Heilongjiang Provincial Key University Laboratory of Processing Agricultural Products, College of Food and Bioengineering, Qiqihar University , 42, Wenhua Street, Qiqihar 161006, China.
J Agric Food Chem. 2015 Mar 4;63(8):2215-24. doi: 10.1021/jf505717e. Epub 2015 Feb 23.
A novel fibrinolytic enzyme from Cordyceps militaris was produced by submerged culture fermentation, purified, and biochemically characterized. The enzyme was purified to homogeneity, with an overall yield of 4.0% and a specific activity of 1682 U/mg. The molecular weight and pI of the enzyme were 32 kDa and 9.3 ± 0.2, respectively. The optimal pH and temperature of the enzyme were 7.4 and 37 °C, respectively. The enzyme activity was inhibited by Fe(2+), phenylmethane sulfonyl fluoride (PMSF), aprotinin, and pepstatin but not by N-tosyl-L-phenylalanine chloromethyl ketone (TPCK) and ethylenediamine tetracetic acid (EDTA). Three internal peptides of the enzyme, APQALTVAAVGATWAR, EKNVGSTVNLLSYDGNK, and TDATSVLLDGYNVSAVNDLVAK, were obtained. The enzyme could hydrolyze fibrin(ogen) directly and cleave the α-chains more efficiently than β- and γ-chains, suggesting that it is a plasmin like protein. It degraded thrombin, which indicated that it can act as an anticoagulant and prevent thrombosis. Intravascular thrombosis is one of the major reasons of cardiovascular diseases. On the basis of these results, the purified enzyme can be developed as a natural agent for oral fibrinolytic therapy or prevention of thrombosis.
蛹虫草来源的新型纤维蛋白溶酶的发酵生产、分离纯化及生化性质研究。该酶经发酵、提取、分离和纯化后获得电泳纯，总产率为 4.0%，比活力为 1682 U/mg。酶的分子量和等电点分别为 32 kDa 和 9.3 ± 0.2。该酶的最适 pH 和温度分别为 7.4 和 37°C。酶活性被 Fe(2+)、苯甲基磺酰氟（PMSF）、抑肽酶和胃蛋白酶抑制剂抑制，但不受 N-甲苯磺酰-L-苯丙氨酸氯甲基酮（TPCK）和乙二胺四乙酸（EDTA）的影响。得到了该酶的 3 个内部肽段，APQALTVAAVGATWAR、EKNVGSTVNLLSYDGNK 和 TDATSVLLDGYNVSAVNDLVAK。该酶可直接水解纤维蛋白（原），并优先切割 α-链，而对 β-和 γ-链的切割效率较低，表明它是一种类纤溶酶。它还可降解凝血酶，这表明它可以作为抗凝剂，预防血栓形成。血管内血栓形成是心血管疾病的主要原因之一。基于这些结果，该纯化酶可作为口服纤维蛋白溶解治疗或预防血栓形成的天然药物。
