大肠杆菌GalP-GlK途径功能分析OA

The metabolic pathway of glucose in Escherichia coli is mainly initialized through a phosphotransferase sys-tem(PTS)for substrate level phosphorylation of glucose.In order to expand the ability of glucose transport and metabolism in E.coli,a PTS-independent glucose transport pathway(GalP-GlK)was established through the overexpression of galactose transporter(GalP)and glucokinase(GlK)on the basis of deleting the PTS system of E.coli.The new initiation mode and efficiency of glucose metabolism of E.coli were investigated,and the changes of physiological metabolism were further explored.Mutant strain 025P was obtained by deleting ptsHI-crr,ptsG and mlc genes from E.coli B0013-025 genome by gene recombinant technology.The mutant strain 025P grew hardly in the medium with glucose as the only carbon source,indicating a significant loss of glucose transport capacity after the destruction of PTS system.The expression of galP and glk was further enhanced by promoter gap,and the ability to initiate glucose metabolism by oxidative phosphorylation of the recombinant strain 025PG was restored.Strengthening the GalP-GlK pathway could not only initiate glucose metabolism,but also galactose and arabinose metabolism.The order of metabolic strength was glucose(µ=0.31 h-1)＞galactose(µ=0.27 h-1)＞arabinose(µ=0.21 h-1)＞fructose=xylose(µ=0.01 h-1).In conclusion,the PTS-independent strain 025PG was able to re-metabolize glucose after overexpression of GalP-GlK pathway,which has provided a theoretical basis for the sub-sequent genetic selection of related industrial strains.