Abstract
In recent years, natural deep eutectic solvents (NADESs) have gained increasing attention as promising nontoxic solvents for biotechnological applications, due to their compatibility with enzymes and ability to enhance their activity. Betaine-based NADESs at a concentration of 25 wt % in a buffered aqueous solution were used as media to inhibit thermal inactivation of POXA1b laccase and its five variants when incubated at 70 and 90 °C. All the tested laccases showed higher residual activity when incubated in NADES solutions, with a further enhancement achieved also for the most thermostable variant. Furthermore, the residual activity of laccases in the presence of NADESs showed a clear advantage over the use of NADESs’ individual components. Molecular docking simulations were performed to understand the role of NADESs in the stabilization of laccases toward thermal inactivation, evaluating the interaction between each enzyme and NADESs’ individual components. A correlation within the binding energies between laccases and NADES components and the stabilization of the enzymes was demonstrated. These findings establish the possibility of preincubating enzymes in NADESs as a facile and cost-effective solution to inhibit thermal inactivation of enzymes when exposed to high temperatures. This computer-aided approach can assist the tailoring of NADES composition for every enzyme of interest.
Highlights
Laccases are widespread multicopper oxidases catalyzing the oxidation of a broad range of phenolic and nonphenolic substrates with the concomitant reduction of oxygen to water, the only reaction byproduct.[1]
natural deep eutectic solvents (NADESs) are based on betaine as the hydrogen bond acceptor (HBA) and sorbitol, recent results obtained with T. versicolor laccase,[20] five betaine- xylitol, glycerol, ethylene glycol, or erythritol as hydrogen bond donor (HBD), in the based NADES media were chosen and tested at 25 wt % aqueous dilution for their thermostabilizing effect on P
Similar beneficial effects due to incubation in the same NADES solution were observed for EV1 and EV5, while the 25 wt % 2Bet:1Xyl NADES solution was the most advantageous NADES media for EV3 and EV4. This is in accordance with Delorme et al.,[20] which described an increase in T. versicolor laccase residual activity of nearly 40% and 50% when incubated in 25 wt % 1Bet:3Sor and 25 wt % 2Bet:1Xyl NADES solutions, respectively. These findings demonstrates that the NADES incubation is effective on various laccases, including ones already endowed with high thermal stability, such as EV5
Summary
Laccases are widespread multicopper oxidases catalyzing the oxidation of a broad range of phenolic and nonphenolic substrates with the concomitant reduction of oxygen to water, the only reaction byproduct.[1]. One of the challenges in the implementation of laccases in industrial processes is their ability to remain active for a longer time or survive harsh operative conditions, resulting in the need of reengineer enzymes to fine-tune their properties toward the end application, boosting both productive efficiency and enzyme performance.[4,5] Recently, flexible surface loops have been identified as potential targets to improve enzyme thermostability by applying modifications aimed at their stiffening.[6,7] An additional and complementing approach to boost enzyme thermostability consists of exploring microbial diversity by culture mining or metagenomic approaches[8] searching for extremozymes. The immobilization of enzymes onto solid supports has become a key enabling technology to implement postsynthesis their exploitation in industrial processes, promoting enzyme reusability and recovery,[9,10] while enhancing their thermostability.[11]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
