Immobilisation of cellulase on vermiculite and the effects on enzymatic kinetics and thermodynamics

Abstract

In this work, we attached cellulase onto vermiculite by adsorption and crosslinking via glutaraldehyde, and examined the properties of the immobilised enzyme. The results indicated that immobilisation was achived with 35% yield. Reusability of immobilised enzyme was determined as 18 times with 40% residual activity. Even after 100 min incubation with substrate at 50 °C, 6 and 9 cycles, 56% and 30% of activity of the immobilised enzyme was retained, respectively. Optimum pH for free cellulase was 5, while that for immobilised cellulase was 7, however, optimum temperature didn't change both free and immobilised cellulase. The Michaelis-Menten constant (Km) were found as 0.194 mg mL−1 and 0.025 mg mL−1 for free and vermiculite immobilised cellulase,respectively. The maximum reaction rate (Vmax) for the free and vermiculite immobilised cellulase were calculated as 7.42 U/min and 12.5 U/min,respectively. Furthermore, the other enzymatic kinetic parameters (kcat, and kcat/Km) suggested the affinity and specificity of vermiculite immobilised cellulase to the substrate was 1.64 and 1.14 times improved, respectively. The immobilised enzyme had a lower activation energy (8.73, kJ/mol) than the free enzyme (9.73,kJ/mol). Calculated thermodynamic parameters (ΔH≠, ∆G≠∆G≠E-S, ∆G≠E-T) (kJ/mol) indicated that the immobilised enzyme performs biocatalysis more conveniently than the free enzyme.