A fibrinolytic protease AfeE from Streptomyces sp. CC5, with potent thrombolytic activity in a mouse model

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• 作者：Zhibin Sun^a ; Pingping Liu^a ; Guangyan Cheng^b ; Biying Zhang^a ; Weiliang Dong^a ; Xingli Su^b ; Yan Huang^a ; Zhongli Cui^a ; ^{czl@njau.edu.cn" class="auth_mail" title="E-mail the corresponding author} ; Yi Kong^b ; ^{yikong668@163.com" class="auth_mail" title="E-mail the corresponding author}
• 关键词：Fibrinolytic protease ; Streptomyces sp. CC5 ; Thrombolytic activity
• 刊名：International Journal of Biological Macromolecules
• 出版年：2016
• 出版时间：April 2016
• 年：2016
• 卷：85
• 期：Complete
• 页码：346-354
• 全文大小：1452 K

文摘

Fibrinolytic proteases have potential applications in cardiovascular disease therapy. A novel fibrinolytic protease, AfeE, with strong thrombolytic activity was purified from Streptomyces sp. CC5. AfeE displayed maximum activity at 40 °C in the pH range of 7.0–12.0. It was strongly inhibited by serine protease inhibitor phenylmethanesulfonylfluoride, soybean trypsin inhibitor, tosyl-l-lysine chloromethyl ketone and tosyl-l-phenylalanine chloromethyl ketone. The activity of the enzyme was partially inhibited by Cu²⁺, Co²⁺ and Zn²⁺. AfeE exhibited higher substrate specificity for fibrin than fibrinogen, which has rarely been reported in fibrinolytic enzymes. AfeE also showed high thrombolytic activity in a carrageenan-induced mouse tail thrombosis model. AfeE prolonged prothrombin time, activated partial thromboplastin time, and thrombin time in rat blood. A bleeding time assay revealed that AfeE did not prolong bleeding time in mice at a dose of 1 mg/kg. No acute cytotoxicity was observed for AfeE at 320 μg/well in human umbilical vein endothelial cells. The afeE gene was cloned from the genome of Streptomyces sp. CC5. Full-length AFE-CC5E contained 434 amino acids and was processed into a mature form consisting 284 amino acids by posttranslational modification, as revealed by high-resolution mass spectrometry analysis. These results indicate that AfeE is a prospective candidate for antithrombotic drug development.