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KMID : 0545120150250091449
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Journal of Microbiology and Biotechnology
2015 Volume.25 No. 9 p.1449 ~ p.1459
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Purification and Biochemical Characterization of a Novel Fibrinolytic Enzyme from Streptomyces sp. P3
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Guangyan Cheng
Liying He
Zhibin Sun
Zhongli Cui
Yingxiang Du
Yi Kong
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Abstract
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A novel proteolytic enzyme with fibrinolytic activity, FSP3, was purified from the recently isolated Streptomyces sp. P3, which is a novel bacterial strain isolated from soil. FSP3 was purified to electrophoretic homogeneity by ammonium sulfate precipitation, anion exchange, and gel filtration. FSP3 is considered to be a single peptide chain with a molecular mass of 44 kDa. The maximum activity of the enzyme was observed at 50°C and pH 6.5, and the enzyme was stable between pH 6 and 8 and below 40°C. In a fibrin plate assay, FSP3 showed more potent fibrinolytic activity than urokinase, which is a clinical thrombolytic agent acting as a plasminogen activitor. The activity was strongly inhibited by the serine protease inhibitor PMSF, indicating that it is a serine protease. Additionally, metal ions showed different effects on the activity. It was significantly suppressed by Mg2+ and Ca2+ and completely inhibited by Cu2+, but slightly enhanced by Fe2+. According to LC-MS/MS results, its partial amino acid sequences are significantly dissimilar from those of previously reported fibrinolytic enzymes. The sequence of a DNA fragment encoding FSP3 contained an open reading frame of 1287 base pairs encoding 428 amino acids. FSP3 is a bifunctional enzyme in nature. It hydrolyzes the fibrin directly and activates plasminogen, which may reduce the occurrence of side effects. These results suggest that FSP3 is a novel serine protease with potential applications in thrombolytic therapy.
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KEYWORD
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Streptomyces, Fibrinolytic activity, Puri?cation, Serine protease
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