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A one-step enzymatic desymmetrization process was developed to produce the direct chiral precursor for S-pregabalin synthesis with high yield and high eep value.

Hazardous materials, such as heavy metals, are the major sources of health risk. Using genetically modified organisms (GMOs) to dispose heavy metals has the advantages of strong environmental compatibility and high efficiency. However, the biosecurity of GMOs used in the environment is a major concern. In this study, a self-controlled genetic circuit was designed and carefully fine-tuned for programmable expression in Pseudomonas putida KT2440, which is a widely used strain for environmental bioremediation. The cell behaviours were controlled by automatically sensing the variation of Hg

Zearalenone (ZEN) is one of the common mycotoxins with quite high occurrence rate and is harmful to animal and human health. Lactobacillus reuteri is known as a probiotic bacterium with active immune stimulating and high inhibitory activity against pathogenic microorganisms. In this study, we expressed the lactonohydrolase from Rhinocladiella mackenziei CBS 650.93 (RmZHD) in L. reuteri via secretion and surface-display patterns, respectively. Endogenous signal peptides from L. reuteri were first screened to achieve high expression for efficient ZEN hydrolysis. For secretion expression, signal peptide from collagen-binding protein showed the best performance, while the one from fructose-2,6-bisphosphatase worked best for surface-display expression. Both of the engineered strains could completely hydrolyze 5.0 mg/L ZEN in 8 h without detrimental effects on bacterial growth. The acid and bile tolerance assay and anchoring experiment on Caco-2 cells indicated both of the abovementioned engineered strains could survive during digestion and colonize on intestinal tract, in which the surface-displayed strain had a better performance on ZEN hydrolysis. Biodetoxification of model ZEN-contaminated maize kernels showed the surface-displayed L. reuteri strain could completely hydrolyze 2.5 mg/kg ZEN within 4 h under low water condition. The strain could also efficiently detoxify natural ZEN-contaminated corn flour in the in vitro digestion model system. The colonized property, survival capacity, and the efficient hydrolysis performance as well as probiotic functionality make L. reuteri strain an ideal host for detoxifying residual ZEN in vivo, which shows a great potential for application in feed industry.

Plants produce a wide variety of secondary metabolites that are highly nutraceutically and pharmaceutically important. Isothiocyanates, which are found abundantly in cruciferous vegetables, are believed to reduce the risk of several types of cancers and cardiovascular diseases. The challenges arising from the structural diversity and complex chemistry of these compounds have spurred great interest in producing them in large amounts in microbes. In this study, we aimed to synthesize benzyl isothiocyanate in Escherichia coli via gene mining, pathway engineering, and protein modification. Two chimeric cytochrome P450 enzymes were constructed and functionally expressed in E. coli. The E. coli cystathionine β-lyase was used to replace the plant-derived C-S lyase; its active form cannot be expressed in E. coli. Suitable desulfoglucosinolate:PAPS sulfotransferase from Arabidopsis thaliana ecotype Col-0 and myrosinase from Brevicoryne brassicae were successfully mined from the database. Biosynthesis of benzyl isothiocyanate by the combined expression of the optimized enzymes in vitro was confirmed by gas chromatography-mass spectrometry analysis. This study provided a proof of concept for the production of benzyl isothiocyanate by microbially produced enzymes and, importantly, laid the groundwork for further metabolic engineering of microbial cells for the production of isothiocyanates.

Epidemiological data confirmed a strong correlation between regular consumption of cruciferous vegetables and lower cancer risk. This cancer preventive property is mainly attributed to the glucosinolate products, such as glucoraphanin found in broccoli that is derived from methionine. Here we report the first successful reconstruction of the complete biosynthetic pathway of glucoraphanin from methionine in Escherichia coli via gene selection, pathway design, and protein engineering. We used branched-chain amino transferase 3 to catalyze two transamination steps to ensure the purity of precursor molecules and used cysteine as a sulfur donor to simplify the synthesis pathway. Two chimeric cytochrome P450 enzymes were engineered and expressed in E. coli functionally. The original plant C-S lyase was replaced by the Neurospora crassa hercynylcysteine sulfoxide lyase. Other pathway enzymes were successfully mined from Arabidopsis thaliana, Brassica rapa, and Brassica oleracea. Biosynthesis of glucoraphanin upon coexpression of the optimized enzymes in vivo was confirmed by liquid chromatography-tandem mass spectrometry analysis. No other glucosinolate analogues (except for glucoiberin) were identified that could facilitate the downstream purification processes. Production of glucoraphanin in this study laid the foundation for microbial production of such health-beneficial glucosinolates in a large-scale.

Background Chlorophenols are environmental contaminants, which are highly toxic to living beings due to their carcinogenic, mutagenic and cytotoxic properties. Bacterial degradation has been considered a cost-effective and eco-friendly method of removing chlorophenols, compared to the traditional physical–chemical processes. Results In this study, we first developed an efficient process for the biodegradation of chlorophenols by magnetically immobilized Rhodococcus rhodochrous cells. R. rhodochrous DSM6263 degrades chlorophenols following the first step of hydroxylation at the ortho-positions of chlorophenolic rings. The cells immobilized by k-carrageenan with 9 g/L Fe3O4 nanoparticles could efficiently degrade 2-chlorophenol, 4-chlorophenol, 2,3-dichlorophenol and their mixture, which were even higher than those by free cells. The magnetically nanoparticle-immobilized cells could be used at least for six cycles. Conclusion Given the much easier separation by an external magnetic field and high degradation efficiency, this study provides a promising technique for improving biocatalysts used in the bioremediation process for chlorophenols in wastewater.

Reuterin is a compound that contains the monomeric, hydrated monomeric, and cyclic dimeric forms of 3-hydroxypropionaldehyde (3-HPA). It is widely used as a food preservative due to its antimicrobial properties, and 3-HPA is also an important precursor for several industrial chemicals. In this study, we first developed an efficient immobilization process involving magnetic cell entrapment for production of reuterin from glycerol by Lactobacillus reuteri. The cell growth condition was optimized by statistical approaches. High conversion efficiency was achieved using k-carrageenan as the immobilization support and Fe3O4 as magnetic nanoparticles. Furthermore, addition of 0.02 g/L vitamin B12, 10 mmol/L Mg(2+), and glucose in a molar ratio of 0.15 to glycerol significantly increased the conversion rate. Under optimal conditions, 235.9 mmol/L 3-HPA was produced from 250 mmol/L initial glycerol in 1.5 h, with a molar yield of 94.4 % and a productivity of 15.4 mmol/(g dry cell weight∙L∙h), which were the highest values to date. Thus, this study demonstrated a promising process for the improvement of biocatalyst efficiency in biotransformation.

The purpose of this study was to identify main influencing parameters to the responses of the rear-seat female occupants in frontal crash and enhance the protection for rear-seat female occupants with specific design. The results suggested parameters of the restraint system exerted significant influence on the injury severity of the female occupant in rear seats. After parameter optimization, the neck score, the chest score and the general score of the female dummy based on the assessment protocol for adult occupant protection of Euro-NCAP increased by 3.6%, 13.3% and 3.73%, respectively. Protection of female occupants in rear seat could be enhanced by systematically matching the parameters of the restraint system.