Aug , 2021, Volume : 2 Article : 7
Nanozymes – A breakthrough in enzyme research
Author : Prithusayak Mondal
Cite this article as:
Mondal, P. (2021) Nanozymes – A breakthrough in enzyme research. Food and Scientific Reports. 2 (8) 39-41.
ABSTRACT
The drawbacks of natural enzymes, such as low stability, high cost, and difficult storage, compelled the researchers to think about promising alternatives. In the past few decades, a lot of artificial enzymes have been fabricated to mimic the structures and catalytic actions of natural enzymes. The escalating growth in the field of nanoscience and nanotechnology has recently been able to formulate nanozymes, engineered nanomaterials with enzyme-like characteristics, those are considered as next-generation of artificial enzymes due to their unique attributes. Varieties of nanozymes have been fabricated till now based on their configurations, such as metal-based, metal oxide-based and carbon-based. In this article, recent applications of nanozymes in diverse biological fields have been discussed. The current challenges of nanozyme research and possible future directions are also addressed.
Keywords: Nanomaterials, Artificial enzyme, Nanotechnology, Enzyme mimics, Catalytic activities
Nanozymes are engineered nanomaterials with enzyme mimicking capabilities. Natural enzymes play vital roles in the biological systems. However, some intrinsic drawbacks, such as ease of denaturation, laborious preparation, high cost, and difficulty of recycling, have limited their practical applications. Intensive efforts have been made to fabricate artificial enzymes to overcome these drawbacks of natural enzymes. As an emerging research area of artificial enzymes, nanozymes, the catalytic nanomaterials with enzyme-like characteristics, have attracted enormous attentions among the researchers. Nanozymes are built of nanostructured materials (organic or inorganic) having unique physicochemical features owing to their typical nanoscale factors, such as size, morphology and surface characteristics, those cater excellent strategies for adjusting their activities. They catalyze biochemical reactions of the substrates for natural enzymes under physiological conditions including mild temperature and physiological pH, and the nature of catalysis is totally intrinsic i.e. devoid of interference of additional natural enzymes or chemical catalysts. Like natural enzymes, they obey the same enzyme kinetics (e.g. Michaelis–Menten equation) and also have active centers or electron-transport structures useful for catalytic functions. Nanozymes can be suggested as an enzyme alternatives that outpaced natural enzymes in numerous aspects such as stability, low cost, and ease of production.
These are basically nanomaterials capable to mimic the activities of various natural enzymes, such as oxidases (oxidase, glucose oxidase, sulphite oxidase and ascorbate oxidase), peroxidases (peroxidase, glutathione peroxidase, horseradish peroxidase and haloperoxidase), catalase, superoxide dismutase and others (nuclease, phosphatase etc.). The intrinsic enzyme-like catalytic activities of nanozymes are generally thought to be originated from the ions or atoms present both on the surface and inside the core. Thus, the composition and arrangement of atoms present in nanozymes are vital factors in determining their catalytic activity, however, other influencing factors such as size, morphology, surface coating and modification, pH, light and temperature can play roles. Based on the type of configuration, nanozymes can be grouped into three broad categories: metal oxide-based (Fe3O4, CeO2, MnO2, V2O5 etc.), metal-based (Au, Pt, Au-Pt, Ag-Pd, Ag-Au etc.), and carbon-based (CNT, fullerene and graphene) nanozymes.
Modern applications of nanozymes
We are surrounded by various nanosized materials, and our body is composed of and controlled by numerous biological nanomachines. Nanozymes have emerged as useful tools in numerous fields.
Biosensors
Of late, horseradish peroxidase (HRP) capable of catalyzing the oxidation of various organic substrates by hydrogen peroxide (H2O2) has become one of the most commonly used enzymes for the construction of biosensors. Since H2O2 is a by-product of numerous biological pathways and potential contaminant of several industrial products, its detection is vital in the field of biosensors. The peroxidase-like catalytic activity of Fe3O4 magnetic nanoparticles (MNPs) has made them robust nanozyme for H2O2 detection using colorimetric signal readout strategy, based on a redox reaction between HRP and colorimetric substrates. Similarly, Peroxidase-like nanozymes coupled with glucose oxidase (GOx) have been frequently employed in the construction of glucose biosensors. A nanocomposite was developed through entrapment of MNPs and GOx in the pores of mesoporous carbon, where GOx generated H2O2, which was readily reduced to H2O with the electrocatalytic reduction mediated by MNPs. A unique cholesterol detection system was developed using MNPs incorporated in the wall of mesoporous silica, forming magnetic mesoporous silica (MMS), and cholesterol oxidases. In this multicatalyst system, cholesterol oxidase immobilized in the MMS initiated a reaction with cholesterol to generate H2O2, which subsequently activated MNPs in the mesocellular silica pores to convert a colorimetric substrate into a colored product.
Immunoassays
In the research of biological sciences, immunoassays are routine biochemical tests for tracking various molecules using antibody (usually) or antigen (sometimes). The most commonly used enzymes in immunoassay include HRP (horseradish peroxidase) and ALP (alkaline phosphatase). Chitosan-based MNPs (CS-MNPs) were explored as an alternative for HRP and were able to detect mouse immunoglobulin G (IgG) and carcinoembryonic antigen (CEA). Chitosan modified on the surface of MNPs prevented aggregation of MNPs in aqueous solutions, and amino groups in the chitosan catered a convenient site for covalent linking of antibodies to MNPs, thereby replacing the linkage of HRP-conjugated antibodies to CS-MNP-conjugated antibodies in the immunoassay. Utilizing the enzyme-mimic activity of ferric nanocore located in ferritins, antigen-down type and sandwich type immunoassays were conducted taking avidin and nitrated human ceruloplasmin respectively as target molecules. In these assays, ferritin oxidizes p-hydroxyphenylpropionic acid (p-HPPA), a common fluorescence substrate for peroxidase, in the presence of H2O2 to generate a fluorescent product, and appeared as more thermally stable and pH-tolerant in comparison with HRP.
