In order to substitute microbial enzyme for the plantoriginated proteolytic enzyme, papain, which has been traditionally used for chillproofing beer, 148 protease-producing strains were isolated from natural sources such as soil and sewage. Among them a strain designated S-103 showed the highest chillproofing activity and was partially identified as $\mbox{\underline{Bacillus sp}}$.
S-103 was shaked in a modified nutrient broth(PH 7.0)containing 10g of beef extract, 7g of peptone, 3g of Nacl in 1000 ml of tap water under the culture temperature of 30 ℃. The exzyme was harvested after 72 hours of fermentation and was subjected to salting-out with saturated ammonium sulfate. After dialysis against phosphate buffer solution(0.1 M, Ph 7.0) a gel-filtration on Sephadex G-75 was followed. Through the procedure the enzyme activity was found to be increased by 60 folds of the original culture with recovery rate of 25%.
The optimum temperature and PH for the enzyme production were 25-30 ℃ and PH 7-8. The enzyme was stable at PH range of 7-9 and at temperature below 35 ℃ and a complete in activation was observed in the presence of Hg,Ag,Cd,Zn,Cu and Fe. However, the presence of Ca, especially in the form of $CaCl_2$, enhanced the proteolytic activity of the enzyme strongly.
The chillproofing activity in the beer of the partially purified enzyme from S-103 was compared with a commercial brand chillproofer, Protesal, and found to be almost equivalent in the strength each other.
