中国医科大学学报

中国医科大学学报
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中国医科大学学报 ›› 2017, Vol. 46 ›› Issue (12): 1117-1123.doi: 10.12007/j.issn.0258-4646.2017.12.013

• 论著 • 上一篇    下一篇

表达人源SOD的毕赤酵母重组子的构建

李新鸣1,2, 孙冶1,2, 肖纯凌2   

  1. 1. 沈阳医学院基础医学院病原生物学教研室, 沈阳 110034;
    2. 辽宁省环境污染与微生态重点实验室, 沈阳 110034
  • 收稿日期:2017-07-06 出版日期:2017-12-30 发布日期:2017-12-08
  • 通讯作者: 肖纯凌 E-mail:xiaochunling@symc.edu.cn
  • 作者简介:李新鸣(1973-),女,副教授,博士.
  • 基金资助:
    沈阳市科学技术计划(F15-199-1-29)

Construction of Recombinant Pichia pastoris Expressing Human SOD

LI Xinming1,2, SUN Ye1,2, XIAO Chunling2   

  1. 1. Department of Pathogenic Biology, College of Basic Medical Sciences, Shenyang Medical College, Shenyang 110034, China;
    2. Liaoning Province Key Lab of Environment Pollution and Microecology, Shenyang 110034, China
  • Received:2017-07-06 Online:2017-12-30 Published:2017-12-08

摘要: 目的 研究在毕赤酵母重组子中高表达有活性人源超氧化物歧化酶(SOD)方法。方法 在毕赤酵母构建表达重组人源SOD,对培养时间、培养基中铜离子浓度和甲醇诱导条件进行优化,并初步纯化SOD。结果 获得了5个稳定表达人源SOD的毕赤酵母重组子。色谱层析方法分离纯化重组子培养上清中的SOD。在培养48 h,培养基中铜离子浓度0.5%、甲醇诱导浓度1%时上清中SOD分泌量多,同时活力最好。结论 建立毕赤酵母表达可溶性人源SOD的有效方法,实现了在毕赤酵母系统高效表达有活性人源SOD。为发酵生产人源SOD提供了研究基础。

关键词: 毕赤酵母, 重组子, 人源超氧化物歧化酶

Abstract: Objective To optimize the expression of active human Cu,Zn-superoxide dismutase (SOD) in recombinant Pichia pastoris (P.pastoris). Methods A recombinant human SOD expression construct was generated and expressed in P. pastoris. The culture time,concentration of copper ions in the medium,and conditions for methanol induction were optimized,and SOD was preliminarily purified. Results Five P. pastoris recombinants that stably expressed human SOD were obtained. SOD could be purified from the culture supernatants of the recombinant strains by chromatography. The amount of SOD secreted into the supernatant was high with optimal activity,when the incubation time was 48 h,copper ion concentration in the medium was 0.5%,and methanol concentration was 1%. Conclusion An effective method for the expression of soluble human Cu,Zn-SOD in P. pastoris was established,and the enzyme produced using this method exhibited high activity. This method provides a foundation for further research on the production of human SOD by fermentation.

Key words: Pichia pastoris, recombinant, human superoxide dismutase

中图分类号: 

  • Q93-335
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