Optimization of the fermentation conditions for transglutaminase by recombinant Escherichia coli
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摘要: 以谷氨酰胺转氨酶生产菌株重组大肠杆菌BL21（DE3）/mtg为研究对象,对工程菌的发酵产酶条件进行了优化。首先通过单因素实验法,对培养基碳、氮源种类及浓度、培养温度、初始p H、装液量、接种量、诱导时机、诱导浓度、诱导温度及诱导时间进行了优化;随后采用Plackett-Burman设计从中筛选出3个关键影响因子:初始p H、培养温度及诱导温度;最后通过Box-Benhnken设计建立上述3个因子对谷氨酰胺转氨酶比酶活的响应面模型。结果表明,最佳发酵条件为:葡萄糖5 g/L,酵母粉28 g/L,蛋白胨14 g/L,培养温度34℃,初始p H7.0,装液量50 m L（250 m L三角瓶）,接种量5%,诱导时机4 h,IPTG浓度0.6 mmol/L,诱导温度25℃,诱导时间14 h。在该条件下,优化后的谷氨酰胺转氨酶比酶活达1.507 U/mg,为未优化前的1.56倍。Abstract: The paper investigated the improvement of the production of microbial transglutaminase( MTGase,MTG)by recombinant Escherichia coli BL21( DE3) / mtg in 250 m L shake flasks.Using single factor experiment,the types and the concentrations of carbon sources and nitrogen sources of the media,the inoculum sizes,the medium volumes,the culture temperature,the initial p H,the inducer concentration,the adding time of inducer,the induction temperature and the induction time were optimized,respectively. Among the above factors,3 key factors,the culture temperature,the initial p H and the induction temperature,were selected by Plackett- Burman design.And a response surface model of the 3 factors and the specific activity of MTG were constructed by Box- Benhnken design.The optimal conditions were as follows,the media contained 5 g / L of glucose,28 g / L of yeast extract and14 g / L of tryptone,and the bacteria were cultured in 50 m L( 250 m L triangle bottle),p H7.0 optimized media at34 ℃ for 4 h with 5%( v / v) inocula and were induced by 0.6 mmol / L IPTG at 25 ℃ for 14 h. Under the optimal conditions,the maximum specific activity of MTG was 1.507 U / mg,which was 1.56 times larger than the MTG produced in initial conditions.
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