Protease producing bacteria was isolated from soil of Tatapani area of Mandi district of Himachal Pradesh. These bacteria were screened in skim milk agar medium using skim milk as the substrate. The highest clear zone producing bacterial isolate P5 was selected for further optimization studies. The isolate was identified as Bacillus subtilis based on morphological, biochemical and molecular characterizations. The isolate was able to grow under alkaline conditions at pH 9.0 and a temperature of 50°C. 1.9 fold purification of enzyme following ammonium sulphate precipitation and DEAE-cellulose chromatography was achieved. The molecular weight of the enzyme was estimated to be approximately 29 kDa as shown by electrophoresis. Interestingly ca²⁺ (5mM) activated enzyme activity, while Mg^2+, Mn²⁺ Co²⁺ moderately activated enzyme activity, where as Fe²⁺ and Cu²⁺ inhibited the activity of enzyme.