Search

Background: Azotobacter vinelandii is a bacterium that produces alginate and polyhydroxybutyrate (P3HB); however, the role of NAD(P)H/NAD(P)+ ratios on the metabolic fluxes through biosynthesis pathways of these biopolymers remains unknown. The aim of this study was to evaluate the NAD(P)H/NAD(P)+ ratios and the metabolic fluxes involved in alginate and P3HB biosynthesis, under oxygen-limiting and non-limiting oxygen conditions. Results: The results reveal that changes in the oxygen availability have an important effect on the metabolic fluxes and intracellular NADPH/NADP+ ratio, showing that at the lowest OTR (2.4 mmol L-1 h-1), the flux through the tricarboxylic acid (TCA) cycle decreased 27.6-fold, but the flux through the P3HB biosynthesis increased 6.6-fold in contrast to the cultures without oxygen limitation (OTR = 14.6 mmol L-1 h-1). This was consistent with the increase in the level of transcription of phbB and the P3HB biosynthesis. In addition, under conditions without oxygen limitation, there was an increase in the carbon uptake rate (twofold), as well as in the flux through the pentose phosphate (PP) pathway (4.8-fold), compared to the condition of 2.4 mmol L-1 h-1. At the highest OTR condition, a decrease in the NADPH/NADP+ ratio of threefold was observed, probably as a response to the high respiration rate induced by the respiratory protection of the nitrogenase under diazotrophic conditions, correlating with a high expression of the uncoupled respiratory chain genes (ndhII and cydA) and induction of the expression of the genes encoding the nitrogenase complex (nifH). Conclusions: We have demonstrated that changes in oxygen availability affect the internal redox state of the cell and carbon metabolic fluxes. This also has a strong impact on the TCA cycle and PP pathway as well as on alginate and P3HB biosynthetic fluxes. [ABSTRACT FROM AUTHOR]

Background: Understanding the effects of oxygen levels on yeast xylose metabolism would benefit ethanol production. In this work, xylose fermentative capacity of Schefersomyces stipitis, Spathaspora passalidarum, Spathaspora arborariae and Candida tenuis was systematically compared under aerobic, oxygen-limited and anaerobic conditions. Results: Fermentative performances of the four yeasts were greatly influenced by oxygen availability. S. stipitis and S. passalidarum showed the highest ethanol yields (above 0.44 g g-1) under oxygen limitation. However, S. passalidarum produced 1.5 times more ethanol than S. stipitis under anaerobiosis. While C. tenuis showed the lowest xylose consumption rate and incapacity to produce ethanol, S. arborariae showed an intermediate fermentative performance among the yeasts. NAD(P)H xylose reductase (XR) activity in crude cell extracts correlated with xylose consumption rates and ethanol production. Conclusions: Overall, the present work demonstrates that the availability of oxygen influences the production of ethanol by yeasts and indicates that the NADH-dependent XR activity is a limiting step on the xylose metabolism. S. stipitis and S. passalidarum have the greatest potential for ethanol production from xylose. Both yeasts showed similar ethanol yields near theoretical under oxygen-limited condition. Besides that, S. passalidarum showed the best xylose consumption and ethanol production under anaerobiosis. [ABSTRACT FROM AUTHOR]

Background: Ribofavin, an intermediate of primary metabolism, is one kind of important food additive with high economic value. The microbial cell factory Bacillus subtilis has already been proven to possess significant importance for the food industry and have become one of the most widely used ribofavin-producing strains. In the practical fermentation processes, a sharp decrease in ribofavin production is encountered along with a decrease in the dissolved oxygen (DO) tension. Influence of this oxygen availability on ribofavin biosynthesis through carbon central metabolic pathways in B. subtilis is unknown so far. Therefore the unveiled effective metabolic pathways were still an unaccomplished task till present research work. Results: In this paper, the microscopic regulation mechanisms of B. subtilis grown under different dissolved oxygen tensions were studied by integrating 13C metabolic flux analysis, metabolomics and transcriptomics. It was revealed that the glucose metabolic flux through pentose phosphate (PP) pathway was lower as being confirmed by smaller pool sizes of metabolites in PP pathway and lower expression amount of ykgB at transcriptional level. The latter encodes 6-phosphogluconolactonase (6-PGL) under low DO tension. In response to low DO tension in broth, the glucose metabolic flux through Embden-Meyerhof-Parnas (EMP) pathway was higher and the gene, alsS, encoding for acetolactate synthase was significantly activated that may result due to lower ATP concentration and higher NADH/NAD+ ratio. Moreover, ResE, a membrane-anchored protein that is capable of oxygen regulated phosphorylase activity, and ResD, a regulatory protein that can be phosphorylated and dephosphorylated by ResE, were considered as DO tension sensor and transcriptional regulator respectively. Conclusions: This study shows that integration of transcriptomics, 13C metabolic flux analysis and metabolomics analysis provides a comprehensive understanding of biosynthesized ribofavin's regulatory mechanisms in B. subtilis grown under different dissolved oxygen tension conditions. The two-component system, ResD--ResE, was considered as the signal receiver of DO tension and gene regulator that led to differences between biomass and ribofavin production after triggering the shifts in gene expression, metabolic flux distributions and metabolite pool sizes. [ABSTRACT FROM AUTHOR]

