N2O Reduction During Denitrifying Phosphorus Removal With Propionate as Carbon Source

Denitrifying phosphorus removal was realized in sequencing batch reactors using different carbons sources (acetate, propionate, and a mixture of acetate/propionate). Nutrient removal and N2O production were investigated, and the factors affecting N2O production were explored. Nitrogen removal was 40.6% lower when propionate was used as the carbon source instead of acetate, while phosphorus removal was not significantly different. N2O production was greatly reduced when propionate was used as the carbon source instead of acetate. The emission factor in the propionate system was only 0.43%, while those in the acetate and mixed-carbon source system were 16.3% and 1.9%, respectively. Compared to the propionate system, ordinary heterotrophic organisms (i.e., glycogen-accumulating organisms) were enriched in the acetate system, explaining the higher N2O production in the acetate system. The lower nitrite accumulation in the propionate system compared to the acetate system was the dominant factor leading to the lower N2O production.

Efficient production of myo-inositol in Escherichia coli through metabolic engineering

Background: The biosynthesis of high value-added compounds using metabolically engineered strains has received wide attention in recent years. Myo-inositol (inositol), an important compound in the pharmaceutics, cosmetics and food industries, is usually produced from phytate via a harsh set of chemical reactions. Recombinant Escherichia coli strains have been constructed by metabolic engineering strategies to produce inositol, but with a low yield. The proper distribution of carbon flux between cell growth and inositol production is a major challenge for constructing an efficient inositol-synthesis pathway in bacteria. Construction of metabolically engineered E. coli strains with high stoichiometric yield of inositol is desirable.Results: In the present study, we designed an inositol-synthesis pathway from glucose with a theoretical stoichiometric yield of 1 mol inositol/mol glucose. Recombinant E. coli strains with high stoichiometric yield (>0.7 mol inositol/mol glucose) were obtained. Inositol was successfully biosynthesized after introducing two crucial enzymes: inositol-3-phosphate synthase (IPS) from Trypanosoma brucei, and inositol monophosphatase (IMP) from E. coli. Based on starting strains E. coli BW25113 (wild-type) and SG104 (ΔptsG::glk, ΔgalR::zglf, ΔpoxB::acs), a series of engineered strains for inositol production was constructed by deleting the key genes pgi, pfkA and pykF. Plasmid-based expression systems for IPS and IMP were optimized, and expression of the gene zwf was regulated to enhance the stoichiometric yield of inositol. The highest stoichiometric yield (0.96 mol inositol/mol glucose) was achieved from recombinant strain R15 (SG104, Δpgi, Δpgm, and RBSL5-zwf). Strain R04 (SG104 and Δpgi) reached high-density in a 1-L fermenter when using glucose and glycerol as a mixed carbon source. In scaled-up fed-batch bioconversion in situ using strain R04, 0.82 mol inositol/mol glucose was produced within 23 h, corresponding to a titer of 106.3 g/L (590.5 mM) inositol.Conclusions: The biosynthesis of inositol from glucose in recombinant E. coli was optimized by metabolic engineering strategies. The metabolically engineered E. coli strains represent a promising method for future inositol production. This study provides an essential reference to obtain a suitable distribution of carbon flux between glycolysis and inositol synthesis.

Efficient production of myo-inositol in Escherichia coli through metabolic engineering

Background: The biosynthesis of high value-added compounds using metabolically engineered strains has received wide attention in recent years. Myo-inositol (inositol), an important compound in the pharmaceutics, cosmetics and food industries, is usually produced from phytate via a harsh set of chemical reactions. Recombinant Escherichia coli strains have been constructed by metabolic engineering strategies to produce inositol, but with a low yield. The proper distribution of carbon flux between cell growth and inositol production is a major challenge for constructing an efficient inositol-synthesis pathway in bacteria. Construction of metabolically engineered E. coli strains with high stoichiometric yield of inositol is desirable.Results: In the present study, we designed an inositol-synthesis pathway from glucose with a theoretical stoichiometric yield of 1 mol inositol/mol glucose. Recombinant E. coli strains with high stoichiometric yield (>0.7 mol inositol/mol glucose) were obtained. Inositol was successfully biosynthesized after introducing two crucial enzymes: inositol-3-phosphate synthase (IPS) from Trypanosoma brucei, and inositol monophosphatase (IMP) from E. coli. Based on starting strains E. coli BW25113 (wild-type) and SG104 (ΔptsG::glk, ΔgalR::zglf, ΔpoxB::acs), a series of engineered strains for inositol production was constructed by deleting the key genes pgi, pfkA and pykF. Plasmid-based expression systems for IPS and IMP were optimized, and expression of the gene zwf was regulated to enhance the stoichiometric yield of inositol. The highest stoichiometric yield (0.96 mol inositol/mol glucose) was achieved from recombinant strain R15 (SG104, Δpgi, Δpgm, and RBSL5-zwf). Strain R04 (SG104 and Δpgi) reached high-density in a 1-L fermenter when using glucose and glycerol as a mixed carbon source. In scaled-up fed-batch bioconversion in situ using strain R04, 0.82 mol inositol/mol glucose was produced within 23 h, corresponding to a titer of 106.3 g/L (590.5 mM) inositol.Conclusions: The biosynthesis of inositol from glucose in recombinant E. coli was optimized by metabolic engineering strategies. The metabolically engineered E. coli strains represent a promising method for future inositol production. This study provides an essential reference to obtain a suitable distribution of carbon flux between glycolysis and inositol synthesis.

