高密度培养大肠杆菌YK537/pSBHL-11生产重组人细胞白介素

High density Escherichia coli YK537/pSBHL 11 Cultivation Process for Hyperexpression of Recombinant Human Interleukin 3

在 Ｂ． Ｂｒａｕｎ Ｅ Ｓ１０ 型１５ Ｌ 和 Ｎ Ｂ Ｓ Ｂｉｏ Ｆｌｏ ３０００ 型５ Ｌ 发酵罐中，利用补料分批培养技术高密度表达培养含重组质粒ｐ Ｓ Ｂ Ｈ Ｌ１１ 的大肠杆菌 Ｙ Ｋ５３７ ，生产重组人白细胞介素３（ Ｉ Ｌ３） ，发现在发酵过程中，限制性流加甘油，控制溶解氧在３０ ％ ～４０ ％ 左右、３０ ℃生长１１ｈ ，４２ ℃诱导培养４ｈ ，能将发酵液中最终菌体密度从 Ｏ Ｄ１６６００ 提高到 Ｏ Ｄ５３６００（ 相当于每升发酵液含１０６ 克湿菌体） ，并且保持了白细胞介素３ 的表达量，占菌体总蛋白的３０ ％ 左右，含量超过３３ ％ ｇ／ Ｌ，使 Ｉ Ｌ３ 包涵体产量从湿重２２ｇ／ Ｌ 提高到８５ ｇ／ Ｌ，纯化步骤比较简单，超声破菌后经两次洗涤纯度就达到７０ ％ 以上。

High density cultivation of Escherichia coli YK537/pSBHL 11 in a B. Braun ES 10 type and a NBS BioFlo 3000 type fermentor was carried out to overexpress recombinant human Interleukin 3(IL 3) using fed batch method. The results indicated that limiting glycerol at low concentration and keeping dissolved oxygen at 30%～40%, temperature at 30℃ for 11 hours in the growth phase then at 42℃ for 4 hours could increase the final cell density from 16 OD 600 to 53 OD 600 (106g wet cell weight/L). Expressed IL 3 was about 30% of the total amount of protein in E.coli , 3.3g IL 3/L, and the wet weight of IL 3 inclusion body increased from 2.2g/L to 8.5g/L. After ultrasonication and two steps of washing, the purity of IL 3 inclusion body exceeded 70%.