanomaterials with enzyme mimicking activities.

Nanozymes: Classification, Catalytic Mechanisms, Activity Regulation, and Applications

The terms 'antioxidant', 'oxidative stress' and 'oxidative damage' are widely used but rarely defined. This brief review attempts to define them and to examine the ways in which oxidative stress and oxidative damage can affect cell behaviour both in vivo and in cell culture, using cancer as an example.

Biochemistry of oxidative stress

Nanozyme, a term defined for nanomaterial with enzyme-like properties, has attracted significant research attention owing to its striking merits. Recently, a surge of nanozymes have been demonstrated to catalyze some typical enzymatic reactions mimicking oxidase, peroxidase and catalase. Especially, nanozymes with peroxidase-like activity have grown into a big family due to their broad range of applications in the field of biosensing and immunoassay. Since inorganic nanoparticles possess the advantages of high stability and easy surface modification, nanozymes have been emerging alternatives to natural enzymes to some extent. In this Review, we briefly summarize several typical nanozymes and then focus our attention on their enormous applications with respect to analytical chemistry. Representative examples would be discussed in detail from the literatures of last 10 years. Additionally, the current challenges and future directions about nanozymes are speculated at the end of this review.

Nanozyme: An emerging alternative to natural enzyme for biosensing and immunoassay

As a new generation of artificial enzymes, nanozymes have the advantages of high catalytic activity, good stability, low cost, and other unique properties of nanomaterials. Due to their wide range of potential applications, they have become an emerging field bridging nanotechnology and biology, attracting researchers in various fields to design and synthesize highly catalytically active nanozymes. However, the thorough understanding of experimental phenomena and the mechanisms beneath practical applications of nanozymes limits their rapid development. Herein, the progress of experimental and computational research of nanozymes on two issues over the past decade is briefly reviewed: (1) experimental development of new nanozymes mimicking different types of enzymes. This covers their structures and applications ranging from biosensing and bioimaging to therapeutics and environmental protection. (2) The catalytic mechanism proposed by experimental and theoretical study. The challenges and future directions of computational research in this field are also discussed.

Recent Advances in Nanozyme Research

Reactive oxygen species (ROS)-induced oxidative stress is linked to various diseases, including cardiovascular disease and cancer Though highly efficient natural ROS scavenging enzymes have been evolved, they are sensitive to environmental conditions and hard to mass-produce Therefore, enormous efforts have been devoted to developing artificial enzymes with ROS scavenging activities Among them, ROS scavenging nanozymes have recently attracted great interest owing to their enhanced stability, multi-functionality, and tunable activity It has been implicated that Mn-contained nanozymes would possess efficient ROS scavenging activities, however only a few such nanozymes have been reported To fill this gap, herein we demonstrated that Mn3O4 nanoparticles (NPs) possessed multiple enzyme mimicking activities (ie, superoxide dismutase and catalase mimicking activities as well as hydroxyl radical scavenging activity) The Mn3O4 nanozymes therefore significantly scavenged superoxide radical as well as hydrogen peroxide and hydroxyl radical Moreover, they were not only more stable than the corresponding natural enzymes but also superior to CeO2 nanozymes in terms of ROS elimination We showed that the Mn3O4 NPs not only exhibited excellent ROS removal efficacy in vitro but also effectively protected live mice from ROS-induced ear-inflammation in vivo These results indicated that Mn3O4 nanozymes are promising therapeutic nanomedicine for treating ROS-related diseases

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ROS scavenging Mn3O4 nanozymes for in vivo anti-inflammation

Nanomaterials with enzyme-like activities (nanozymes) attracts significant interest due to their therapeutic potential for the treatment of various diseases. Herein, we report that a Mn3 O4 nanozyme functionally mimics three major antioxidant enzymes, that is, superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) and the multienzyme activity is size as well as morphology-dependent. The redox modulatory effect of Mn3 O4 plays a crucial role in protecting the cells from MPP+ induced cytotoxicity in a Parkinson disease (PD)-like cellular model, indicating that manganese-based nanomaterials having multi-enzyme activity can robustly rescue the cells from oxidative damage and thereby possess therapeutic potential to prevent ROS-mediated neurological disorders.

