EXTRACTION, PARTIAL PURIFICATION AND CHARACTERIZATION OF KINEMA PROTEASE PRODUCED FROM WHITE SOYABEAN

Abstract

Protease constitute a large group of highly demanded enzymes having huge application in various industries. This study was carried out with an aim to extract protease from kinema prepared from white soybean, partial purify the extracted protease by ammonium sulphate precipitation followed by dialysis and to determine general characteristics of the purified protease. The kinema, which has highest proteolysis activity after completion of three days fermentation was taken for the study. The data were analyzed by using one way ANOVA using Genstat at 5% level of significance level. About 2.5 fold of purification with overall recovery of 71.7% was achieved after precipitation with ammonium sulphate at 30-70% saturation followed by dialysis of crude extracted protease. The specific activity was found to be 7.67 U/mg/min. The enzyme was found active over various natural substrates; showing highest activity on casein, followed by BSA, gelatin, hemoglobin and whey protein. Metal ions such as Iron, Mercury and aluminum shows the inhibition effect whereas presence of sodium, cobalt, zinc, copper, calcium, potassium shows the activation effect on protease activity. Furthermore, the enzyme activity was highly inhibited with EDTA and then PMSF at 5mM concentration, suggesting the protease from kinema was a metal dependent serine protease.