Immobilization and characterization of recombinant Candida antarctica lipase B on poly(glycidyl methacrylate-ter-divinyl benzene-ter-ethylene dimethacrylate) beads, “DILBEADS™TA”

Abstract

Recombinant Candida antarctica lipase B (rCALB) expressed in methylotrophic yeast, Pichia pastoris was covalently immobilized on epoxy-activated macroporous poly(glycidyl methacrylate-ter-divinyl benzene-ter-ethylene dimethacrylate) beads, namely DILBEADS™TA with activity recovery of 39.2%. Tributyrin hydrolysis activity (TBU) of the optimum immobilized enzyme catalyst DILBEADSCB10K was 3380.4 TBU/g dry beads, which is approximately 1.5 times that of Novozym 435 (2592.92 TBU/g dry beads) under similar conditions of TBU activity analysis. Though, the optimum pH for both free as well as immobilized enzyme was found to be 7.0 and optimum temperature was found to be 37°C, the immobilized enzyme catalyst DILBEADSCB10K showed better stability over the pH ranges from 3 to 6 as well as 8 to 10. At extreme acidic and basic pH values, activity of DILBEADSCB10K was 3% more than that of free rCALB enzyme. After 1 h incubation at 50°C, the activity of DILBEADSCB10K was 9% more than that of free rCALB. The improved pH and thermal stability of immobilized enzyme over the free enzyme indicates that immobilization on DILBEADS™TA imparted structural and conformational stability to this enzyme. The immobilization procedure developed is simple, easily reproducible and scalable.

Keywords::

• CALB
• tributyrin
• polyacrylate beads
• epoxy
• kinetic assay
• Novozym 435

Acknowledgments

We would like to thank our biotech lab colleagues and management team of Fermenta Biotech Ltd for their continued support in carrying out this research work.

Declaration of interest: The authors report no declarations of interest. The authors alone are responsible for the content and writing of the paper.