【摘要】 以重组枯草芽孢杆菌谷氨酰胺合成酶(L-谷氨酸:氨连接酶,G lutam ine Synthetase,GS EC 6.3.1.2)蛋白表达宿主菌株BL21(DE3)(pET3C/谷氨酰胺合成酶)作为研究对象,借助SDS-PAGE分析方法,对于用乳糖诱导由T7 lac启动子控制的重组目的产物蛋白的表达规律进行了优化研究,分别比较了最佳诱导起始生长量、所需乳糖的浓度、诱导持续时间、补加乳糖和氨苄青霉素及不同培养基等参数对重组产物表达的影响.实验结果表明,对于T7 lac启动子控制的重组目的产物,乳糖作为诱导剂也可以起到很好的诱导表达效果,诱导产物的表达量、可溶性及酶活与IPTG诱导产物基本接近.本研究结果为乳糖作为诱导剂应用于重组大肠杆菌谷氨酰胺合成酶工业化生产提供了一定的参考依据.更多还原

【Abstract】 L-Glutamine is mainly produced by fermentation,rarely by enzyme reaction.Glutamine Synthetase gene is amplified from Bacillus Subtilis gnd cloned into prokaryotic expression vector pET3C.After the transformation of this recombinant plasmid into BL-21(DE3),Lactose and IPTG(isopropyl-β-D-thiogalacto-pyranoside isopropylthio-β-D-galactoside) are used to induce gene expression at 37℃ respectively.Both of them can induce the expression of the Glutamine Synthetase gene efficiently.The influences of culture condition such as the lactose concentration,growth medium composition,the duration of the induction and the re-addition of lactose on the expression of the recombinant protein are analyzed in detail.The induction efficiency of lactose is basically the same as that of IPTG, which provides the evidence that lactose could be a promising inducer in the future large scale production of recombined Glutamine Synthetase.更多还原