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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
【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.
- 【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