Du Juan, Wu Jian, Dai Guifu, Wang Chunyang, Zhou Xinqin, Song Minghui, Li Jue, Li Jilun
Department of Biotechnology, Zhengzhou University, Zhengzhou 450001, China.
Sheng Wu Gong Cheng Xue Bao. 2009 Apr;25(4):626-31.
A recombinant strain Escherichia coli DH5alpha(pMD19-glnA) including Bacillus subtilis glnA gene was constructed. Capillary electrophoresis and nuclear magnetic resonance were used to determine qualitatively the product of transformation by recombinant strain, and the relative level of mRNA expression of glnA was also determined by fluorescence quantitative RT-PCR. Subsequently, SDS-PAGE (polyacrylamide gel electrophoresis) was used to analysis the relative level of protein. Surprisingly, there was no increase of glutamine production in this recombinant strain, but an obvious increase in the GABA (gamma-aminobutyric acid ) production. It was showed in the experiment that protein expression of the glutamine synthetase did not increase, although glnA gene can be transcribed normally in this recombined strain. The phenomenon of exogenous glnA gene interfering metabolism of Escherichia coli was worthy of further study.
构建了一种包含枯草芽孢杆菌谷氨酰胺合成酶基因的重组大肠杆菌DH5α(pMD19-glnA)。采用毛细管电泳和核磁共振对重组菌株转化产物进行定性测定，并通过荧光定量RT-PCR测定谷氨酰胺合成酶基因mRNA的相对表达水平。随后，用SDS-PAGE（聚丙烯酰胺凝胶电泳）分析蛋白质的相对水平。令人惊讶的是，该重组菌株中谷氨酰胺产量没有增加，但γ-氨基丁酸（GABA）产量明显增加。实验表明，尽管谷氨酰胺合成酶基因在该重组菌株中能正常转录，但谷氨酰胺合成酶的蛋白表达并未增加。外源谷氨酰胺合成酶基因干扰大肠杆菌代谢的现象值得进一步研究。
