Stabilization of cutinase by covalent attachment on magnetic nanoparticles and improvement of its catalytic activity by ultrasonication

Abstract

This paper reports on stabilization of serine cutinase activity by immobilizing it through cross linking with glutaraldehyde on magnetic nanoparticles (Fe-NPs) and intensification of catalytic activity by ultrasonic treatment. The optimum parameters were cross linking with 10.52 mM glutaraldehyde for 90 min using 1:2 (w/w) ratio of enzyme:Fe-NPs. The characterization of cutinase-Fe-NPs was done by different instrumental analysis. Ultrasonic power showed a beneficial effect on the activity of free and immobilized cutinase at 5.76 and 7.63 W, respectively, after 12 min. Immobilization and ultrasonic treatment led to increments in kinetic parameters (K_m and V_max) along with noticeable changes in the secondary structural fractions of cutinase. Cutinase-Fe-NPs showed augmented pH (4-8) and thermal stability (40-60 °C). Considerably higher thermal inactivation kinetic constants (k_d, t_1/2 and D-value) and thermodynamic constants (E_d, ΔH°, ΔG° and ΔS°) highlighted superior thermostability of cutinase-Fe-NPs. Cutinase-Fe-NPs and ultrasound treated cutinase-Fe-NPs retained 61.88% and 38.76% activity during 21-day storage, and 82.82 and 80.69% activity after fifth reusability cycle, respectively.