School of Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, 250355, China.
School of Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, 250355, China.
Protein Expr Purif. 2020 Oct;174:105680. doi: 10.1016/j.pep.2020.105680. Epub 2020 Jun 2.
Developing an effective fibrinolytic drug for treating thrombolysis with minimal undesirable side effects is of great importance. In the current study, an optimum solvent was selected for the extraction of fibrinolytic active components. Furthermore, a strong fibrinolytic enzyme named WPI01 was purified from Whitmania pigra Whitman through various chromatographic steps. WPI01 has a molecular mass of 27044.297 Da, and the N-terminal 8 amino acid sequence was determined as VVGGVEAR. WPI01 was stable within the pH range of 6.0-10.0 and with maximum fibrinolytic activity at 40 °C and a pH of 8.0. At 500 U/mL, WPI01 induced 50.59% blood clot reduction in vitro within 6 h, which was higher than that induced by urokinase at 1000 U/mL. In an analysis of the plasminogen activator activity, WPI01 produced obvious halos on heated and unheated fibrin plates, suggesting that WPI01 may not only act as a plasminogen activator but also degrade fibrin clots directly, and more study is needed to support this. In conclusion, WPI01 is obviously different from known fibrinolytic enzymes in terms of substrate specificity and fibrinolytic mode of action, suggesting that it is a novel fibrinolytic enzyme with potential applications in the treatment and prevention of thrombosis.
开发一种能够有效治疗血栓溶解且副作用最小的纤维蛋白溶解药物非常重要。在本研究中，选择了一种最佳溶剂来提取纤维蛋白溶解活性成分。此外，通过多种色谱步骤，从Whitmania pigra Whitman 中纯化出一种名为 WPI01 的强纤维蛋白溶解酶。WPI01 的分子量为 27044.297 Da，N 端 8 个氨基酸序列被确定为 VVGGVEAR。WPI01 在 pH 6.0-10.0 范围内稳定，在 40°C 和 pH 8.0 时具有最大的纤维蛋白溶解活性。在 500 U/mL 时，WPI01 在体外 6 h 内诱导 50.59%的血凝块减少，高于 1000 U/mL 的尿激酶诱导的减少。在纤溶酶原激活剂活性分析中，WPI01 在加热和未加热的纤维蛋白平板上产生明显的晕圈，表明 WPI01 不仅可以作为纤溶酶原激活剂，还可以直接降解纤维蛋白凝块，需要更多的研究来支持这一点。总之，WPI01 在底物特异性和纤维蛋白溶解作用方式方面与已知的纤维蛋白溶解酶明显不同，表明它是一种新型的纤维蛋白溶解酶，具有在血栓形成的治疗和预防方面的潜在应用价值。
