Purification, characterization and crystallization of an extracellular alkaline protease from Aspergillus nidulans HA-10.

Aspergillus nidulans is a highly potent fungus used in the production of alkaline protease. Extracellular alkaline protease was purified from A. nidulans in a two-step procedure involving ammonium sulphate precipitation and Sephadex G-100 column chromatography. The molecular mass of the enzyme was determined to be 42 kDa by SDS-PAGE. The enzyme activity was also analyzed by zymogram with gelatin. The enzyme was more stable over a wide range of pH (6-10) and the temperatures up to 50 degrees C. It showed optimum enzyme activity at pH 8.0 and a temperature of 35 degrees C. The protease enzyme was completely inhibited by the serine protease inhibitor of phenylmethylsulfonyl fluoride (PMSF). The crystallization of the purified enzyme was performed by hanging drop vapour diffusion method using PEG 6000 as the precipitant. The micro crystals occurred in 40% of PEG 6,000.