Enzymatic Cascade Reactions inside Polymeric Nanocontainers: A Means to Combat Oxidative Stress

Abstract

Oxidative stress, which is primarily due to an imbalance in reactive oxygen species, such as superoxide radicals, peroxynitrite, or hydrogen peroxide, represents a significant initiator in pathological conditions that range from arthritis to cancer. Herein we introduce the concept of enzymatic cascade reactions inside polymeric nanocontainers as an effective means to detect and combat superoxide radicals. By simultaneously encapsulating a set of enzymes that act in tandem inside the cavities of polymeric nanovesicles and by reconstituting channel proteins in their membranes, an efficient catalytic system was formed, as demonstrated by fluorescence correlation spectroscopy and fluorescence cross-correlation spectroscopy. Superoxide dismutase and lactoperoxidase were selected as a model to highlight the combination of enzymes. These were shown to participate in sequential reactions in situ in the nanovesicle cavity, transforming superoxide radicals to molecular oxygen and water and, therefore, mimicking their natural behavior. A channel protein, outer membrane protein F, facilitated the diffusion of lactoperoxidase substrate/products and dramatically increased the penetration of superoxide radicals through the polymer membrane, as established by activity assays. The system remained active after uptake by THP-1 cells, thus behaving as an artificial organelle and exemplifying an effective approach to enzyme therapy.

Polymersomes containing quantum dots for cellular imaging

“…Ps remained stable inside cells after having escaped from endosomes without releasing the encapsulated compounds. 24,25,45 Therefore, it is possible that in our experiments, QDs were partially degraded inside the Ps without being released. Due to the stability of Ps in cellular conditions, degradation of QDs and background signals were dramatically lower in Ps than in liposomes.…”

“…22,23 Numerous types of Ps have been proposed as nanocarriers for encapsulating small drug molecules, larger therapeutic proteins, and other types of nanoparticles. [24][25][26] Reports on the use of Ps as carriers of water-soluble nanoparticles such as QDs are scarce. 27 Recently, Ps prepared from amphiphilic poly (D,Llactide)-poly(2-methacryloyl-oxy-ethylphosphorylcholine) (PLA-PMPC) diblock copolymers have been used to encapsulate phosphorylcholine-coated QDs.…”

“…28 Molecular brightness measurements have been used to calculate the number of fluorescent molecules encapsulated inside each polymersome. 25,38 We estimated the average number of QDs per polymersome based on brightness obtained from the count rate per molecule (cpm, kHz). We calculated an average of four QDs/polymersome based on the brightness of free QDs and QDs encapsulated in Ps.…”

Artificial Organelles: Reactions inside Protein–Polymer Supramolecular Assemblies

“…Upon uptake by THP-1/HeLa cells, this simple peroxisome efficiently detoxified both O 2 -and H 2 O 2 . [32] Development of the artificial peroxisome using tandem enzymes found in the natural peroxisomes opens a form a mechanically more stable, but still biocompatible, polymer compartment. The role of the polymersome is to simultaneously protect the encapsulated active compounds from environmental conditions (e.g.…”

Nucleocytoplasmic Transport: A Paradigm for Molecular Logistics in Artificial Systems

“…[22] As a step towards multicompartmentalization, two different nanoreactors bearing two different enzymes could be encapsulated into a larger polymersome [23] and used to control a cascade reaction across the compartmental boundaries. [24] We refer the interested reader to Table 1 Ta ble 1. Selection of polymersomes that harbor transmembrane channels and their cargoes.…”