Purification, Crystallization and Some Properties of Low-Temperature-Active Intracellular Proteinase from Streptococcus lactis

Abstract

An intracellular proteinase has been separated in crystalline from Streptococcus lactis (IAM 1198), and some enzymatic properties of it were studied. The procedure resulting in 45.4-fold purification of the proteinase included protamine sulfate fractionation, DEAE-52 column chromatography and Sephadex G-50 column chromatography. It was purified to a specific activity of 4.72U/mg protein. The proteinase was freezedried, and then dispersed in deionized water containing ethanol to make a proteinase suspension, which was gradually cooled. The enzyme was allowed to be crystallized after storage. Approximately 2% of the original enzyme activity was recovered in the purified fraction. The crystalline proteinase was shown to be homogeneous by polyacrylamide gel electrophoresis. The molecular weight was about 12, 500 and the absorption maximum spectrum was at 278nm and the value of E ^1% _1cm at 278nm was 10.83. The optimum activity was at pH5.5 and 6°C.