Engineering Systems with Spatially Separated Enzymes via Dual-Stimuli-Sensitive Properties of Microgels

Abstract

This work examines the adsorption regime and the properties of microgel/enzyme thin films deposited onto conductive graphite-based substrates. The films were formed via two-step sequential adsorption. A temperature- and pH-sensitive poly­(N-isopropylacrylamide)-co-(3-(N,N-dimethylamino)­propylmethacrylamide) microgel (poly­(NIPAM-co-DMAPMA microgel) was adsorbed first, followed by its interaction with the enzymes, choline oxidase (ChO), butyrylcholinesterase (BChE), or mixtures thereof. By temperature-induced stimulating both (i) poly­(NIPAM-co-DMAPMA) microgel adsorption at T > VPTT followed by short washing and drying and then (ii) enzyme loading at T < VPTT, we can effectively control the amount of the microgel adsorbed on a hydrophobic interface as well as the amount and the spatial localization of the enzyme interacted with the microgel film. Depending on the biomolecule size, enzyme molecules can (in the case for ChO) or cannot (in the case for BChE) penetrate into the microgel interior and be localized inside/outside the microgel particles. Different spatial localization, however, does not affect the specific enzymatic responses of ChO or BChE and does not prevent cascade enzymatic reaction involving both BChE and ChO as well. This was shown by the methods of electrochemical impedance spectroscopy (EIS), atomic force microscopy (AFM), and amperometric analysis of enzymatic responses of immobilized enzymes. Thus, a novel simple and fast strategy for physical entrapment of biomolecules by the polymeric matrix was proposed, which can be used for engineering systems with spatially separated enzymes of different types.

Microgel PAINT – nanoscopic polarity imaging of adaptive microgels without covalent labelling

“…It seems that the core collapse induced structural changes in the surroundings of Nile Red labels adsorbed to the shell, a fact that is also highly important for microgels as encapsulating agents, for example, for drug delivery and for their application in catalysis. 43,44 Especially close to the coverslip, we observe a signicantly higher polarity. This increased polarity is reasonable since the coverslips are slightly negatively charged due to our surface cleaning procedure (see the ESI ‡).…”

“…This result has to be taken into account, for example, for the design of microgels as drug delivery systems and for catalysis. 43,44…”

Electroactive, Mediating and Thermosensitive Microgel Useful for Covalent Entrapment of Enzymes and Formation of Sensing Layer in Biosensors

“…In the simplest cases the electroactive species find it harder to penetrate the shrunken polymer layer and to reach the electrode surface. If the enzyme was placed deep in the layer, the shrinking process would depress the electrocatalysis [26][27][28]. A more complicated situation takes place when the polymer layer contains catalytic centers (organic or inorganic).…”

Microgels at fluid-fluid interfaces for food and drinks

“…Drying of microgels onto solid surfaces and the factors that control the structure of the films that form and their barrier properties is an interesting area discussed elsewhere (Horigome and Suzuki, 2012); (Mayarani et al, 2018) . Similarly, there are many studies investigating microgel absorption into gels or release from them (Yaroslavov et al, 2017), plus adsorption of biopolymers and bioactive compounds onto (Kumar and Singh, 2010), (Kureha et al, 2017), (Liu et al, 2014), (Morisada et al, 2010) and absorption into microgels, such as enzymes (Sigolaeva et al, 2014, Sigolaeva et al, 2015, betacarotene (Tan et al, 2014) and DNA (Ozdemir et al, 2006), including the interaction of microgels and their payloads with membranes (Nordstrom et al, 2018); (Mihut et al, 2013) and cells (Vihola et al, 2007). Of particular importance to foods is the ease of enzymatic digestion of materials encapsulated within the microgel particles (Mansson et al, 2013), (Torres et al, 2016).…”