Darobactin is a ribosomally synthesized and post-translationally modified peptide (RiPP), which possesses potent activity against various Gram-negative bacteria. Darobactin features a highly unique bicyclic scaffold, consisting of an ether crosslink between two Trp residues and a C–C crosslink between a Lys and a Trp. Here we report in vivo and in vitro activity of darobactin synthase DarE. We show DarE is a radical S-adenosylmethionine (rSAM) enzyme and is solely responsible for forming the bicyclic scaffold of darobactin. DarE mainly produced the ether-crosslinked product in vitro, and when the assay was performed in H218O, apparent 18O incorporation was observed into the ether-crosslinked product. These observations suggested an rSAM-dependent process in darobactin biosynthesis, involving a highly unusual oxygen insertion step from a water molecule and subsequent O–H and C–H activations. Genome mining analysis demonstrates the diversity of darobactin-like biosynthetic gene clusters, a subclade of which likely encode monocyclic products with only an ether linkage. We propose the name daropeptide for this growing family of ether-containing RiPPs produced by DarE enzymes.

Pharmacotherapy using natural substances can be currently regarded as a very promising future alternative to conventional therapy. With the rapid development of biotechnologies and analytical techniques, a great number of methods have been developed for the identification and quantification of the material, extracts, and products of natural ingredients. The advances available today. The need for safer drugs without side effects has led to the use of natural ingredients with proven safety. In recent years, some bioactive polysaccharides isolated from natural sources have attracted much attention in the field of biochemistry and pharmacology. As an example, polysaccharides or their glycoconjugates were shown to exhibit multiple biological activities including anticarcinogenic, anticoagulant, immunostimulating, antioxidant, etc. During the last several years, we have witnessed a steady expansion in the number of publications that focus in antioxidant polysaccharides. This review presents current findings on the latest advancements and trends in antioxidant polysaccharides isolated from the following: plants, fungi, bacteria, animal sources, and algae. Some interesting studies focus on investigation of the relationship between their structure and antioxidant activity, elucidation of their antioxidant mechanism at the molecular level, and improvement of their various biological activities by chemical modifications. Although the mechanism of their antioxidant action is still not completely clear, these polysaccharides are suggested to enhance cell-mediated immune responses in vivo and in vitro and act as biological response modifiers.

A batch fermentation utilizing Saccharomyces cerevisiae BY4742 was conducted to determine the inhibitory effects of highly concentrated substrate and product levels on yeast. Experiments were performed to determine the largest dosage of substrate and the largest product concentration that the yeast could tolerate in a very high gravity fermentation process. The yeast's growth and fermentation activities were characterized by changes in the biomass and ethanol yield under different substrate and product concentrations during fermentation. All of the experiments were performed at a pH of 5.0 and a temperature of 35°C with a stirring rate of 180 r/min and a fermentation time of 96 h. Furthermore, five cycles of acclimatization were conducted to improve the yeast's tolerance to ethanol. Ethanol yield was maximized at 95% with a product concentration of 39 g/L and substrate dosage of 80 g/L. The system exhibited an obvious increase in cell growth and ethanol production with increasing substrate dosage up to a critical point of 160 g/L glucose (53 g/L ethanol fermented and an ethanol yield of 65%). Above this point, cell growth and ethanol production were inhibited with the final product concentration increasing only slightly with an increase in the initial substrate concentration. The end product (ethanol) was shown to be the primary factor inhibiting yeast growth and fermentation activity because the yeast would completely stop growing and fermenting when the initial exogenous ethanol concentration exceeded 70 g/L. The endogenous ethanol exerted a greater impact on yeast performance during anaerobic fermentation than exogenous ethanol. Five cycles of acclimatization significantly improved the yeast density, cell morphology, and ethanol production during very high gravity fermentation. The ethanol yield increased from 6% to 30% under an initial exogenous ethanol concentration of 60 g/L.

Linaridin is a small class of peptide natural products belonging to the ribosomally synthesized and post-translationally modified peptides (RiPPs) superfamily. By an extensive genome-wide survey of linaridin biosynthetic genes, we show that this class of natural products is widespread in nature and possesses vast structural diversity. The linaridin precursor peptides are relatively conserved in the N-termini but have diverse sequences in the core region, which appear to have coevolved with the biosynthetic enzymes. Using the prototypic linaridin cypemycin as a model, we have explored the structure-activity relationships involved in precursor peptide maturation and generated a diverse set of novel cypemycin variants, among which the T2S variant exhibits enhanced activity against Micrococcus luteus. Our results reveal valuable insights into linaridin biosynthesis and highlight the potential to explore this class of natural products by genome mining and by biosynthetic engineering studies.

Aminomalonate (Ama) is aw idespread structural motif in Nature,whereas its biosynthetic route is only partially understood. In this study,w es howt hat ar adical S-adenosylmethionine (rSAM) enzyme involved in cyclophane biosynthesis exhibits remarkable catalytic promiscuity.This enzyme, named three-residue cyclophane forming enzyme (3-CyFE), mainly produces cyclophane in vivo,w hereas it produces formylglycine (FGly) as am ajor product and barely produce cyclophane in vitro.I mportantly,t he enzyme can further oxidize FGly to produce Ama. Bioinformatic study revealed that 3-CyFEs have evolved from ac ommon ancestor with anaerobic sulfatase maturases (anSMEs), and possess asimilar set of catalytic residues with anSMEs.Remarkably,the enzyme does not need leader peptide for activity and is fully active on at runcated peptide containing only 5a mino acids of the core sequence.Our work discloses the first ribosomal path towards Ama formation, providing ap ossible hint for the rich occurrence of Ama in Nature.