Background: Azotobacter vinelandii is a bacterium that produces alginate and polyhydroxybutyrate (P3HB); however, the role of NAD(P)H/NAD(P) + ratios on the metabolic fluxes through biosynthesis pathways of these biopolymers remains unknown. The aim of this study was to evaluate the NAD(P)H/NAD(P) + ratios and the metabolic fluxes involved in alginate and P3HB biosynthesis, under oxygen-limiting and non-limiting oxygen conditions., Results: The results reveal that changes in the oxygen availability have an important effect on the metabolic fluxes and intracellular NADPH/NADP + ratio, showing that at the lowest OTR (2.4 mmol L -1  h -1 ), the flux through the tricarboxylic acid (TCA) cycle decreased 27.6-fold, but the flux through the P3HB biosynthesis increased 6.6-fold in contrast to the cultures without oxygen limitation (OTR = 14.6 mmol L -1  h -1 ). This was consistent with the increase in the level of transcription of phbB and the P3HB biosynthesis. In addition, under conditions without oxygen limitation, there was an increase in the carbon uptake rate (twofold), as well as in the flux through the pentose phosphate (PP) pathway (4.8-fold), compared to the condition of 2.4 mmol L -1  h -1 . At the highest OTR condition, a decrease in the NADPH/NADP + ratio of threefold was observed, probably as a response to the high respiration rate induced by the respiratory protection of the nitrogenase under diazotrophic conditions, correlating with a high expression of the uncoupled respiratory chain genes (ndhII and cydA) and induction of the expression of the genes encoding the nitrogenase complex (nifH)., Conclusions: We have demonstrated that changes in oxygen availability affect the internal redox state of the cell and carbon metabolic fluxes. This also has a strong impact on the TCA cycle and PP pathway as well as on alginate and P3HB biosynthetic fluxes.

Background: Understanding the effects of oxygen levels on yeast xylose metabolism would benefit ethanol production. In this work, xylose fermentative capacity of Scheffersomyces stipitis, Spathaspora passalidarum, Spathaspora arborariae and Candida tenuis was systematically compared under aerobic, oxygen-limited and anaerobic conditions., Results: Fermentative performances of the four yeasts were greatly influenced by oxygen availability. S. stipitis and S. passalidarum showed the highest ethanol yields (above 0.44 g g -1 ) under oxygen limitation. However, S. passalidarum produced 1.5 times more ethanol than S. stipitis under anaerobiosis. While C. tenuis showed the lowest xylose consumption rate and incapacity to produce ethanol, S. arborariae showed an intermediate fermentative performance among the yeasts. NAD(P)H xylose reductase (XR) activity in crude cell extracts correlated with xylose consumption rates and ethanol production., Conclusions: Overall, the present work demonstrates that the availability of oxygen influences the production of ethanol by yeasts and indicates that the NADH-dependent XR activity is a limiting step on the xylose metabolism. S. stipitis and S. passalidarum have the greatest potential for ethanol production from xylose. Both yeasts showed similar ethanol yields near theoretical under oxygen-limited condition. Besides that, S. passalidarum showed the best xylose consumption and ethanol production under anaerobiosis.

Background: Riboflavin, an intermediate of primary metabolism, is one kind of important food additive with high economic value. The microbial cell factory Bacillus subtilis has already been proven to possess significant importance for the food industry and have become one of the most widely used riboflavin-producing strains. In the practical fermentation processes, a sharp decrease in riboflavin production is encountered along with a decrease in the dissolved oxygen (DO) tension. Influence of this oxygen availability on riboflavin biosynthesis through carbon central metabolic pathways in B. subtilis is unknown so far. Therefore the unveiled effective metabolic pathways were still an unaccomplished task till present research work., Results: In this paper, the microscopic regulation mechanisms of B. subtilis grown under different dissolved oxygen tensions were studied by integrating 13 C metabolic flux analysis, metabolomics and transcriptomics. It was revealed that the glucose metabolic flux through pentose phosphate (PP) pathway was lower as being confirmed by smaller pool sizes of metabolites in PP pathway and lower expression amount of ykgB at transcriptional level. The latter encodes 6-phosphogluconolactonase (6-PGL) under low DO tension. In response to low DO tension in broth, the glucose metabolic flux through Embden-Meyerhof-Parnas (EMP) pathway was higher and the gene, alsS, encoding for acetolactate synthase was significantly activated that may result due to lower ATP concentration and higher NADH/NAD + ratio. Moreover, ResE, a membrane-anchored protein that is capable of oxygen regulated phosphorylase activity, and ResD, a regulatory protein that can be phosphorylated and dephosphorylated by ResE, were considered as DO tension sensor and transcriptional regulator respectively., Conclusions: This study shows that integration of transcriptomics, 13 C metabolic flux analysis and metabolomics analysis provides a comprehensive understanding of biosynthesized riboflavin's regulatory mechanisms in B. subtilis grown under different dissolved oxygen tension conditions. The two-component system, ResD-ResE, was considered as the signal receiver of DO tension and gene regulator that led to differences between biomass and riboflavin production after triggering the shifts in gene expression, metabolic flux distributions and metabolite pool sizes.