Trans-4-Hydroxy-l-proline (trans-Hyp) is a valuable chiral building block for the synthesis of pharmaceutical intermediates. Unsatisfactory microbial bioconversion resulted in a low yield of trans-Hyp. In this study, we blocked the unwanted blunting pathways of host strain and make the cell growth couple with the trans-Hyp synthesis from glucose. Finally, a recombinant Escherichia coli with short-cut and efficient trans-Hyp biosynthetic pathway was obtained. It provided a theoretical basis for commercial production of trans-Hyp.

Aims:We aimed to explore Yarrowia lipolytica carbonyl reductases as effective biocatalysts and to develop efficient asymmetric reduction systems for chiral alcohol synthesis. Methods and Results: Yarrowia lipolytica carbonyl reductase genes were obtained via homologous sequence amplification strategy. Two carbonyl reductases, YaCRI and YaCRII, were identified and characterized, and used to catalyse the conversion of 2-hydroxyacetophenone (2-HAP) to optically pure (S)-1-phenyl-1,2-ethanediol. Enzymatic assays revealed that YaCRI and YaCRII exhibited specific activities of 6Á96 U mg À1 (99Á8% e.e.) and 7Á85 U mg À1 (99Á9% e.e.), respectively, and showed moderate heat resistance at 40-50°C and acid tolerance at pH 5Á0-6Á0. An efficient whole-cell two-phase system was established using reductase-expressing recombinant Escherichia coli. The conversion of 2-HAP (20Á0 g l À1 ) conversion with the solvent of dibutyl phthalate was approximately 70-fold higher than in water. Furthermore, the two recombinant E. coli displayed biocatalyst activity and enantioselectivity towards several different carbonyl compounds, and E. coli BL21 (DE3)/pET-28a-yacrII showed a broad substrate spectrum. Conclusions: A new whole-cell recombinant E. coli-based bioreduction system for enantiopure alcohol synthesis with high enantioselectivity at high substrate concentrations was developed. Significance and Impact of the Study: We proposed a promising approach for the efficient preparation of enantiopure chiral alcohols.

Medicago sativa L. (alfalfa) is an important forage crop throughout the world. Despite the abiotic nutritional components of alfalfa having been extensively studied, there is only limited information on alfalfa endophytes. In this study, thirteen endophytic bacteria were isolated from alfalfa seeds. Bacillus (76·9%) was the most abundant genus, followed by Enterobacter (15·4%), Brevibacterium (7·7%), Geobacillus (7·7%) and Staphylococcus (7·7%). Four of the 13 endophytic bacteria, including Bacillus amyloliquefaciens EnB‐alf1, Bacillus subtilis EnB‐alf3, EnB‐alf5 and EnB‐alf13, were capable of significantly extending the lifespan of Caenorhabditis elegans. In addition, B. amyloliquefaciens EnB‐alf1 enhanced the resistance of C. elegans to thermal stress whereas B. subtilis EnB‐alf3 enhanced the resistance to oxidative stress. Further studies demonstrated that the enhanced lifespan of the worm was depended on the function of DAF‐2/DAF‐16 and was associated with the colonization of strain in the worms’ intestines when strain EnB‐alf1 or strain EnB‐alf3 was presented to the worms as food sources. Our results suggest that some endophytic Bacillus strains isolated from alfalfa are beneficial on C. elegans health. Significance and Impact of the Study Medicago sativa L. (alfalfa) is an important forage crop throughout the world. Despite the abiotic nutritional components of alfalfa having been extensively studied, there is only limited information available on alfalfa endophytes. Beneficial bacteria residing in the host intestine have been shown to affect host longevity. However, there is limited information available on the functions of alfalfa seed endophytes to nematodes. In this study, four endophytic Bacillus strains isolated from alfalfa seeds were found to significantly extend the lifespan of Caenorhabditis elegans and enhance resistance to thermal and oxidative stress. Our results suggest that some endophytic Bacillus strains isolated from alfalfa seeds can promote good health in C. elegans.

Amphiphilic core–shell (ACS) nanoparticles are gaining increasing research interest for multi-drug delivery in cancer therapy. In this work, a new cationic peptide-coated PHA nanosphere was prepared by self-assembly of a hydrophobic core of biodegradable poly (3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) and a hydrophilic shell of fusion proteins of PHA granule-associated protein (PhaP) and cationic peptide RALA through a strong hydrophobic effect. The hydrophobic drug curcumin (Cur) was encapsulated in PHBHHx nanoparticles. The chemotherapy drug 5-fluorouracil (5-FU) was administered in the form of its metabolite oligomeric 5-fluorodeoxyuridine (FUdR). Fifteen consecutive FUdR (FUdR15S) were adsorbed on the surface of PHBHHx nanoparticles by electrostatic interaction with RALA to form Cur@PHBX-PR/FUdR15S. Such amphiphilic cationic nanospheres had 88.3% EE of Cur and the drug loading of Cur and FUdR were 7.8% and 12.1%. The dual-drug-loaded nanospheres showed a time-differential release of Cur and FUdR. In addition, Cur@PHBX-PR/FUdR15S exhibited excellent anticancer activity and played a vital role in promoting the synergistic effect of FUdR and Cur in gastric cancer cells. The exploration of antitumor mechanisms demonstrated that Cur improved the activity of apoptosis-related proteins and cancer cells sensitized to FUdR. This amphiphilic core–shell system can serve as a general platform for sequential delivery of multiple drugs to treat several cancer cells.

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