Mechanism of Low Temperature Inactivation of a Heat-Resistant Bacterial Protease in Milk

Abstract

The heat-resistant protease MC60 underwent a conformational transition at 55 C which resulted in loss of enzyme activity. Under certain conditions, this loss was irreversible. In the absence of sufficient substrate, the transition rendered the altered protease susceptible to autolysis. Heating high concentrations of enzyme in milk resulted in loss of enzyme activity by the formation of an enzymecasein aggregate and by autolysis. Experiments with radioactively-labeled protease showed that low concentrations of enzyme in milk were inactivated only by aggregation with casein micelles. Aggregation and autolysis in milk were prevented by high concentrations of sodium chloride or urea, and aggregation was reversed with 6 M urea at elevated temperatures. The possibility of inactivating heat-resistant enzymes at low temperatures (55 C) may be applicable to other food systems.