Purification and biochemical characterization of a trypsin-like enzyme present in the sperm of the rock shrimp, <i>Rhynchocinetes typus</i>
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Date
2001
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Abstract
The aim of the present work was to isolate, purify and characterize a trypsin-like enzyme from the sperm of the rock shrimp, Rhynchocinetes typus. Sperm proteins were extracted with 1 mM HCl in 10% glycerol at pH 3.0. Purification of the trypsin-like substance was effected by affinity chromatography using SBTI-agarose, yielding a specific activity on BAEE as substrate of 787 U/mg, with a recovery rate of 34%. Enzymatic activity was maximal at 27 degreesC, pH of 8.0, 50 mM Ca2+ and 30 mM Mg2+. One hundred percent inhibition of enzymatic activity was obtained at 0.05 mM Zn2+. Kinetic analysis showed that the K-M on BAEE as substrate at pH 8.0 was 2.5 x 10(-5) M and the V-MAX reached was 198 U. It was also found that the enzyme had a substrate inhibition at concentrations higher than 0.06 mM of BAEE. These findings suggest that thin enzyme has similar characteristics to other trypsin-like enzymes including acrosin.
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Keywords
sperm, trypsin-like, enzyme, shrimp