• 主题
高级搜索  >
历史搜索 清空历史
首页> 美国卫生研究院文献>other >Expression Platforms for Producing Eukaryotic Proteins: A Comparison of E. coli Cell-Based and Wheat Germ Cell-Free Synthesis Affinity and Solubility Tags and Cloning Strategies
【2h】

Expression Platforms for Producing Eukaryotic Proteins: A Comparison of E. coli Cell-Based and Wheat Germ Cell-Free Synthesis Affinity and Solubility Tags and Cloning Strategies

机译：产生真核蛋白的表达平台：大肠杆菌细胞和小麦胚无细胞合成亲和力和可溶性标签以及克隆策略的比较。

代理获取
本网站仅为用户提供外文OA文献查询和代理获取服务，本网站没有原文。下单后我们将采用程序或人工为您竭诚获取高质量的原文，但由于OA文献来源多样且变更频繁，仍可能出现获取不到、文献不完整或与标题不符等情况，如果获取不到我们将提供退款服务。请知悉。
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

Vectors designed for protein production in Escherichia coli and by wheat germ cell-free translation were tested using 21 well-characterized eukaryotic proteins chosen to serve as controls within the context of a structural genomics pipeline. The controls were carried through cloning, small-scale expression trials, large-scale growth or synthesis, and purification. Successfully purified proteins were also subjected to either crystallization trials or 1H-15N HSQC NMR analyses. Experiments evaluated: (1) the relative efficacy of restriction/ligation and recombinational cloning systems; (2) the value of maltose-binding protein (MBP) as a solubility enhancement tag; (3) the consequences of in vivo proteolysis of the MBP fusion as an alternative to post-purification proteolysis; (4) the effect of the level of LacI repressor on the yields of protein obtained from E. coli using autoinduction; (5) the consequences of removing the His tag from proteins produced by the cell-free system; and (6) the comparative performance of E. coli cells or wheat germ cell-free translation. Optimal promoter/repressor and fusion tag configurations for each expression system are discussed.
机译：使用在结构基因组学管道中被选作对照的21种特征充分的真核蛋白，测试了设计用于在大肠杆菌中生产蛋白质并通过小麦胚无细胞翻译的载体。通过克隆，小规模表达试验，大规模生长或合成以及纯化进行对照。成功纯化的蛋白质也要进行结晶试验或 1 H- 15 N HSQC NMR分析。实验评估：（1）限制/连接和重组克隆系统的相对功效； （2）麦芽糖结合蛋白（MBP）作为溶解度增强标签的价值； （3）MBP融合蛋白在体内蛋白水解作为纯化后蛋白水解的替代方法的后果； （4）LacI阻遏物水平对使用自动诱导从大肠杆菌获得的蛋白质的产量的影响； （5）从无细胞系统产生的蛋白质中去除His标签的后果； （6）大肠杆菌细胞或小麦生殖细胞无翻译的比较性能。讨论了每个表达系统的最佳启动子/阻遏物和融合标签配置。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
代理获取

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号