Extracellular Heat-Resistant Proteases of Psychrotrophic Pseudomonads

Several bacterial isolates from raw milk produced proteases. Most of such 28 isolates were gram-negative rods which were oxidase- and catalase-positive. All the isolates grew at temperatures in the range of 0-35°C, but failed to grow at 37°C. Nineteen of these isolates were tentatively assigned to genus Pseudomonas , and were used in the present investigation. Extracellular proteases from these psychrotrophic pseudomonads were heat-resistant, being able to retain partial activity even after heat-treatment at 120°C for 10 min. Milk proteins were preferred substrates by these proteases although some also hydrolysed bovine serum albumin, hemoglobin and ovalbumin. The optimum pH for the maximum activity was between pH 7.2 and 7.4. Divalent metal ions like Cu²⁺, Co²⁺, Hg²⁺, and Zn²⁺ were inhibitory to protease activity while Ca²⁺, Mg²⁺, and Mn²⁺ had little or no inhibitory effect on the proteases. Induced levels of protease production were observed when cultures were grown in minimal media containing either casein or nonfat dried milk powder. Glucose, citrate and lactose repressed enzyme synthesis in a minmal salts medium containing either casein or nonfat dried milk powder. Protease activity was also detected in cultures grown in minimal medium containing glutamine. Proteases from different isolates varied in their molecular weights.