重组N-乙酰鸟氨酸脱酰基酶的表达条件优化-食品科学.PDFVIP

※生物工程 食品科学 2008, Vol. 29, No. 04 239 重组N- 乙酰鸟氨酸脱酰基酶的表达条件优化 李 环，朱大伟，陈 悦，任 真，韦 萍 (南京工业大学制药与生命科学学院，江苏 南京 210009) 摘 要：N-乙酰鸟氨酸脱酰基酶(N-acetylornithine deacetylase，NAOase)为新型氨基酸拆分酶，可在重组菌BL21 (DE3)-pET22b-argE中胞内诱导表达。为降低生产成本并提高酶活，优化了培养基成分及诱导条件。结果表明，进 口的蛋白胨和酵母膏明显优于国产原料。5.0g/L 的乳糖可以代替昂贵的IPTG(isopropy1 β-D-1-Thiogalacto pyranoside，异丙基β-D硫代半乳糖苷)起到有效的诱导效果。最佳碳源为1.0g/L甘油，氮源为 15.0ml/L玉米浆和 5.0g/L蛋白胨。NAOase大多以不可溶的包涵体形式表达，少量为可溶的活性表达。降低诱导温度可增大活性表达 的比例，而整菌表达量也随之降低。22℃诱导10h为活性表达的最适条件，生物量为6.91，酶活为392.65U/ml， 分别为优化前的1.65倍和5.72倍。 关键词：N- 乙酰鸟氨酸脱乙酰基酶；表达优化；包涵体；可溶性部分酶活 Expression Conditions Optimization of Recombiant N-acetylornithine Decatylase LI Huan，ZHU Da-wei，CHEN Yue，REN Zhen，WEI Ping (College of Life Science and Pharmacy, Nanjing University of Technology, Nanjing 210009, China) Abstract ：N-acetylornithine decatylase(NAOase), a new kind of amino acids resolving enzyme, could be cellular expressed by the recombinant strain BL21(DE3)-pET22b-argE. In order to reduce producing cost and increase soluble NAOase activity, the culture medium and inducing conditions were optimized. The results showed that the imported peptone and yeast extracts were more suitable than the native materials. Lactose of 5.0g/L could act as inducer in place of expensive IPTG The optimized carbon and nitrogen sources are: 1.0 g/L glycerol, 15.0 ml/L corn steep liquor and 5.0 g/L peptone. Most of NAOase were expressed as bodies, little as active enzymes. The proportion of the soluble expression could be increased by lowering the inducing inclusion temperature, but the yield of target protein followed was decreased. With the optimized active inducing condition of 22 ℃for 10 h, the biomass is enhanced by