Purification and characterization of an extracellular protease produced by psychrotolerant Clostridium sp. LP3 from lake sediment of Leh, India

An anaerobic, proteolytic bacterium isolated from lake sediments of Leh, India, was characterized with respect to morphology, biochemical characteristics, and 16S rRNA sequence and was identified as Clostridium species, with closest similarity to Clostridium subterminale. Isolate LP3 was psychrophilic, forming maximum cell mass between 10 and 20 degrees C, and produced extracellular protease. Growth was observed in the pH range of 7.0-8.5, with optimum at pH 7.5. Protease was purified 62.4-fold with a total yield of 17.5%. The effects of temperature, pH, and salt concentration on enzyme activity were studied. Protease was found to be a serine-type metallo-enzyme, active in a broad range of pHs. It was thermolabile and resistant to sodium dodecyl sulfate. Enzyme kinetics showed a tendency to increase Km with an increase in temperature for casein substrate.