Enhanced Thermostability and Anticancer Activity in Breast Cancer Cells of Laccase Immobilized on Pluronic-Stabilized Nanoparticles

Abstract

Laccases are multi-copper oxidase enzymes having widespread applications in various biotechnological fields. However, low stability of free enzymes restricts their industrial use. Development of effective methods to preserve and even increase the enzymatic activity is critical to maximize their use, though this remains a challenge. In the present study we immobilized Trametes versicolor laccase on pH-responsive (and charge-switchable) Pluronic-stabilized silver nanoparticles (AgNPsTrp). Our results demonstrate that colloidal stabilization of AgNPsTrp with the amphiphilic copolymer Pluronic F127 enhances enzyme activity (AgNPsTrpF1 + Lac6) by changing the active site microenvironment, which is confirmed by circular dichroism (CD) and fluorescence spectroscopy. Detailed kinetic and thermodynamic studies reveal a facile strategy to improve the protein quality by lowering the activation energy and expanding the temperature window for substrate hydrolysis. The immobilized nanocomposite did not show any change in flow behavior which indirectly suggests that the enzyme stability is maintained, and the enzyme did not aggregate or unfold upon immobilization. Finally, assessing the anticancer efficacy of this nanocomposite in breast cancer MCF-7 cells shows the inhibition of cell proliferation through β-estradiol degradation and cells apoptosis. To understand the molecular mechanism involved in this process, semi qRT-PCR experiments were performed, which indicated significant decrease in the mRNA levels of anti-apoptotic genes, for example, BCL-2 and NF-kβ, and increase in the mRNA level of pro-apoptotic genes like p53 in treated cells, compared to control. Overall, this study offers a completely new strategy for tailoring nano-bio-interfaces with improved activity and stability of laccase.

Balancing loading, cellular uptake, and toxicity of gelatin‐pluronic nanocomposite for drug delivery: Influence of HLB of pluronic

“…24,25 Pluronic as a stabilizing agent can successfully be used to overcome this limitation as shown by us and several other groups. 12,26 Apart from biological stability, the other most important parameter affecting the efficacy of a drug delivery system is its uptake by the target cells. 27 Extensive research in last two decades or so have revealed that properties of nanoparticles such as bulk composition, surface parameters (charge, hydrophobicity, elasticity, and targeting ligand), size, and morphology can influence their uptake by the target cells.…”

“…However, one of the major limitations of using protein nanoparticles as drug delivery system is their hydrophobicity resulting into aggregation and/or opsonization and in turn low circulation half‐life 24,25 . Pluronic as a stabilizing agent can successfully be used to overcome this limitation as shown by us and several other groups 12,26 . Apart from biological stability, the other most important parameter affecting the efficacy of a drug delivery system is its uptake by the target cells 27 .…”

Compare the physicochemical properties of engineered polymer-functionalized silver nanoparticles and evaluate their biological properties against Porphyromonas gingivalis

“…The comparative analysis of the hydrated size of AgNPs-1/2 and P-AgNPs-1/2 shows the degree of increase in hydrated particle size is not consistent with the molecular weight of polymer. Chauhan et al found that this may be relate to the e ciency of polymer adsorption on the surface of AgNPs [35]. Hence, the increase in the nano size of P-SH5000 and PVP indicates that they have strong adsorption to AgNPs.…”

In vitro cytotoxicity of Reishi mushroom extract against two human cell lines

“…Recent study of [19] which the inhibition cell growth against MCF-7 cell line by Coriolus versicolor extract. Moreover, the percentages of MCF7 cell proliferation inhibition by Xanthium strumarium extract have maximum compared with doxorubicin [20].…”