Immobilization of Alkaline Protease From Bacillus brevis Using Ca-Alginate Entrapment Strategy for Improved Catalytic Stability, Silver Recovery, and Dehairing Potentialities

摘要

Proteases are one of the most important biocatalysts with an industrial perspective due to their high production capability, cost-effective, and eco-friendly nature. In this study, an alkaline protease produced by Bacillus brevis (228.31 +/- 6.2 U/mL) in liquid-state fermentation at pH 7.0 and 47 degrees C was further enhanced by optimizing various physical parameters. The maximum protease activity (631.09 +/- 3.7 U/mL) was recorded at 45 degrees C, pH 8.0, and using 3 mL of inoculum size after 72 h. The optimally-produced alkaline protease was immobilized on Ca-alginate beads, and the process was optimized using a central composite design-based response surface methodology. The maximum immobilization yield (> 70%) was achieved using a 2-3.0% gelling agent (Na-alginate) and 2.5-3.0% binder (CaCl2), with 400-600 mg/L of protease concentration. Characterization revealed that immobilization improves the pH and thermal stability of protease, as the maximum activity was recorded at pH 10 and 65 degrees C. The kinetic studies of protease revealed higher V-max (454.5 U/mL) and lower K-m (0.09 mu M) values after Ca-alginate immobilization as compared with the free enzyme (V-max 333.3 U/mL and K-m 0.16 mu M). Ca-alginate immobilized protease also possessed good recyclability potential in several consecutive substrate hydrolysis experiments. By applying on goatskin and X-ray film, the immobilized biocatalyst showed improved activity than the free enzyme, revealing its potential biotechnological applications.