Development and characterization of cross-linked enzyme aggregates of thermotolerant alkaline protease from Bacillus licheniformis

An alkaline protease was produced by B. licheniformis with 132.43 +/- 3.4 U/mL activity in LSF which was further enhanced by optimizing culture conditions. The optimum enzyme activity (148.9 +/- 4.1 U/mL) was harvested at pH 7.5; temperature, 40 degrees C and inoculum, 15 mL after 48 h incubation. Alkaline protease was immobilization by forming cross linked enzyme aggregates (CLEAs) and the processes of CLEAs formation was also optimized. The protease CLEAs developed using 80% ammonium sulfate, 65 mM glutaraldehyde and 0.11 mM BSA showed best activity recovery (39.76%). Free protease and CLEAs were characterized and compared. It was observed that CLEAs of protease exhibited broad pH range with best activity at pH 10. The immobilized protease was also thermo-tolerant with optimum activity at 65 degrees C temperature. The V-max and K-m of protease-CLEAs were 1255 U/mL and 18.97 mu M, respectively as compared to 104.9 U/mL and 29 mu M, respectively for free protease. It was concluded that immobilized enzyme in the form of CLEAs is a valuable catalyst for potential industrial applications. (C) 2018 Elsevier B.V. All rights reserved.