Metabolic Engineering of Escherichia coli for High-Level Production of Lacto‑<i>N</i>‑neotetraose and Lacto‑<i>N</i>‑tetraose

Lacto-N-neotetraose (LNnT) and lacto-N-tetraose (LNT) are important oligosaccharides found in breast milk and are commonly used as nutritional supplements in infant formula. We used metabolic engineering techniques to optimize the modified Escherichia coli BL21 star (DE3) strain for efficient synthesis of LNnT and LNT using β-1,4-galactosyltransferase (HpgalT) from Helicobacter pylori and β-1,3-galactosyltransferase (SewbdO) from Salmonella enterica subsp. salamae serovar, respectively. Further, we optimized the expression of three key genes, lgtA, galE, and HpgalT (SewbdO), to synthesize LNnT or LNT and deleted several genes (ugd, ushA, agp, wcaJ, otsA, and wcaC) to block competition in the UDP-galactose synthesis pathway. The optimized strain produced LNnT or LNT with a titer of 22.07 or 48.41 g/L, respectively, in a supplemented batch culture, producing 0.41 or 0.73 g/L/h, respectively. The strategies used in this study contribute to the development of cell factories for high-level LNnT and LNT and their derivatives

A Highly Ordered Meso@Microporous Carbon-Supported Sulfur@Smaller Sulfur Core–Shell Structured Cathode for Li–S Batteries

For lithium–sulfur batteries, commercial application is hindered by the insulating nature of sulfur and the dissolution of the reaction intermediates of polysulfides. Here, we present an ordered meso-microporous core–shell carbon (MMCS) as a sulfur container, which combines the advantages of both mesoporous and microporous carbon. With large pore volume and highly ordered porous structure, the “core” promises a sufficient sulfur loading and a high utilization of the active material, while the “shell” containing microporous carbon and smaller sulfur acts as a physical barrier and stabilizes the cycle capability of the entire S/C composite. Such a S/MMCS composite exhibits a capacity as high as 837 mAh g^–1 at 0.5 C after 200 cycles with a capacity retention of 80% vs the second cycle (a decay of only 0.1% per cycle), demonstrating that the diffusion of the polysulfides into the bulk electrolyte can be greatly reduced. We believe that the tailored highly ordered meso-microporous core–shell structured carbon can also be applicable for designing some other electrode materials for energy storage

Free-Standing Mn₃O₄@CNF/S Paper Cathodes with High Sulfur Loading for Lithium–Sulfur Batteries

Free-standing paper cathodes with layer-by-layer structure are synthesized for high-loading lithium–sulfur (Li–S) battery. Sulfur is loaded in a three-dimensional (3D) interconnected nitrogen-doped carbon nanofiber (CNF) framework impregnated with Mn₃O₄ nanoparticles. The 3D interconnected CNF framework creates an architecture with outstanding mechanical properties. Synergetic effects generated from physical and chemical entrapment could effectively suppress the dissolution and diffusion of the polysulfides. Electrochemical measurements suggest that the rationally designed structure endows the electrode with high utilization of sulfur and good cycle performance. Specifically, the cathode with a high areal sulfur loading of 11 mg cm^–2 exhibits a reversible areal capacity over 8 mAh cm^–2. The fabrication procedure is of low cost and readily scalable. We believe that this work will provide a promising choice for potential practical applications

Thermodynamic Difference between Protocatechualdehyde and <i>p</i>‑Hydroxybenzaldehyde in Aqueous Sodium Chloride Solutions

The enthalpies of dilution of protocatechualdehyde and <i>p</i>-hydroxybenzaldehyde in the aqueous sodium chloride solutions were measured by using a mixing-flow microcalorimeter at 298.15 K. Densities of the ternary homogeneous systems at different temperatures (293.15, 298.15, 303.15, 308.15, and 313.15 K) were also measured with a quartz vibrating-tube densimeter. The homogeneous enthalpic interaction coefficients (<i>h</i>₂, <i>h</i>₃, and <i>h</i>₄) were calculated according to the excess enthalpy concept based on the calorimetric data. The apparent molar volumes (<i>V</i>_ϕ) and standard partial molar volumes (<i>V</i>_ϕ⁰) of the investigated system were computed from their density data. The variation trends in <i>h</i>₂ and <i>V</i>_ϕ⁰ with increasing salt molality were obtained and discussed in terms of the (solute + solute) and (solute + solvent) interactions. The experimental results showed that the molecular structures of protocatechualdehyde and <i>p</i>-hydroxybenzaldehyde, especially the number of hydroxyl groups, have evident influence on their thermodynamic properties. The thermodynamic data obtained in this work may be helpful for exploring the structure–function relationship of protocatechualdehyde and <i>p</i>-hydroxybenzaldehyde

High-Yield Synthesis of Lacto‑<i>N</i>‑Neotetraose from Glycerol and Glucose in Engineered Escherichia coli

Lacto-N-neotetraose (LNnT) is a neutral human milk oligosaccharide with important biological functions. However, the low LNnT productivity and the incomplete conversion of the intermediate lacto-N-tetraose II (LNT II) currently limited the sustainable biosynthesis of LNnT. First, the LNnT biosynthetic module was integrated in Escherichia coli. Next, the LNnT export system was optimized to alleviate the inhibition of intracellular LNnT synthesis. Furthermore, by utilizing rate-limiting enzyme diagnosis, the expressions of LNnT synthesis pathway genes were finely regulated to further enhance the production yield of LNnT. Subsequently, a strategy of cofermentation using a glucose/glycerol (4:6, g/g) mixed feed was employed to regulate carbon flux distribution. Finally, by overexpressing key transferases, LNnT and LNT II titers reached 112.47 and 7.42 g/L, respectively, in a 5 L fermenter, and 107.4 and 2.08 g/L, respectively, in a 1000 L fermenter. These are the highest reported titers of LNnT to date, indicating its significant potential for industrial production

Additional file 1 of Chemoproteomics-based profiling reveals potential antimalarial mechanism of Celastrol by disrupting spermidine and protein synthesis

Additional file 1: Fig. S1. The antimalarial activity of Cel and Cel-P against P. falciparum Dd2 strain. Fig. S2. Heatmap representation of the proteome after Celastrol treatment. Fig. S3. The absorbance spectra of increasing concentration Celastrol. Fig. S4. A Heatmap representation of the decreased expression of parasite proteins after Cel treatment. B GO enrichment analysis of the decreased expression proteins. Fig. S5. The antimalarial activity of Cel against artemisinin-sensitive (P. falciparum 3D7) and artemisinin-resistant strains (P. falciparum 6320). Fig. S6. Raw data of all gel images and Western blots