RESUMO
Aims@#Microorganisms play a vital role in the breakdown of natural organic compounds and are valuable objects for worldwide enzyme production. The aim of this study was to identify favorable production conditions for Bacillus amyloliquefaciens D19 protease, followed by the purification and chemical characterization of this novel enzyme to assess its potential applications in various fields.@*Methodology and results@#In this study, favorable conditions of protease production from B. amyloliquefaciens D19 were determined using a medium containing soluble starch (1.5%), earthworm extract (1.0%), yeast extract (0.5%), NaCl (1.0%), at pH 7.0-8.0, 37 °C for 36 h with 150 rpm shaking condition. The protease was purified and had a molecular weight of about 23 kDa. The optimum condition for casein hydrolysis was at 40 °C and pH 6.5-7.0 in the presence of 1.0 mM Na+ or 5.0 mM Zn2+. The enzymatic activity was maintained at 75-100% at 30-50 °C and in pH 6.0-10.0. The values of Vmax and KM were also determined as 1547 U/mg and 6.33 mg/mL, respectively.@*Conclusion, significance and impact of study@#The identified optimal conditions will serve as the foundation for the production of the 23 kDa B. amyloliquefaciens D19 protease, one of the smallest proteases within the Bacillus genus. Moreover, its notable heat resistance, broad pH tolerance, high substrate catalysis and moderate substrate binding affinity make this enzyme a promising candidate for various applications in the food-feed and brewing industries.