摘要
目的获取活性高的重组SOD 1/3杂合酶。方法用含pET-SOD 1/3的工程菌诱导表达重组SOD 1/3杂合酶,通过分析SDS-PAGE及SOD酶活性,考察诱导温度、诱导时间、诱导剂浓度、Cu 2+、Zn 2+浓度对重组酶表达量和活性的影响。结果正交试验表明,用1 mmol/L异丙基-β-D-硫代半乳糖苷(IPTG)在A600=1.2时诱导,Cu2+、Zn2+浓度分别为2.4 mmol/L,诱导温度15℃,诱导时间12 h,能获得高活性重组SOD 1/3,其比活性为1681U/mg。结论培养温度和Cu2+、Zn2+浓度是优化表达的最主要因素。降低诱导温度有利于重组酶的可溶性表达,添加Cu2+、Zn2+能促进重组酶活性提高。
Objective To obtain recombinant hybrid enzyme SOD1/3 with higher activity in E. coli. Methods The recombinant bacteria containing expression plasmid pET-SOD1/3 was used to express hybrid enzyme SOD1/3. The effects of induction temperature, induction time, IPTG concentration and Cu^2＋ or Zn^2＋concentration on the expression amount and activity of recombinant hybrid enzyme SOD1/3 were investigated by SDS-PAGE and SOD activity assay. Results When the recombinant bacteria were cultured at 15 ℃ for 12 h with 2.4 mmol/L Cu^2＋ or Zn^2＋ and 1 mmol/L IPTG at A600=1.2, the activity of hybrid enzyme SOD 1/3 was higher with specific activity of 1 681U/ mg. Conclusion The optimal expression of hybrid enzyme SOD1/3 is mainly affected by induction temperature and Cu^2＋ or Zn^2＋ concentration. Hybrid enzyme SOD1/3 can be expressed in supernatant at low temperature. The addition of Cu^2＋ or Zn^2＋ can increase the activity of hybrid enzyme SOD1/3.
出处
《食品与药品》
CAS
2008年第6期8-11,共4页
Food and Drug
关键词
超氧化物歧化酶
杂合酶
表达
优化
superoxide dismutase
hybrid enzyme
expression
optimization
