A novel neutral protease from thermophilic Bacillus strain HUTBS62

A novel neutral highly thermostable protease was detected in the culture medium of thermophilic Bacillus strain HUTBS62 isolated from hot-spring located near to the Dead Sea, Jordan. The enzyme was purified by precipitation with 55-60% ammonium sulfate, gel filtration on Sephadex G-100 and DEAE ion exchange chromatography. The enzyme was purified 53-fold with 2% yield. The optimum pH and temperature for catalytic activity of protease was pH 6.8 and 80oC, respectively, and 31% activity of protease remained even after heat treatment at 100oC for 60 min. The relative activity of the enzyme was highly stable (90%) at 50oC for 2 h. The half-life of the enzyme at 90oC, 80oC and 70oC was estimated to be 3, 4 and 6 h, respectively. The activation energy of denaturation of purified enzyme was 21.7 kJmol-1. Iron, sodium, calcium, and manganese increased protease activity. On the other hand, magnesium, cobalt and zinc variably decreased the residual activity. But cadmium and copper drastically inhibited the enzyme activity. The enzymatic activity was highly stable in the presence of 1 and 2 mM EDTA at pH 6.8 and 80oC. The neutral protease therefore could be defined as a highly thermostable with new properties make the present enzyme applicable for many biotechnological purposes.