Production of Alkaline Proteases using Aspergillus sp. Isolated from Injera: RSM-GA Based Process Optimization and Enzyme Kinetics Aspect.

Alkaline proteases are well known to be significant industrial enzymes. This study focused on isolating the fungus producing proteases from, a typical Ethiopian food, Injera. Further, the process optimization for protease production using response surface methodology (RSM) and the characterization of the acquired protease were investigated. The 18S rDNA gene sequence homology of the fungus isolate revealed that it was Aspergillus sp. Further, it was deposited in NCBI GenBank with accession number MK4262821. Using the isolate, owing to maximize the protease production, the independent process parameters, temperature, pH, and sucrose concentration were optimized using RSM followed by a genetic algorithm (GA). Based on the statistical approach by RSM-GA optimization, maximum enzyme activity (166.4221 U/ml) was found at 30.5 °C, pH 8.24, and 0.316% sucrose concentration. Also, the crude cocktail of enzymes acquired from optimal condition was partially purified using ammonium which showed that the increased activity by 1.96-fold. Considerably, the partially purified enzyme exhibited stable performance during the temperature range 30-60 °C, pH range 7-10, and NaCl concentration of 0.5-2 mM. Also, the antioxidant activity, degree hydrolysis for the protein, Michaelis-Menten (M-M) kinetic parameters, and activation energy were determined for the obtained enzyme cocktail. It showed that the M-M kinetic parameters, Km (5.54 mg/ml), and Vmax (24.44 mg/ml min) values were observed. Using Arrhenius law, the value of activation energy for the enzyme cocktail was determined as 32.42 kJ/mol.