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Purification and characterization of a fibrinolytic enzyme from Streptomyces sp. XZNUM 00004

Author:
Ju, Xiuyun, Cao, Xiaoying, Sun, Yong, Wang, Zhe, Cao, Chengliang, Liu, Jinjuan, Jiang, Jihong
Source:
World journal of microbiology & biotechnology 2012 v.28 no.7 pp. 2479-2486
ISSN:
0959-3993
Subject:
molecular weight, fibrinogen, polyacrylamide gel electrophoresis, EDTA (chelating agent), isoelectric point, amino acids, gel chromatography, pH, anion exchange, Streptomyces, temperature, ammonium sulfate, fibrin, enzyme activity, ethylene glycol tetraacetic acid
Abstract:
A fibrinolytic enzyme (SFE1) from Streptomyces sp. XZNUM 00004 was purified to electrophoretic homogeneity with the methods including ammonium sulfate precipitation, polyacrylamide gel, DEAE-Sepharose Fast Flow anion exchange and gel-filtration chromatography. The molecular weight of SFE1 was estimated to be 20 kDa by SDS-PAGE, fibrin zymography, and gel filtration chromatography. The isoelectric point was 4.9. K m and V max values were 0.96 mg/ml and 181.8 unit/ml, respectively. It was very stable at pH 5.0–8.0 and below 65 °C. The optimum pH for enzyme activity was 7.8. The optimum temperature was 35 °C. The fibrinolytic activity of SFE1 was enhanced by Na+, K+, Mn2+, Mg2+, Zn2+ and Co2+. Conversely, Cu2+ showed strong inhibition. Furthermore, the fibrinolytic activity was strongly inhibited by PMSF, and partly inhibited by EDTA and EGTA. SFE1 rapidly hydrolyzed the Aα-chain of fibrinogen, followed by the Bβ-chain and finally the γ-chain. The first 15 amino acids of the N-terminal sequence were APITLSQGHVDVVDI. Additionally, SFE1 directly digested fibrin and not by plasminogen activators in vitro. SFE1 can be further developed as a potential candidate for thrombolytic therapy.
Agid:
396983