Elementary flux mode （EFM） analysis was used in the metabolic analysis of central carbon metabolism in Saccharomyces cerevisiae based on constructed cellular network. Calculated from the metabolic model, the ethanol...Elementary flux mode （EFM） analysis was used in the metabolic analysis of central carbon metabolism in Saccharomyces cerevisiae based on constructed cellular network. Calculated from the metabolic model, the ethanol-producing pathway No. 37 furthest converts the substrate into ethanol among the 78 elementary flux modes. The in silico metabolic phenotypes predicted based on this analysis fit well with the fermentation performance of the engineered strains, KAM3 and KAMll, which confirmed that EFM analysis is valid to direct the construction of Saccharomyces cerevisiae engineered strains, to increase the ethanol yield.展开更多
L-Serine plays a critical role as a building block for cell growth, and thus it is difficult to achieve the direct fermentation of L-serine from glucose. In this study, Corynebacterium glutamicum ATCC 13032 was engine...L-Serine plays a critical role as a building block for cell growth, and thus it is difficult to achieve the direct fermentation of L-serine from glucose. In this study, Corynebacterium glutamicum ATCC 13032 was engineered de novo by blocking and at- tenuating the conversion of L-serine to pyruvate and glycine, releasing the feedback inhibition by L-serine to 3-phosphoglycerate dehydrogenase （PGDH）, in combination with the co-expression of 3-phosphoglycerate kinase （PGK） and feedback-resistant PGDH （PGDHr）. The resulting strain, SER-8, exhibited a lower specific growth rate and significant differ- ences in L-serine levels from Phase I to Phase V as determined for fed-batch fermentation. The intracellular L-serine pool reached （14.22_＋1.41） ~trnol gcoM-1, which was higher than glycine pool, contrary to fermentation with the wild-type strain. Furthermore, metabolic flux analysis demonstrated that the over-expression of PGK directed the flux of the pentose phosphate pathway （PPP） towards the glycolysis pathway （EMP）, and the expression of PGDHr improved the L-serine biosynthesis pathway. In addition, the flux from L-serine to glycine dropped by 24%, indicating that the deletion of the activator GlyR re- sulted in down-regulation of serine hydroxymethyltransferase （SHMT） expression. Taken together, our findings imply that L-serine pool management is fundamental for sustaining the viability of C. glutamicum, and improvement of C1 units genera- tion by introducing the glycine cleavage system （GCV） to degrade the excessive glycine is a promising target for L-serine pro- duction in C. glutamicum.展开更多
基金Supported by the National Natural Science Foundation of China （No.2002AA647040）
文摘Elementary flux mode （EFM） analysis was used in the metabolic analysis of central carbon metabolism in Saccharomyces cerevisiae based on constructed cellular network. Calculated from the metabolic model, the ethanol-producing pathway No. 37 furthest converts the substrate into ethanol among the 78 elementary flux modes. The in silico metabolic phenotypes predicted based on this analysis fit well with the fermentation performance of the engineered strains, KAM3 and KAMll, which confirmed that EFM analysis is valid to direct the construction of Saccharomyces cerevisiae engineered strains, to increase the ethanol yield.
基金supported by grants from Ministry of Science and Technology of China (Grant Nos.2008ZX09401-05 and 2010ZX09401-403)the National Natural Science Foundation of China (Grant No. 31100074)Chinese Academy of Sciences (Grant No. XBXA-2011-009)
文摘L-Serine plays a critical role as a building block for cell growth, and thus it is difficult to achieve the direct fermentation of L-serine from glucose. In this study, Corynebacterium glutamicum ATCC 13032 was engineered de novo by blocking and at- tenuating the conversion of L-serine to pyruvate and glycine, releasing the feedback inhibition by L-serine to 3-phosphoglycerate dehydrogenase （PGDH）, in combination with the co-expression of 3-phosphoglycerate kinase （PGK） and feedback-resistant PGDH （PGDHr）. The resulting strain, SER-8, exhibited a lower specific growth rate and significant differ- ences in L-serine levels from Phase I to Phase V as determined for fed-batch fermentation. The intracellular L-serine pool reached （14.22_＋1.41） ~trnol gcoM-1, which was higher than glycine pool, contrary to fermentation with the wild-type strain. Furthermore, metabolic flux analysis demonstrated that the over-expression of PGK directed the flux of the pentose phosphate pathway （PPP） towards the glycolysis pathway （EMP）, and the expression of PGDHr improved the L-serine biosynthesis pathway. In addition, the flux from L-serine to glycine dropped by 24%, indicating that the deletion of the activator GlyR re- sulted in down-regulation of serine hydroxymethyltransferase （SHMT） expression. Taken together, our findings imply that L-serine pool management is fundamental for sustaining the viability of C. glutamicum, and improvement of C1 units genera- tion by introducing the glycine cleavage system （GCV） to degrade the excessive glycine is a promising target for L-serine pro- duction in C. glutamicum.
