Synthesis of Daidzein and Thiophene Containing Benzoxazine Resin and Its Thermoset and Carbon Material.

In this work, a novel bio-based high-performance bisbenzoxazine resin was synthesized from daidzein, 2-thiophenemethylamine and paraformaldehyde. The chemical structure was confirmed using nuclear magnetic resonance spectroscopy (NMR) and Fourier-transform infrared spectroscopy (FT-IR). The polymerization process was systematically studied using differential scanning calorimetry (DSC) and in situ FT-IR spectra. It can be polymerized through multiple polymerization behaviors under the synergistic reaction of thiophene rings with benzopyrone rather than a single polymerization mechanism of traditional benzoxazines, as reported. In addition, thermogravimetric analysis (TGA) and a microscale combustion calorimeter (MCC) were used to study the thermal stability and flame retardancy of the resulting polybenzoxazine. The thermosetting material showed a high carbon residue rate of 62.8% and a low heat release capacity (HRC) value of 33 J/gK without adding any flame retardants. Based on its outstanding capability of carbon formation, this newly obtained benzoxazine resin was carbonized and activated to obtain a porous carbon material doped with both sulfur and nitrogen. The CO2 absorption of the carbon material at 0 °C and 25 °C at 1 bar was 3.64 mmol/g and 3.26 mmol/g, respectively. The above excellent comprehensive properties prove its potential applications in many advanced fields.

[Metabolic engineering of Escherichia coli for L-homoserine production].

L-Homoserine is a non-essential amino acid that is often used as an important platform compound and additive in industrial production. To improve the production efficiency, a previously constructed L-homoserine producing strain E. coli H0-0 was used as a chassis for further metabolic modification. Firstly, the ppc and pyccgP458S genes were overexpressed to optimize the Kreb's cycle. Subsequently, thrAC1034T and lysCcgC932T were overexpressed to improve the product synthesis, followed by inactivation of iclR gene to reduce the accumulation of by-products. The introduction of three sucrose metabolism genes, scrA, scrB and scrK, enabled E. coli to ferment sucrose. The titer of L-homoserine increased from 3.2 g/L to 11.1 g/L.

Multiplex Design of the Metabolic Network for Production of l-Homoserine in Escherichia coli.

l-Homoserine, which is one of the few amino acids that is not produced on a large scale by microbial fermentation, plays a significant role in the synthesis of a series of valuable chemicals. In this study, systematic metabolic engineering was applied to target Escherichia coli W3110 for the production of l-homoserine. Initially, a basic l-homoserine producer was engineered through the strategies of overexpressing thrA (encoding homoserine dehydrogenase), removing the degradative and competitive pathways by knocking out metA (encoding homoserine O-succinyltransferase) and thrB (encoding homoserine kinase), reinforcing the transport system, and redirecting the carbon flux by deleting iclR (encoding the isocitrate lyase regulator). The resulting strain constructed by these strategies yielded 3.21 g/liter of l-homoserine in batch cultures. Moreover, based on CRISPR-Cas9/dCas9 (nuclease-dead Cas9)-mediated gene repression for 50 genes, the iterative genetic modifications of biosynthesis pathways improved the l-homoserine yield in a stepwise manner. The rational integration of glucose uptake and recovery of l-glutamate increased l-homoserine production to 7.25 g/liter in shake flask cultivation. Furthermore, the intracellular metabolic analysis further provided targets for strain modification by introducing the anaplerotic route afforded by pyruvate carboxylase to oxaloacetate formation, which resulted in accumulating 8.54 g/liter l-homoserine (0.33 g/g glucose, 62.4% of the maximum theoretical yield) in shake flask cultivation. Finally, a rationally designed strain gave 37.57 g/liter l-homoserine under fed-batch fermentation, with a yield of 0.31 g/g glucose.IMPORTANCE In this study, the bottlenecks that sequentially limit l-homoserine biosynthesis were identified and resolved, based on rational and efficient metabolic-engineering strategies, coupled with CRISPR interference (CRISPRi)-based systematic analysis. The metabolomics data largely expanded our understanding of metabolic effects and revealed relevant targets for further modification to achieve better performance. The systematic analysis strategy, as well as metabolomics analysis, can be used to rationally design cell factories for the production of highly valuable chemicals.