Encapsulation of Spherical Cross-Linked Phenylalanine Ammonia Lyase Aggregates in Mesoporous Biosilica

Abstract

Cross-linked enzyme aggregates (CLEAs) have recently emerged as a promising tool for enzyme immobilization because of their simplicity and low cost. However, a lack of good size and morphological control over the as-prepared CLEAs has limited their practical applications. For example, the prepared CLEAs exhibit amorphous large clusters that would cause significant mass-transfer limitations, which lead to a low catalytic efficiency. Here, inspired by biomineralized core-shell structures in nature, we develop a novel mesoporous spherical CLEA with a biosilica shell by using phenylalanine ammonia lyase based on CaCO microtemplates and biomimetic mineralization. The resultant CLEAs exhibited a spherical structure with good monodispersity instead of the amorphous clusters of conventional CLEAs and showed activity higher than that of conventional CLEAs. Moreover, the thermostability, tolerance against denaturants, and mechanical stability of the spherical CLEAs with a biosilica shell were enhanced significantly compared with those of conventional CLEAs. In particular, the spherical CLEAs with a biosilica shell retained 70% of their original activity after 13 cycles, whereas the conventional CLEAs retained only 35% of their original activity. This approach could be an efficient strategy for improving the catalytic properties of CLEAs.

Bio-based and cost effective method for phenolic compounds removal using cross-linked enzyme aggregates

“…The resulting CLEAs enhanced the enzyme stability to elevated temperature, increased storage stability while allowing their separation in a simple and easy manner. The same observations were concluded in the synthesis of spherical CLEAs with biosilica shell which turned out stable against denaturants, while maintaining 70% of its activity after 13 cycles (Cui et al, 2017). In addition, the combination of traditional techniques with CLEAs technology may prove useful, as in the case of the adsorbed cross-linked phenylalanine ammonia lyase aggregate on the crude-pored microspherical silica core.…”

Production of Trans-Cinnamic Acid by Immobilization of the Bambusa oldhamii BoPAL1 and BoPAL2 Phenylalanine Ammonia-Lyases on Electrospun Nanofibers

“…In free enzymes, there was no significant difference between BoPAL proteins expressed in E. coli and P. pastoris. However, the Km value of immobilized BoPAL proteins were slightly higher than that of free BoPAL proteins, which might be due to the cross-linking effect that limits the permeability of the matrix [39,40]. In addition, the high porosity of PAL/nylon 6/CS nanofibers allowed the matrix to readily access the active site of PAL.…”

“…The optimum pH of immobilized pBoPAL1 compared to free enzyme was changed from pH 9.0 to pH 8.5 (Figure 7C). It might be caused by the ionization changes of acidic and amino acid side chains in the surrounding environment of the active site of PAL [39,40]. The optimum pH working conditions for eBoPAL1/2 and pBoPAL1/2 were assessed at the pH from 5 to 11.…”

Biopolymer-Based Multilayer Capsules and Beads Made via Templating: Advantages, Hurdles and Perspectives

“…CLEAs are popular materials in the field of biocatalysis as reusable catalysts [194], and the control of final aggregate size and porosity are important for the relative catalytic activity and practical use in industrial catalysis. Hard-templating may provide an appropriate means of controlling these properties [195][196][197].…”