Stabilization of a psychrotrophic Pseudomonas protease by calcium against thermal inactivation in milk at ultrahigh temperature

Abstract

The heat-stable extracellular protease of Pseudomonas sp. (isolate MC60) was investigated. Heat resistance of the enzyme in milk at sterilization temperature was dependent on the presence of Ca2+. The half-life of the enzyme at ultrahigh temperature (149.degree. C) in skim milk or milk-salts buffer with Ca2+ was approximately 7.0 s. Treatment of milk with chelators completely removed the heat-stabilizing effect of milk. The enzyme was partially purified by ammonium sulfate precipitation and column chromatography on Sephadex G-100. At 21.degree. C the enzyme retained > 85% activity after exposure to pH values between 5-10. Enzyme activity was reduced by metal chelating agents. Both Ca2+ and Zn2+ were required for optimal enzyme activity. MW was estimated at 48,000 by gel filtration.