Self-Assembled Nano-Bioreactor from Block Ionomers with Elevated and Stabilized Enzymatic Function

Abstract

Core-cross-linked polyion complex (PIC) micelles entrapping trypsin in the core were prepared by mixing trypsin and poly(ethylene glycol)-block-poly(alpha,beta-aspartic acid) in aqueous medium, followed by the introduction of glutaraldehyde cross-linkages. Trypsin incorporated into the core-cross-linked micelles showed high storage stabilities, and the initial enzymatic activity of trypsin was maintained even after standing for one week at ambient temperature. Further, stable compartmentalization of trypsin into the core-cross-linked micelles led to a unique modulation in the enzymatic functions including an improved thermal tolerability with an increased maximum reaction rate compared to native trypsin.

Complex coacervate core micelles

“…Besides co-micellization with DNA and RNA, other biopolymers, such as proteins, peptides, and enzymes, may also be encapsulated to preserve and/or promote stability and function, as was demonstrated for KALA, [88,140] lysozyme, [121,122,[141][142][143][144][145][146]150] trypsin, [148,149] myoglobin, [104] and BSA [62]. Circular dichroism (CD) experiments on lysozyme-containing C3Ms demonstrated that the secondary structure of free and C3M incorporated lysozyme is identical [144].…”

“…Circular dichroism (CD) experiments on lysozyme-containing C3Ms demonstrated that the secondary structure of free and C3M incorporated lysozyme is identical [144]. As is the case for DNA-containing C3Ms, complexation with oppositely charged copolymers, typically improves protein/enzyme stability with respect to ionic strength, dilution, denaturation by urea, [149] protease attack, [104] which may be further enhanced by core crosslinking [122,148,149] or end-functionalization of the core-block of the copolymer with hydrophobic groups, such as pyrenyl [121]. Trypsin and lysozyme activity toward small substrates (i.e., substrates that can diffuse into the micellar core) is promoted when embedded in C3Ms; i.e.…”

“…Trypsin and lysozyme activity toward small substrates (i.e., substrates that can diffuse into the micellar core) is promoted when embedded in C3Ms; i.e. higher initial catalysis [148,149] and overall reaction [122,144,149] rates are observed due to substrate accumulation within the C3M [122,144]. Discrete on-off control of the elevated initial velocity of lysozyme activity in C3Ms of lysozyme/PEO-b-PAsp could be achieved by applying voltages over 63 V cm − 1 , presumably due to micellar dissociation or a change in core microenvironment [145].…”

Cross-linked antioxidant nanozymes for improved delivery to CNS

“…This approach was previously used by us and others to immobilize various biopolymers in micelle-like, core-shell particles with high colloidal stability. 25-29 In particular, Kataoka and coworkers formulated positively-charged lysozyme with negatively-charged poly(ethylene glycol)-poly(α,β-aspartic acid) block copolymer. Noteworthy, the activity of lysozyme incorporated in such complex was higher compared to the native enzyme 26,27 and it was hypothesized that the elevated enzymatic activity is a result of substrate concentration in the polyion complexes species 26 and/or alterations in binding mode between lysozyme and substrate.…”

Towards new functional nanostructures for medical imaging

“…40,[110][111][112][113][114][115] Transient, metastable nanostructures can be constructed from biological or exogenous molecules that will self-assemble in vivo for short periods of time, depending on the local environment or external application of energy. This concept could be expanded further such that nanoscale reaction vessels [116][117][118] for the fabrication of contrast nanostructures could be assembled within the biological system. The use of smaller molecular components in vivo that can assemble directly at the target site may allow larger nanostructures to be built at the target without prior elimination by the MPS.…”