Cross-Linked Enzyme Aggregates of Feruloyl Esterase Preparations from Thermothelomyces thermophila and Talaromyces wortmannii

Abstract

Cross-linked enzyme aggregates (CLEA ®) technology is a well-established method in the current literature for the low-cost and effective immobilization of several enzymes. The main advantage of this particular method is the simplicity of the process, since it consists of only two steps. However, CLEA immobilization must be carefully designed for each desired enzyme, since the optimum conditions for enzymes can vary significantly, according to their physicochemical properties. In the present study, an investigation of the optimum CLEA immobilization conditions was carried out for eight feruloyl esterase preparations. Feruloyl esterases are a very important enzyme group in the valorization of lignocellulosic biomass, since they act in a synergistic way with other enzymes for the breakdown of plant biomass. Specifically, we investigated the type and concentration of precipitant and the crosslinker concentration, for retaining optimal activity. FAE68 was found to be the most promising enzyme for CLEA immobilization, since in this case, the maximum retained activity, over 98%, was observed. Subsequently, we examined the operational stability and the stability in organic solvents for the obtained CLEA preparations, as well as their structure. Overall, our results support that the maximum activity retaining and the stability properties of the final CLEAs can vary greatly in different FAE preparations. Nevertheless, some of the examined FAEs show a significant potential for further applications in harsh industrial conditions.

Biochemical and Structural Characterization of Cross-Linked Enzyme Aggregates (CLEAs) of Organic Solvent Tolerant Protease

“…The loss of enzyme activity after each cycle has been linked to the leaching of the enzyme or mechanical stress during centrifugation causing the CLEA pellet to clump [36]. Our SEM images showed that CLEA-elastase exhibits a type 2 aggregate and this coheres with studies suggesting that type 2 aggregates are often formed by hydrophilic enzymes [45]. The DLS analysis provided insight into the diameter size of the aggregate in aqueous media, where the diameter of CLEA-elastase appeared to be significantly larger than that of a free elastase.…”

Crosslinked Enzyme Aggregates (CLEAs) of Laccases from Pleurotus citrinopileatus Induced in Olive Oil Mill Wastewater (OOMW)

“…Among the various immobilization methods described in the literature, crosslinked enzyme aggregates (CLEAs) technology is one of the most prominent [ 20 ]. Many different enzymes have been immobilized in CLEA form, among them esterases [ 21 ], lipases [ 22 ], cellulases [ 23 ], and xylanases [ 24 ], which were used for various bioconversion applications, such as the production of cosmeceuticals [ 21 ], nutraceuticals [ 24 ], or even biofuels [ 22 ]. The advantages of CLEA technology include the simplicity of the method, the low overall cost of the procedure, and the fact that there is no need for solid support or dedicated specialized equipment.…”

Long chain capsaicin analogues synthetized by CALB-CLEAs show cytotoxicity on glioblastoma cell lines

“…The morphology of CLEAs depends on enzyme type, precipitant agent and the microenvironment of the enzyme when cross-linking (Zerva et al 2018 ). SEM micrographies revealed the formation of clusters with irregular, cluster shapes and low porosity for CALB-CLEAs (Fig.…”