Efficient production of myo-inositol in Escherichia coli through metabolic engineering

Background The biosynthesis of high value-added compounds through metabolically engineered strains has received widely attention in recent years. As an effective compound in pharmaceutical, cosmetic and food industry, myo-inositol (inositol) is mainly produced via a harsh set of chemical reactions from phytate. The proper distribution of carbon flux between cell growth and inositol production was a major challenge for constructing an efficient inositol-synthetic pathway. Recombinant E. coli strains have been constructed by metabolic engineering strategies to produce inositol, yet with a low yield. Therefore, construction of E. coli metabolically engineered strains with high stoichiometric yield will be attractive. Results In the present study, the recombinant E. coli strains with high stoichiometric yield (> 0.7 mol inositol/mol glucose) were obtained to efficiently synthesize inositol. Inositol was successfully biosynthesized after introducing two crucial enzymes, inositol-3-phosphate synthase (IPS) from Trypanosoma brucei , and inositol monophosphatase (IMP) from E. coli. Based on starting strains E. coli BW25113 (wild type) and SG104 ( ΔptsG::glk , ΔgalR::zglf , ΔpoxB::acs ), a series of engineered strains for inositol production were constructed by deleting the key genes pgi, pfkA or pykF . Furthermore, the plasmid expression systems of IPS and IMP were optimized, and the gene zwf was regulated to enhance stoichiometric yield. The highest stoichiometric yield (0.96 mol inositol/mol glucose) was achieved with the combined strain R15 of SG104, Δpgi , Δpgm , and RBSL5-zwf. Simultaneously, the engineered strain R04 reached high-density fermentation level in a 1-L fermenter by using glucose and glycerol as mixed carbon source. In the scale-up bioconversion in situ with R04, 0.82 mol inositol/mol glucose was produced by fed-batch within 23 h, corresponding to a titer of 106.3 g/L (590.5 mM). Conclusions The biosynthetic pathway of inositol from glucose in recombinant E. coli was optimized by metabolic engineering strategies. The metabolically engineered E. coli strains represent a promising method for future inositol production. This study provided an essential reference to obtain a suitable distribution of carbon flux between glycolysis pathway and product synthetic pathway.

Mixed carbon source improves deep denitrification performance in up-flow anaerobic sludge bed reactor

To investigate the advantages of mixed carbon source over a single one in deep denitrification, sodium acetate, glucose and their mixture were used as carbon sources in present study. Denitrification performance, effluent pH, microbial community and carbon source cost were taken into account. With the same influent NO3–-N concentration of 50 mg/L and the same C/N ratio of 1.5, the NO3–-N removal rate with the mixed carbon source (96.53%) was slightly lower than that with sodium acetate (98.15%), but significantly higher than that with glucose (74.69%). The specific denitrification rates of the sodium acetate, glucose and sodium acetate/glucose reactor were 47.7, 29.7 and 45.4 mg N/g VSS d, respectively. The effluent pH with sodium acetate varied in the range of 9.13–9.60, exceeding the discharge standard limit of 9.0, whereas the sodium acetate/glucose reactor could keep pH in the range of 7.80–8.23. The 16S rRNA gene-based high-throughput sequencing revealed that carbon sources determined the microbial community structure and the sludge Shannon index with the mixed carbon source was the highest. Furthermore, cost estimation indicated that the mixed carbon source was the cheapest. This study is significant as it tests reasonable selection of carbon sources for deep denitrification in practice.