中国传统细菌型豆豉溶栓酶的提取纯化技术研究(简报)

郝征红; 张炳文; 庞庆芳

中国传统细菌型豆豉溶栓酶的提取纯化技术研究(简报)

Separation and purification of fibrinolytic enzyme from Chinese traditional bacteria-type Douchi

摘要

摘要: 为深入开发利用中国传统发酵细菌型豆豉溶栓酶，对豆豉溶栓酶分离纯化工艺进行了研究探讨，确认了最佳的纯化条件：首先用饱和度75％的硫酸铵沉淀豆豉溶栓酶；然后利用DEAE-Sepharose FF（Diethylaminoethyl Sepharose Fast Flow，二乙基氨基琼脂糖快速凝胶）阴离子交换柱进行层析，优化的层析条件为用10 mmol/L Tris（Trishydroxymethylaminomen，三羟甲基氨基甲烷）-HCl（pH 8.7）作为上样缓冲液，采用线性梯度洗脱，洗脱液为10 mmol/L Tris-HCl（pH 8.7）和0.6 mol/L NaCl，洗脱流速为1 mL/min；最后经过Sephadex G-75（葡聚糖凝胶G-75）凝胶过滤，得到纯化倍数为11.29倍的豆豉溶栓酶。利用SDS－PAGE（Sodium dodecyl sulphate-polylamide gel electroresis，十二烷基硫酸钠—聚丙烯酰胺凝胶电泳）显示经纯化后的酶无杂蛋白，初步鉴定其分子量接近31 ku。

Abstract: In order to exploit fibrinolysis components of Chinese traditional bacteria-type Douchi, separation and purification of fibrinolytic enzyme were discussed and the best purification method was determined. Firstly, fibrinolytic enzyme was separated by ammonium sulfate with concentration of 75% saturation. Secondly, purification was achieved by Diethylaminoethyl Sepharose Fast Flow(DEAE-Sepharose FF) anion exchange chromatography with 10mmol/L Tris-HCl (pH 8.7) as sample buffer. The column was eluted stepwise with 0.6 mol/L NaCl and 10 mmol/L Trishydroxymethylaminomen(Tris-HCl) buffer (pH 8.7) at flow rate of 1 mL/min. Finally, fibrinolytic enzyme was further filtered by Sephadex G-75 chromatography and 11.29 times purified fibrinolytic enzyme was gained. Sodium dodecyl sulphate-polylamide gel electroresis (SDS-PAGE) showed that there was no unpurposed protein in purified fibrinolytic enzyme and the molecular weight was identified approximately 31ku preliminarily.

HTML全文

参考文献(0)

施引文献

资源附件(0)