A Redox Modulatory Mn3O4 Nanozyme with Multi-Enzyme Activity Provides Efficient Cytoprotection to Human Cells in a Parkinson's Disease Model

Biocompatible nanoparticles with an intrinsic ability to mimic the cellular antioxidant enzymes are potential candidates for the development of new therapeutics for various oxidative stress related disorders. However, the understanding of the interaction and the mechanistic crosstalk between the nanoparticles and the cellular biomolecules is limited. Here we show that the multienzyme mimic manganese(ii,iii) oxide, Mn3O4, in nanoform (Mp) rescues the cells from oxidative damage induced by reactive oxygen species (ROS). The nanoparticles provide remarkable protection to biomolecules against the ROS-mediated protein oxidation, lipid peroxidation and DNA damage. Interestingly, the endogenous antioxidant machinery remains unaltered in the presence of these nanozymes, indicating the small molecule targeting of these nanoparticles in the cellular redox modulation. This study delineates the possible mechanism by which the nanoparticles provide protection to the cells against the adverse effects of oxidative stress. Based on our observation, we suggest that the multienzyme mimic Mn3O4 nanoparticles possess great potential in suppressing the oxidative stress-mediated pathophysiological conditions under which the antioxidant system is overwhelmed.

A manganese oxide nanozyme prevents the oxidative damage of biomolecules without affecting the endogenous antioxidant system.

Nanoparticles that functionally mimic the activity of metal-containing enzymes (metallo-nanozymes) are of therapeutic importance for treating various diseases. However, it is still not clear whether such nanozymes can completely substitute the function of natural enzymes in living cells. In this work, we show for the first time that a cerium vanadate (CeVO4 ) nanozyme can substitute the function of superoxide dismutase 1 and 2 (SOD1 and SOD2) in the neuronal cells even when the natural enzyme is down-regulated by specific gene silencing. The nanozyme prevents the mitochondrial damage in SOD1- and SOD2-depleted cells by regulating the superoxide levels and restores the physiological levels of the anti-apoptotic Bcl-2 family proteins. Furthermore, the nanozyme effectively prevents the mitochondrial depolarization, leading to a significant improvement in the cellular levels of ATP under oxidative stress.

A Cerium Vanadate Nanozyme with Specific Superoxide Dismutase Activity Regulates Mitochondrial Function and ATP Synthesis in Neuronal Cells

Nanomaterials having enzyme-like activity (nanozymes) make them suitable candidates for various biomedical applications. In this study, we demonstrate the morphology-dependent enzyme mimetic activity of Mn3 O4 nanoparticles. It is found that Mn3 O4 nanoparticles mimic the functions of all three cellular antioxidant enzymes: superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx). Interestingly, the nanozyme activity of Mn3 O4 depends on various factors including size, morphology, surface area, and the redox properties of the metal ions. The Mn3 O4 nanoflowers exhibited remarkably high activity in all three enzyme systems and the order of multienzyme activity of different morphologies was: flowers ≫ flakes > hexagonal plates≈polyhedrons≈cubes. Interestingly, all five nanoforms are taken up by the mammalian cells and were found to be biocompatible, with very low cytotoxicity. The activity of the most active nanoflowers was studied in primary human umbilical vein endothelial cells (HUVEC) and human pulmonary microvascular endothelial cells (hPMEC) and it was found that Mn3 O4 does not reduce the level of nitric oxide (NO). This is in contrast to the effect of some of the Mn-porphyrin-based SOD mimetics, which are known to scavenge NO in endothelial cells.

Manganese-Based Nanozymes: Multienzyme Redox Activity and Effect on the Nitric Oxide Produced by Endothelial Nitric Oxide Synthase.

In this study, we report a remarkably active CeVO4 nanozyme that functionally mimics cytochrome c oxidase (CcO), the terminal enzyme in the respiratory electron transport chain, by catalyzing a four-electron reduction of dioxygen to water. The nanozyme catalyzes the reaction by using cytochrome c (Cyt c), the biological electron donor for CcO, at physiologically relevant pH. The CcO activity of the CeVO4 nanozymes depends on the relative ratio of surface Ce3+ /Ce4+ ions, the presence of V5+ and the surface-Cyt c interactions. The complete reduction of oxygen to water takes place without release of any partially reduced oxygen species (PROS) such as superoxide, peroxide and hydroxyl radicals.

Namrata Singh

Papers

A Redox Modulatory Mn3O4 Nanozyme with Multi-Enzyme Activity Provides Efficient Cytoprotection to Human Cells in a Parkinson's Disease Model

A manganese oxide nanozyme prevents the oxidative damage of biomolecules without affecting the endogenous antioxidant system.

A Cerium Vanadate Nanozyme with Specific Superoxide Dismutase Activity Regulates Mitochondrial Function and ATP Synthesis in Neuronal Cells

Manganese-Based Nanozymes: Multienzyme Redox Activity and Effect on the Nitric Oxide Produced by Endothelial Nitric Oxide Synthase.

CeVO 4 Nanozymes Catalyze the Reduction of Dioxygen to Water without Releasing Partially Reduced Oxygen Species