Developing an l-threonine-producing strain from wild-type Escherichia coli by modifying the glucose uptake, glyoxylate shunt, and l-threonine biosynthetic pathway

摘要

Wild-type Escherichia coli MG1655 usually does not accumulate l-threonine. In this study, the effects of 13 genes related to the glucose uptake, glycolysis, TCA cycle, l-threonine biosynthesis, or their regulation on l-threonine accumulation in E. coli MG1655 were investigated. Sixteen E. coli mutant strains were constructed by chromosomal deletion or overexpression of one or more genes of rsd, ptsG, ptsH, ptsI, crr, galP, glk, iclR, and gltA; the plasmid pFW01-thrA*BC-rhtC harboring the key genes for l-threonine biosynthesis and secretion was introduced into these mutants. The analyses on cell growth, glucose consumption, and l-threonine production of these recombinant strains showed that most of these strains could accumulate l-threonine, and the highest yield was obtained in WMZ016/pFW01-thrA*BC-rhtC. WMZ016 was derived from MG1655 by deleting crr and iclR and enhancing the expression of gltA. WMZ016/pFW01-thrA*BC-rhtC could produce 17.98 g/L l-threonine with a yield of 0.346 g/g glucose, whereas the control strain MG1655/pFW01-thrA*BC-rhtC could only produce 0.68 g/L l-threonine. In addition, WMZ016/pFW01-thrA*BC-rhtC could tolerate the high concentration of glucose and produced no detectable by-products; therefore, it should be an ideal platform strain for further development. The results indicate that manipulating the glucose uptake and TCA cycle could efficiently increase l-threonine production in E. coli.