Purification and Partial Characterization of Alkaline Protease from Bacillus subtilis NRRL B-3749 (BIOTECH 1679)

An alkaline protease was isolated form the culture supernatant of Bacillus subtilis NRRL B-3749 (BIOTECH 1679). Purification of the crude enzyme filtrate involved ammonium sulfate precipitation, desalting by dialysis and ion-exchange chromatography using carboxy methyl cellulose (CMC). The purified alkaline protease fraction showed a single protein band after polyacrylamide gel electrophoresis (PAGE). Enzyme assay of the single protoen band showed alkaline protease activity. The purified enzyme had a molecular weight of 31,000 daltons using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Chemical characterization of the purified alkaline protease showed optimum proteolytic activity at 55°C and pH 9.0 using casein as substrate. It was stable at pH 6.5 to 10.5 and at temperatures 50°C and below. Key words: Bacillus subtilis , alkaline protease, gel chromatography, enzyme