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| Purification and Biochemical Properties of Protease from White Leg Shrimp Litopenaeus vannamei |
| PAN Bin,XIE Yue-xia,DAI Jun-yong |
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Abstract The protease was extracted and purified from heads of white leg shrimp Litopenaeus vannamei by Tris-HCl buffer solution, gel-filtertion with Sephadex G-100, and ion-exchange with DEAE-Sepharose F.F. The enzymological nature was studied with biochemical and electrophoretic techniques. The enzyme was purified to an electrophoretically homogenous state, reaching 14.3-fold purification with a yield of 31.9%. The extracted protease showed a single band in its relative molecular weight at 25 kD based on SDS-PAGE.The optimum pH was 7.0 for the enzymatic reaction and the optimum temperature was 60 ℃ for casein as the substrate This protease showed a good thermal stability below 50 ℃, and more than 60% of the activity still remained when the protease was cultured at 50 ℃ for 1 h. But the enzyme became inactivated rapidly at the temperature above 60 ℃, and only 4.7% of protease activity remained at 60 ℃ for 1 h. The protease activity was found to be inhibited by 10 mmol/L of EDTA(at a rate of 84%), and Fe~(2+)(53%), Ba~(2+)(43%)and Zn~(2+)(38%). However, 10 mmol/L of Ca~(2+) increased the enzyme activity,while 10 mmol/L of Mg~(2+) and Cu~(2+) had less effect on the protease activity. It is suggested that the protease from the shrimp be a type of metalloenzyme.
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Received: 25 December 2009
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