Purification and Properties of Cold-active Metalloprotease from Curtobacterium luteum and Effect of Culture Conditions on Production

Abstract:

Curtobacterium luteum, a gram-positive psychrotrophic bacterium, secreting an extracellular protease was isolated from the soil of Gangotri glacier, Western Himalaya. The maximum enzyme production was achieved when isolate was grown in a pH-neutral medium containing skim milk at 15oC over 120 hour. The metal ions such as Zn2+ and Cr2+ enhanced enzyme production. The specific activity of purified enzyme was 8090 u/mg after 34.1 fold purification. The 115 kD enzyme was a metalloprotease (activity inhibited by EDTA and EGTA) and showed maximum activity at 20oC and pH 7. The enzyme was active over a broad pH range and retained 84% of its original activity between pH 6–8. There was no loss in enzyme activity when exposed for 3 hours at 4oC-20oC. However, lost 65% of activity at 30oC, and was almost inactivated at 50oC, but was resistant to repeated freezing and thawing. The enzyme activity was stimulated by manganese ions; however, it was inactivated by copper ions.