Characterization and heat inactivation of extracellular proteinase from Pseudomonas fluorescens TR2

Extracellular proteinase from Pseudomonas fluorescens was purified to SDS-P AGE homogeneity. The enzyme appeared to be a neutral endopeptidase containi ng zinc. It had a molecular weight of 43.5 KDa with K-m 0.07 mM and Vmax 20 .2 units/mu g protein, for casein as substrate. It was heat resistant at te mperatures above 80 degrees C in the presence of calcium, while it appeared to be sensitive at lower temperatures. The enzyme was autolyzed at 60 degr ees C, while it was not in the presence of casein. In the latter case, prot einase hydrolyzed the casein. Proteinase studied hydrolyzed both beta- and alpha(s)- caseins of cow's and ewe's milks in the order beta->alpha(s). Ewe 's caseins appeared to be more resistant to proteinase hydrolysis than cow' s caseins.