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重组人β淀粉样蛋白1-42小规模发酵及小量纯化工艺的摸索

Small scale fermentation and a small purification technology of recombinant human β-amyloid peptide 1-42

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【作者】 申茉函王全才杨丽

【Author】 SHEN Mo-han;WANG Quan-cai;YANG Li;Shengjing Hospital of China Medical University;The People’s Hospital of Liaoning Province;Key Laboratory of Zoonosis Research of Ministry of Education,Jilin University;

【机构】 中国医科大学附属盛京医院辽宁省人民医院吉林大学人兽共患研究教育部重点实验室

【摘要】 目的研究重组人β淀粉样蛋白1-42(rh Aβ1-42)在5 L摇瓶中的高密度发酵实验,以及小量纯化工艺的方法。方法分别从甲醇浓度、p H、诱导时间等方面对毕赤酵母重组菌株产生rh Aβ1-42的发酵过程进行了优化;通过硫酸铵沉淀目的蛋白、透析、脱盐和阴离子交换层析,对rh Aβ1-42小量精确的纯化方法进行了研究。结果确定rh Aβ1-42在毕赤酵母中分泌表达的最佳条件为:在p H 6.0的条件下,以0.5%甲醇诱导72 h,经过硫酸铵沉淀目的蛋白、透析、脱盐和阴离子交换层析,rh Aβ1-42纯度可达94%以上。结论确定了rh Aβ1-42在毕赤酵母中分泌表达的最佳条件,建立了小量纯化rh Aβ1-42的新方法。

【Abstract】 Objective To study the high-cell-density fermentation experiment of human recombinant β-amyloid protein 1-42( rh Aβ1-42) in 5 L shake flask and the method of small scale purification. Methods Fermentation parameters of rh Aβ1-42 was further optimized mainly on concentration of methanol induction,pH value,and induced time. Secreted supernatant of rh Aβ1-42 was purified with ammonium sulphate sediment,dialysis desalination and anionexchange chromatography. Results The optimal condition of rh Aβ1-42 expressed in Pichia pastoris was: under the condition of pH = 6,induced by 0. 5% methanol for 72 h,precipitation of target protein by ammonium sulfate,dialysis desalting,anion exchange chromatography,rh Aβ1-42 purity could reach more than 94%. Conclusion A stable fermentation and new technology of the small scale purification is successfully set up.

【基金】 国家高技术研究发展计划(863计划)(2004AA205020)
  • 【文献出处】 实用药物与临床 ,Practical Pharmacy and Clinical Remedies , 编辑部邮箱 ,2015年03期
  • 【分类号】R749.16
  • 【下载频次】84
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