Disease theranostics
Disease theranostics involve disease diagnosis and therapies, where nanozymes have been started exploring very recently. For determination of both benign and cancerous tumors, nanozymes have made successful applications. A hybrid nanozyme was fabricated through immobilization of Au nanoparticles on periodic mesoporous silica-coated nanosized reduced graphene oxide conjugated with folic acid, which displayed an exceptional peroxidase-like activity and was employed as a selective, quantitative, and rapid colorimetric detection probe for detection of cancer cells. When the nanoparticles were conjugated with folic acid, they attached to folate receptors on the tumor cells due to high expression of folate receptors on the tumor cell surface. Poly(acrylic acid) polymer coated CeO2 nanoparticles (nanoceria) act as oxidase mimic and can directly oxidize colorimetric substrate to a colored product without H2O2 and make colorimetric detection possible. This nanoceria as superoxide dismutase (SOD) mimic may prevent retinal degeneration by inhibiting the production of reactive oxygen intermediates (ROIs). Ferucarbotran, a commercialized superparamagnetic iron oxide (SPIO) nanozyme, was found to expedite cell cycle progression and stimulate cell growth in human mesenchymal stem cells (hMSCs) by lowering intracellular H2O2 with the help of intrinsic peroxidase-like activity.
Environmental engineering
Nanozyme-based techniques have been used in several environmental situations like contaminants detection and removal for the purpose of food safety, pollution control and public health management. In dairy sector, melamine is a toxic contaminant that can be detected by a simple and rapid colorimetric method using peroxidase-mimicking activity of MNP nanozymes. Melamine inhibits the catalytic oxidation of colorimetric substrates (ABTS) by MNPs in the presence of H2O2, because it competitively reacts with H2O2, forming an additional compound which ultimately alters the intensity of ABTS colour signal dependent on the concentration of melamine. This colorimetric method may facilitate safety assurance through detection of melamine concentration in dairy products. A combination of nanocomposite-entrapping MNPs and oxidase in mesoporous carbon facilitated amperometric detection of several phenolic compounds such as phenol, cresol, and cathechol. A common industrial dye, Methylene blue (MB) can be removed at room temperature utilizing the peroxidase-mimicking activity of Fe3O4 MNPs and oxidizing power of H2O2.
Conclusion and future perspectives
Nanozymes, because of their excellent stability, high tunability and robust enzyme-mimicking characteristics, are foreseen as a replacement for natural enzymes in the coming days. However, multi-functionality of nanozymes propels a critical question of regulating their off-target activities. Biosafety issues also of developed nanozymes must be thoroughly investigated as most of the reported nanozymes lack both short-term and long-term biosafety evaluation. Biodegradability and biocompatibility must be of first priority while choosing nanomaterials for nanozyme development. More conclusive reports are needed from the interdisciplinary fields of biology, nanotechnology, medicine, and material science, for wide commercial and practical applications of nanozymes. Many more pertinent techniques for effective management of nanozyme activities in vivo are of utmost need in present situation. Scientists are optimistic that nanozymes in future will certainly open new horizons in the multidisciplinary fields of research.
Abbreviations: ABTS 2,2`-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), ALP Alkaline phosphatase, CEA Carcinoembryonic antigen, CeO2 Cerium oxide, GOx Glucose oxidase, H2O2 Hydrogen peroxide, hMSCs Human mesenchymal stem cells, HRP Horseradish peroxidase, IgG Immunoglobulin G, MB Methylene blue, MMS Magnetic mesoporous silica, MNPs Magnetic nanoparticles, p-HPPA p-hydroxyphenylpropionic acid, ROIs Reactive oxygen intermediates, SOD Superoxide dismutase, SPIO Superparamagnetic iron oxide.
Further Reading
Gao, L., Fan, K., Yan, X. (2017). Iron oxide nanozyme: a multifunctional enzyme mimetic for biomedical applications. Theranostics, 7(13), 3207–3227. https://doi.org/10.7150/thno.19738
Gao, L., Zhuang, J., Nie, L., Zhang, J., Zhang, Y., Gu, N., Wang, T., Feng, J., Yang, D., Perrett, S., Yan, X. (2007). Intrinsic peroxidase-like activity of ferromagnetic nanoparticles. Nature Nanotechnology, 2(9), 577–583. https://doi.org/10.1038/nnano.2007.260
Meng, X.Q., Fan, K.L. (2018). Application of nanozymes in disease diagnosis. Progress in Biochemistry and Biophysics, 45(2), 218–236. https://doi.org/10.16476/j.pibb.2018.0039
Shin, H.Y., Park, T.J., Kim, M.I. (2015). Recent Research Trends and Future Prospects in Nanozymes. Journal of Nanomaterials, vol. 2015, Article ID 756278, 11 pages. https://doi.org/10.1155/2015/756278
Wang, X., Hu, Y., Wei, H. (2016). Nanozymes in bionanotechnology: from sensingto therapeutics and beyond. Inorganic Chemistry Frontiers, 3, 41. https://doi.org/10.1039/c5qi00240k
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