重组人DNaseⅠ不同表达模式的下游工艺研究

Downstream processes for recombinant human DNase Ⅰ from two different expression patterns in Escherichia coli

研究了诱导温度对重组人DNaseⅠ(rhDNaseⅠ)在大肠杆菌中可溶表达的影响,发现加诱导剂后在37℃培养时,rhDNaseⅠ融合蛋白以包涵体形式存在;而20℃培养,rhDNaseⅠ融合蛋白有62.3%分泌到周质,并且具有酶活性。对这两种不同表达模式表达的rhDNaseⅠ融合蛋白进行分离纯化,发现前者最终酶活为592.2 U/mg,回收率为32.1%;后者酶活为1283U/mg,回收率为24.1%。比较这两种纯化工艺,分泌表达模式的分离纯化工艺较好。

The relationship between the induction temperature and the soluble expression of human DNase I in E. coli has been studied. It was found that human DNase I protein assembled in inclusion bodies under 37 ℃ induction, whereas under 20 ℃ induction human DNase I was mostly （about 62.3% ） secreted to the periplasmic space and showed DNase enzyme activity. The two kinds of human DNase I protein from the two different expression patterns were purified. The results indicated that the enzyme activity of recombinant human DNase I from inclusion bodies was 592.2 U/mg, and its recovery rate was 32.1% , whereas the activity of that from the periplasmic space was 1283 U/mg, and the recovery rate was 24.1%. Furthermore, the purification process for DNase I secreted to the periplasmic space was easier than that for DNase I assembled in inclusion bodies.