Studies on Immobilization of Cutinases from Thermobifida fusca on Glutaraldehyde Activated Chitosan Beads

Aims: To evaluate and optimize the activity and stability performance of two recombinant cutinases of Thermobifida fusca, Cut1 and Cut2 on glutaraldehyde activated chitosan beads.

Place and Duration of Study: Biochemical Engineering Laboratory, Department of Biotechnology, Indian Institute of Technology Guwahati, Assam India. Experiment conducted as a partial fulfillment to PhD degree from December, 2010 to January, 2014.

Methodology: Purified cutinase were immobilized on chitosan beads by covalently coupling with glutaraldehyde. The biophysical properties of immobilized cutinase was analysed by FTIR, FESEM and the operational stability and activity of the immobilized cutinase was studied at different pH and temperature.

Results: The optimal immobilization was achieved with 3% (v/v) glutaraldehyde activation and coupling pH of 8.5. Under this condition, 74% and 71% of immobilization was achieved for Cut1 and Cut2, respectively. Immobilized cutinase showed optimal activity at pH 8 with optimal functional range of pH 7.5 to 9 and 55ºC with operational stability in the range of 45ºC to 70ºC. The reusability and storage stability was found to be 80% after 10 reuse cycles and 50% after 13 days, respectively as compared to its initial activity. There was no loss in activity even after repeated freeze drying.

Conclusion: the cutinase immobilized on glutaraldehyde activated chitosan beads demonstrated better operational stability in comparison to free cutinase, showing chitosan as a potential support in the enzyme immobilization technology for industrial applications of T. fusca cutinases.