Biochemical analysis of a fibrinolytic enzyme purified from <Emphasis Type="Italic">Bacillus subtilis</Emphasis> strain A1 |
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Authors: | Won Sik Yeo Min Jeong Seo Min Jeong Kim Hye Hyeon Lee Byoung Won Kang Jeong Uck Park Yung Hyun Choi Yong Kee Jeong |
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Institution: | (1) Department of Biological Pharmacy, Shanghai Institute of Pharmaceutical Industry, 200040 Shanghai, China;(2) Department of Life Science and Biopharmacy, Shenyang Pharmaceutical University, 110016 Shenyang, China;(3) Shanghai Health Creation Center of Biopharmaceutical R&D, 201203 Shanghai, China; |
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Abstract: | A fibrinolytic enzyme from Bacillus subtilis strain Al was purified by chromatographic methods, including DEAE Sephadex A-50 column chromatography and Sephadex G-50 column
gel filtration. The purified enzyme consisted of a monomeric subunit and was estimated to be approximately 28 kDa in size
by SDS-PAGE. The specific activity of the fibrinolytic enzyme was 1632-fold higher than that of the crude enzyme extract.
The fibrinolytic activity of the purified enzyme was approximately 0.62 and 1.33 U/ml in plasminogen-free and plasminogen-rich
fibrin plates, respectively. Protease inhibitors PMSF, DIFP, chymostatin, and TPCK reduced the fibrinolytic activity of the
enzyme to 13.7, 35.7, 15.7, and 23.3%, respectively. This result suggests that the enzyme purified from B. subtilis strain Al was a chymotrypsin-like serine protease. In addition, the optimum temperature and pH range of the fibrinolytic
enzyme were 50°C and 6.0–10.0, respectively. The N-terminal amino acid sequence of the purified enzyme was identified as Q-T-G-G-S-I-I-D-P-I-N-G-Y-N,
which was highly distinguished from other known fibrinolytic enzymes. Thus, these results suggest a fibrinolytic enzyme as
a novel thrombolytic agent from B. subtilis strain Al. |
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