Partial purification and characterization of an extracellular cold-active protease from Flavobacterium azizsancarii, a novel Antarctic isolate.

Cold-active enzymes exhibit high catalytic efficiency at low temperatures, making them valuable biocatalysts for energy-efficient and environmentally friendly industrial processes. The enzymatic activity of bacteria that thrive in low-temperature environments has attracted the attention of researchers. In this study, an extracellular cold-active protease produced by Flavobacterium azizsancarii (F. azizsancarii), a novel Antarctic isolate, was partially purified by ammonium sulfate precipitation and dialysis. The enzyme showed maximal activity toward casein at pH 8.0 and 20 °C. The effects of various metal ions on protease activity indicated that Ca²⁺ did not result in a significant change in activity, whereas Fe²⁺, Zn²⁺, Mg²⁺, and Mn²⁺ significantly inhibited proteolytic function. The enzyme was inhibited by organic solvents, including ethanol, methanol, acetone, toluene, and benzene, and was significantly and partially inhibited by PMSF, suggesting the possible presence of a serine-type protease component. These findings demonstrate that F. azizsancarii produces a cold-active alkaline protease with promising biotechnological potential, particularly for food processing, detergent formulation, and protein hydrolysate production.