食品科学

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解淀粉芽孢杆菌产纤溶酶的19 L发酵罐实验及其酶学性质

刘 林,谢 和   

  1. 贵州大学生命科学学院,贵州 贵阳 550025
  • 出版日期:2014-04-15 发布日期:2014-04-18

Production and Characterization of Fibrinolytic Enzyme from Bacillus amyloliquefaciens in 19 L Fermenter and Investigation of Its Properties

LIU Lin, XIE He   

  1. College of Life Sciences, Guizhou University, Guiyang 550025, China
  • Online:2014-04-15 Published:2014-04-18

摘要:

在摇瓶实验的基础上,对解淀粉芽孢杆菌(Bacillus amyloliquefaciens)GZJSI-12-7产纤溶酶最佳条件进行19 L发酵罐实验,结果表明:GZJSI-12-7在发酵罐中培养6 h时开始产纤溶酶,78 h后纤溶酶产量基本稳定,达到4 178.39 IU/mL,产酶高峰比摇瓶发酵提前12 h左右。将发酵液经离心除菌、硫酸铵分级盐析、透析除盐、SephadexG-75凝胶过滤,得到纤溶酶的纯化倍数为9.84 倍,酶活力回收率为42.52%,比活力为48 073.193 IU/mg。对纤溶酶的酶学性质研究表明,该酶的分子质量约为30.2 kD,最适温度和pH值分别为40 ℃、7.5;在40 ℃以下酶的稳定性良好,超过50 ℃后酶活力迅速降低,耐热性较差;金属离子Mg2+、Ca2+、Mn2+对纤溶酶活性均有较强的激活作用,而Cu2+和Fe3+对酶活性具有明显的抑制作用。

关键词: 解淀粉芽孢杆菌, 纤溶酶, 发酵, 分离纯化, 酶学性质

Abstract:

As an extension of our previous fermentation experiments in shake flasks, the objective of this study was tooptimize the fermentation conditions for the production of fibrinolytic enzyme by Bacillus amyloliquefaciens GZJSI-12-7 ina 19 L fermenter. The results showed that plasmin activity began to appear after incubation for 6 h and remained constantafter 78 h at a level of 4 178.39 IU/mL. The peak of enzyme production occurred about 12 h earlier than in shake flasks.Fibrinolytic enzyme was purified from the fermented liquid medium by bactofugation, ammonium sulfate precipitation,dialysis and Sephadex G-75 gel filtration, with a purification fold of 9.84 and an activity recovery of 42.52%. The specificactivity of the purified fibrinolytic enzyme was 48 073.193 IU/mg. Enzymatic characterization showed that the molecularweight of the fibrinolytic enzyme was about 30.2 kD, and its optimum temperature and pH were 40 ℃ and 7.5, respectively.The purified fibrinolytic enzyme was stable blow 40 ℃ but sharply decreased above 50 ℃. The enzymatic activity wasmarkedly catalyzed by Mg2+, Ca2+ and Mn2+, but strongly inhibited by Cu2+ and Fe3+.

Key words: Bacillus amyloliquefaciens, fibrinolytic enzyme, fermentation, purification, characterization