Chen Zhenya, Li Ye, Sun Xinxiao, Yuan Qipeng
State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, No.15 East Road of North Third Ring, Chao Yang District, Beijing, 100029, China.
Department of Biotechnology, Beijing Polytechnic, No.1 TaiyanggongShaoyaoju, Chao Yang District, Beijing, 100029, China.
J Biotechnol. 2016 Oct 20;236:159-65. doi: 10.1016/j.jbiotec.2016.08.016. Epub 2016 Aug 25.
Escherichia coli (E. coli) is widely used to express a variety of heterologous proteins. Efforts have been made to enhance the expression level of the desired protein. However, problems still exist to regulate the level of protein expression and therefore, new strategies are needed to overcome those issues. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) which is properly expressed in E. coli might play a leading role and increase the expression levels of the target proteins. In this study, GAPDH was fused with a target enzyme, ChSase ABC I, an endoeliminase and polysaceharide lyase. Our results confirmed this hypothesis and indicated that GAPDH boosted the expression level of ChSase ABC I with an increase of 2.25 times, while the enzymatic activity with an increase of 2.99 times. The hypothesis were also supported by RT-PCR study and GAPDH was more effective in enhancing the expression level and enzymatic activity as compared to MBP, which is commonly used as fused tag and can improve the soluble expression of target protein. addition, the expression level and enzymatic activity of other polysaceharide lyases were also improved in the presence of GAPDH. The findings of this study prove that GAPDH has a strong effect on enhancing the expression level and enzymatic activity of the target proteins.
大肠杆菌（E. coli）被广泛用于表达多种异源蛋白。人们已努力提高所需蛋白的表达水平。然而，在调节蛋白表达水平方面仍然存在问题，因此，需要新的策略来克服这些问题。在大肠杆菌中正常表达的甘油醛-3-磷酸脱氢酶（GAPDH）可能发挥主导作用并提高目标蛋白的表达水平。在本研究中，GAPDH与一种目标酶ChSase ABC I（一种内切消除酶和多糖裂解酶）融合。我们的结果证实了这一假设，并表明GAPDH使ChSase ABC I的表达水平提高了2.25倍，而酶活性提高了2.99倍。RT-PCR研究也支持了这一假设，并且与通常用作融合标签并可提高目标蛋白可溶性表达的MBP相比，GAPDH在提高表达水平和酶活性方面更有效。此外，在GAPDH存在的情况下，其他多糖裂解酶的表达水平和酶活性也得到了提高。本研究结果证明，GAPDH对提高目标蛋白的表达水平和酶活性具有显著作用。
