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Carbon dioxide (CO2) emissions are related to global warming. However, CO2 can be used as an abundant and cheap carbon source for production of valuable chemicals using carbon capture and storage technology. Here, the genes related to carbon flux toward pyruvate biosynthesis in E. coli were deleted to enhance the incorporation of CO2 for succinate production. The codonoptimized carbonic anhydrase gene (SP(-)HCCA) derived from Hahella chejuensis KCTC 2396 and the phosphoenolpyruvate carboxylase gene (ppc) of E. coli W3110 were co-overexpressed to enhance carbon flux toward oxaloacetate synthesis in E. coli. Finally, we constructed SGJS134, which shows the highest production of succinate derived from CO2 compared with other strains. SGJS134 produced approximately 6.5 mM succinate from CO2 and yielded approximately 13.0 mM succinate per dry cell weight. These results may be useful for enhancing the incorporation of CO2 for succinate production in E. coli. Additionally, the metabolic engineering method used in this study will propose the potential of E. coli to convert CO2 to valuable chemicals. [ABSTRACT FROM AUTHOR]

In this study, a gas chromatography–mass spectrometry (GC–MS) analysis method was developed and validated for the quantitative analysis of limonene that was attempted to be industrially mass‐produced from genetically engineered Saccharomyces cerevisiae (S. cerevisiae). Owing to the high potential economic value of limonene, (e.g., for biofuels, fragrant additives, pharmaceuticals, or organic solvents), it is important to develop and validate a readily available quantitative analysis method for limonene produced from genetically engineered yeast or microorganisms. The engineered S. cerevisiae was cultured using two‐phase extractive fermentation with n‐dodecane, which is known to alleviate monoterpene toxicity, used as an extractive solvent. The quantitative GC–MS analysis of the n‐dodecane overlay that contained limonene produced from the engineered S. cerevisiae was validated to be in accord with the international standards set forth by the Food and Drug Administration. [ABSTRACT FROM AUTHOR]

Both ThyA and ThyX proteins catalyze the transfer of the methyl group from methylenetetrahydrofolate ( CH2H4-folate) to d UMP, forming d TMP. To estimate the relative steady state expression levels of ThyA and ThyX, Western blot analysis was performed using ThyA or ThyX antiserum on total protein from the wild-type, ΔthyX, and thyX-complemented strains of Corynebacterium glutamicum. The level of ThyA decreased gradually during the stationary growth phase but that of ThyX was maintained steadily. Whereas the expression level of ThyA in a ΔsigB strain was comparable to that of the wild-type, the level of ThyX was significantly diminished in the deletion mutant and was restored to that of the wild-type in the complemented strain, indicating that the level of ThyX was regulated by Sig B. Growth of the C. glutamicum ΔsigB strain was dependent upon coupling activity of dihydrofolate reductase ( DHFR) with ThyA for the synthesis of thymidine, and thus showed sensitivity to the inhibition of DHFR by the experimental inhibitor, WR99210- HCl. These results suggested that the relative levels of ThyA and ThyX differ in response to different growth phases and that Sig B is necessary for maintenance of the level of ThyX during transition into the stationary growth phase. [ABSTRACT FROM AUTHOR]