Unkeshwar hot spring of Maharashtra was explored for the isolation of industrially important efficient thermostable protease producers. These isolates were identified based on their morphological characters, microscopic features, enzymatic profile and biochemical characteristics as Bacillus sphaericus APP21, Staphylococcus auricularis APP32, Microbacterium sp. APP41, and Kurthia sp. APP42 from Unkeshwar hot spring. Thermostable protease production was carried out using casein-yeast extract medium at 50 oC. Maximum specific activity of partially purified thermostable protease was recorded from Bacillus sphaericus APP21 (447.75 U/mg). Molecular weight of partially purified proteases from these isolates has been determined in the range of 14-29 KDa. These proteases have showed excellent thermostability in the range of temperature 45 to 65 oC. These proteases were catalytically active in the range of pH 7 to 9 and belonged to the class serine protease. Km and Vmax values were found in the range 5.26-8.34 mg/mL and 446-1380 U/mg respectively. The divalent cations Mg2+ and Mn2+ enhanced the catalytic efficiency of these proteases. These proteases showed excellent catalytic efficiency in presence of selected solvents, surfactants and detergents. These properties make them excellent candidates to use in detergent formulation, food processing, and in many biotechnologically important industries where proteolysis is needed at an elevated range of temperature.
Thermostable protease; Thermostability; Unkeshwar hot spring; SDS-PAGE; Km; Vmax