Purification and characterization of protease produced by Aspergillus wentti isolated from Korean traditional Meju

摘要

The protease produced by a newly isolated Aspergillus wentti from Korean traditional Meju was purified and characterized. The optimal medium composition and culture conditions for maximum protease production were; bran : 1 percent glucose solution = 1 : 1, pH 9.0, 30 deg C, and 4 days of fermentation. Protease was purified by QAE-Sephadex, SP-Sephadex ion exchange chromatography and Sephadex G-100 chromatography. The specific activity and the purification fold of the purified enzyme were 213 unit/mg protein and 27.3, respectively. The molecular weight of purified protease was found to be 32 kDa by SDS-PAGE. Km and Vmax value's for hammastein milk casein were 3.049 X 10~(-4) M and 151.1 #mu#g/min, respectively. Kinetic parameters showed that the enzyme has higher affinity to casein than isolated soybean protein, hemoglobin and bovine serum albumin. Optimal pH and temperature for reaction of the purified enzyme were 9.0 and 50 deg C, respectively. The enzyme was stable at pH 4.0-11.0, below 40 deg C, and the activity was not stimulated by metal ions. 1 mM phenylmethylsulfonyl fluoride inhibited the enzyme activity by 98.5 percent. It means that the enzyme is one of serine protease.