木瓜蛋白酶的原位固定化及理化性质研究

doi:10.3969/j.issn.1000-2006.2014.06.019

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南京林业大学学报（自然科学版） ›› 2014, Vol. 38 ›› Issue (06): 99-104.doi: 10.3969/j.issn.1000-2006.2014.06.019

木瓜蛋白酶的原位固定化及理化性质研究

苏二正¹, 李明亮², 魏东芝²

1. 南京林业大学轻工科学与工程学院,酶与发酵工程实验室,江苏南京 210037;
2. 华东理工大学生物反应器工程国家重点实验室,鲁华生物技术研究所,上海 200237

出版日期:2014-12-31 发布日期:2014-12-31
基金资助:
收稿日期:2014-02-23 修回日期:2014-08-01
基金项目:国家高技术研究发展计划(2012AA022206B); 南京林业大学引进高层次人才科研基金项目; 南京林业大学“青年拔尖人才培养计划”专项经费资助项目; 江苏高校优势学科建设工程资助项目(PAPD)
第一作者:苏二正,副教授,博士。E-mail: ezhsu@njfu.edu.cn。
引文格式:苏二正, 李明亮, 魏东芝. 木瓜蛋白酶的原位固定化及理化性质研究[J]. 南京林业大学学报:自然科学版,2014,38(6):99-104.

Study on in situ immobilization and characterization of papain

SU Erzheng¹, LI Mingliang², WEI Dongzhi²

1. Enzyme and Fermentation Technology Laboratory, College of Light Industry Science and Engineering, Nanjing Forestry University, Nanjing 210037,China;
2. State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China

Online:2014-12-31 Published:2014-12-31

摘要/Abstract

摘要： 以氨基载体原位固定化PEG相中的木瓜蛋白酶,获得了高固定化率(95.9%)和酶活回收率(38.9%)的固定化木瓜蛋白酶,解决了双水相分离纯化后成相聚合物和目的蛋白难以分离的问题。固定化木瓜蛋白酶的最适pH在7.0左右,最适温度处于60～70 ℃之间。相比游离木瓜蛋白酶,在广泛的pH区间内(3.0～9.0)固定化酶稳定性都较好,热稳定性也有改善。在4 ℃密闭容器内将固定化木瓜蛋白酶以湿润状态保存可以较好地保留其活性。LH-HA固定化木瓜蛋白酶可以特异性地水解单克隆抗体IgG,制备Fab和Fc片段。

Abstract: Amino carriers could immobilize the papain in situ in the PEG phase, which gave high papain immobilization yield(95.9%)and activity recovery(51.3%), and thus solved the difficulty in separation of the target protein and polymer after aqueous two-phase purification. The optimum pH of the immobilized papain was around 7.0, and the optimum temperature was 60-70 ℃. Compared with free enzyme, the immobilized papain was fairly stable in a wide pH(3-9)range, and the thermal stability was also improved. The best way to store immobilized papain was in wet at 4 ℃ and airtight container. The LH-HA immobilized papain could catalyze the hydrolysis of monoclonal antibody IgG to produce Fab and Fc fragments. The immobilized papain prepared in this work could be used in the production of antibody fragment industrially.

中图分类号:

TQ925

苏二正,李明亮,魏东芝. 木瓜蛋白酶的原位固定化及理化性质研究[J]. 南京林业大学学报（自然科学版）, 2014, 38(06): 99-104.

SU Erzheng, LI Mingliang, WEI Dongzhi. Study on in situ immobilization and characterization of papain [J].Journal of Nanjing Forestry University (Natural Science Edition), 2014, 38(06): 99-104.DOI: 10.3969/j.issn.1000-2006.2014.06.019.

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木瓜蛋白酶的原位固定化及理化性质研究

Study on in situ immobilization and characterization of papain

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