A Novel Serine Protease Produced by Marine Isolated Strain of Vibrio anguillarum

A serine protease was purified to homogeniety from the culture supernatant of a marine bacterium, Vibrio anguillarum by a 4-step procedure, ammonium sulfate precipitation, G-100 gel filtration, and two rounds of phenyl-Sepharose chromatography. The molecular weight of the enzyme was estimated to be 30kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration. The enzyme activity was inhibited by diisopropyl fluorophosphate, phenylmethyl sulfonyl fluoride, Leupeptin, and Chymostatin. The optimum temperature was 37°C and the optimum pH of the enzyme was 9.0.Mg2+ and Ca2+ at 10mM enhanced the enzyme activity by 161 and 124%, and stabilized the activity when exposed between 15 to 42°C. The enzyme hydrolyzed strongly butyloxycarbonyl-leucyl-seryl-threonyl-arginyl-4-methylcoumaryl-7-amide among 17 peptydyl-4-methylcoumaryl-7-amides, indicating narrow substrate specificity. The alkaline serine enzyme shows similar substrate specificity to a known metalloprotease. However, this novel serine protease did not show anticoagulation activity as detected in the metalloprotease, suggesting a different function in this bacterium.

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