摘要
研究了重组菌BL21-HTa-cgk Z产κ-卡拉胶酶的分离纯化技术。在发酵罐中制备粗酶液,将粗酶液沉淀、阴离子交换层析、超滤浓缩和葡聚糖凝胶过滤层析,得到高纯度的κ-卡拉胶酶。通过对沉淀剂、沉淀时间、沉淀pH的单因素试验确定κ-卡拉胶酶的最优沉淀条件,即:沉淀剂40%的乙醇,沉淀时间4 h,沉淀pH 6.0。通过各级分离纯化,得到聚丙烯酰胺凝胶电泳分子质量为62 ku的条带,经LC-MS/MS鉴定,确定该条带为含有κ-卡拉胶酶的条带。测定纯化后的κ-卡拉胶酶酶活,结果显示,κ-卡拉胶酶的比活力为94.23 U/mg,纯化倍数12.33倍。重组菌BL21-HTa-cgk Z发酵产生的κ-卡拉胶酶,经各级分离纯化,得到高纯度的κ-卡拉胶酶,为卡拉胶酶的食品工业化应用奠定了基础。
To obtain the purified κ-carrageenase by the recombinant strain BL21-HTa-cgkZ. The purified κ-carrageenase achieved by precipitation, ion exchange chromatography of CM Sepharose Fast Flow, ultrafiltration concentration and gel filtration chromatography of Sephadex G-75. The optimal precipitation conditions of the κ-carrageenase were 40%ethanol, 4 h, pH=6.0. κ-carrageenase was successively purified by using polyacrylamide gel electrophoresis to get a clear band, its molecular weight was 62 ku. The strip was κ-carrageenase by LC-MS/MS identification. The result showed that the specific activity of κ-carrageenase was 94.23 U/mg, 12.33 times that of before. This paper lay the foundation for industrial application of carrageenase.
作者
杨敏
田琳
刘哲民
牟海津
Yang Min;Tian Lin;Liu Zhemin;Mou Haijin(College of Food Science and Engineering, Ocean University of China, Qingdao 266000, Shandon)
出处
《中国食品学报》
EI
CAS
CSCD
北大核心
2018年第5期130-136,共7页
Journal of Chinese Institute Of Food Science and Technology
基金
山东省科技发展计划项目(2014GHY115037)
国家海洋局海洋公益性行业科研专项(201505022)
中央高校基本科研业务费基础性工作专项基金项目(201362041)
