Cross-linked esterase aggregates (CLEAs) using nanoparticles as immobilization matrix

Abstract

The present study focusses on the enhancement of the catalytic activity and stability of an acetylesterase enzyme isolated from Staphylococcus spp. as Cross-Linked Enzyme Aggregates (CLEAs). The various parameters governing the activity of CLEAs were optimized. The magnetite and graphene oxide nanoparticles were successfully prepared via the chemical co-precipitation and Hummer's method, respectively. These nanoparticles supported the preparation as magnetite nanoparticle-supported cross-Linked Enzyme Aggregates (MGNP-CLEAs) and graphene oxide-supported Cross-Linked Enzyme Aggregates (GO-CLEAs). The activity and stability of these immobilized CLEAs were compared with the free enzyme at various temperature, pH, and organic solvents along with its storage stability and reusability. The immobilized preparations were analyzed by Scanning Electron Microscopy (SEM) and Fourier Transform Infrared spectroscopy (FT-IR) techniques. Acetylesterase precipitated with 60% saturated ammonium sulfate salt (SAS) solution and cross-linked with 100 mM glutaraldehyde for 4 h at 30 °C was found to be optimal to produce CLEAs with highest activity recovery of 99.8%. The optimal pH at 8.0 and temperature at 30 °C remained the same for both the free and immobilized enzyme, respectively. Storage stability significantly improved for the immobilized enzyme as compared to free enzyme. SEM showed type-I aggregate and FT-IR revealed the successful immobilization of the enzyme. MGNP-CLEAs were found to have better activity and stability in comparison to other immobilized preparations.

Keywords:

• Acetylesterase
• cross-linked enzyme aggregates
• enhanced stability
• graphene oxide
• immobilization
• magnetite nanoparticle

Acknowledgments

One of the authors, Nithyakalyani. D thanks the University Grants Commission (UGC), Government of India for the CSIR-UGC (NET) fellowship. Dr.PG thanks Department of Biotechnology (DBT) and Department of Science and Technology (DST), Government of India for their continued support to the laboratory. This manuscript has not been published elsewhere and has not been submitted simultaneously for publication elsewhere.