Covalent immobilization of β-1,4-glucosidase from Agaricus arvensis onto functionalized silicon oxide nanoparticles

Abstract

An efficient β-1,4-glucosidase (BGL) secreting strain, Agaricus arvensis, was isolated and identified. The relative molecular weight of the purified A. arvensis BGL was 98 kDa, as determined by sodium dodecylsulfate polyacrylamide gel electrophoresis, or 780 kDa by size exclusion chromatography, indicating that the enzyme is an octamer. Using a crude enzyme preparation, A. arvensis BGL was covalently immobilized onto functionalized silicon oxide nanoparticles with an immobilization efficiency of 158%. The apparent V (max) (k (cat)) values of free and immobilized BGL under standard assay conditions were 3,028 U mg protein(-1) (4,945 s(-1)) and 3,347 U mg protein(-1) (5,466 s(-1)), respectively. The immobilized BGL showed a higher optimum temperature and improved thermostability as compared to the free enzyme. The half-life at 65 °C showed a 288-fold improvement over the free BGL. After 25 cycles, the immobilized enzyme still retained 95% of the original activity, thus demonstrating its prospects for commercial applications. High specific activity, high immobilization efficiency, improved stability, and reusability of A. arvensis BGL make this enzyme of potential interest in a number of industrial applications.

Immobilization of β-glucosidase onto silicon oxide nanoparticles and augment of phenolics in sugarcane juice

“…This increase of optimum temperature range may be due to conformational stability of the enzyme (Wang et al 2011). Singh et al (2011) also observed that the optimum temperature of the free b-glucosidase, isolated from Agaricus arvensis was 65°C, but the b-glucosidase immobilized on SiO 2 nanoparticles had maximal activity at 70°C. They also observed that for the free enzyme, increase in temperature resulted in a sharp decrease in the relative activity of the free enzyme.…”

“…They also observed that for the free enzyme, increase in temperature resulted in a sharp decrease in the relative activity of the free enzyme. In contrast, the immobilized enzyme showed no such tendency and retained 82 % of its optimum activity even at 85°C (Singh et al 2011). …”

“…4b). It has also reported about 288-fold increase in the thermal stability at 65°C, after immobilization of b-glucosidase on SiO 2 nanoparticles (Singh et al 2011). This transition in stability of immobilized b-glucosidase may be attributed to the multiple covalent bonds that alter the conformational flexibility and thermal vibrations, thus preventing the immobilized protein from unfolding and denaturing (Mateo et al 2000;Wong et al 2009).…”

“…Activated SiO 2 nanoparticles are supposed to contain -OH group. The amino group of lysine present on the surface of the b-glucosidase reacts with the activated silicon oxide nanoparticles to form the covalent bond at multiple points (Singh et al 2011). A high surface-tovolume ratio of these nanosized particles induces the increase of specific surface area available to biomolecules which allows easier enzyme anchoring and catalytic reaction.…”

“…The washed carrier was directly used for the determination of activity and stability. The supernatant was also checked for any remaining b-glucosidase activity (Singh et al 2011).…”