Abstract

The purification and characterization of alkaline protease from a Bacillus strain SAL1,   isolated from tannery waste have been reported in this paper. This protease was purified to homogeneity by the combination of ammonium sulfate precipitation, DEAE sephacryl ion exchange and phenyl sepharose hydrophobic interaction chromatography. The protease was purified up to 11.18 fold and had a specific activity of 4250 PU/mg. The enzyme was a monomeric protease with a relative molecular mass of 27 kDa by SDS-PAGE. Proteolytic activity of the enzyme was detected by gelatin zymography, which gave a very clear protease activity zone on gel. Molecular mass of the purified protease was also determined by matrix-assisted laser desorption ionization-time-off light- mass spectrometry (MALDI-TOF-MS) that corresponded to the mass determined by SDS-PAGE. The enzyme exhibited its optimal activity at 60°C and at pH 9. The enzyme was stable in the pH range of 7.0 – 10.0 and was able to maintain its stability at 50°C for 1 h.

Key words: Alkaline protease, Bacillus subtilis, purification, characterization,tannery waste, DEAE sephacryl, chromatography.