Fused radical SAM and αKG-HExxH domain proteins contain a distinct structural fold and catalyse cyclophane formation and β-hydroxylation.

Two of nature's recurring binding motifs in metalloproteins are the CxxxCxxC motif in radical SAM enzymes and the 2-His-1-carboxylate motif found both in zincins and α-ketoglutarate and non-haem iron enzymes. Here we show the confluence of these two domains in a single post-translational modifying enzyme containing an N-terminal radical S-adenosylmethionine domain fused to a C-terminal 2-His-1-carboxylate (HExxH) domain. The radical SAM domain catalyses three-residue cyclophane formation and is the signature modification of triceptides, a class of ribosomally synthesized and post-translationally modified peptides.

Isolation of new indole alkaloid triglucoside from the aqueous extract of Uncaria rhynchophylla.

Uncaria rhynchophylla (Miq.) Miq. (Rubiaceae) is widely used as a botanical raw material for traditional Japanese and Chinese medicines.

Glutathione degradation activity of γ-glutamyl peptidase 1 manifests its dual roles in primary and secondary sulfur metabolism in Arabidopsis.

Glutathione (GSH) functions as a major sulfur repository and hence occupies an important position in primary sulfur metabolism. GSH degradation results in sulfur reallocation and is believed to be carried out mainly by γ-glutamyl cyclotransferases (GGCT2;1, GGCT2;2, and GGCT2;3), which, however, do not fully explain the rapid GSH turnover. Here, we discovered that γ-glutamyl peptidase 1 (GGP1) contributes to GSH degradation through a yeast complementation assay.

The Biosynthetic Landscape of Triceptides Reveals Radical SAM Enzymes That Catalyze Cyclophane Formation on Tyr- and His-Containing Motifs.

Peptide-derived cyclophanes inhabit a unique niche in the chemical space of macrocyclic peptides with several examples of pharmaceutical importance. Although both synthetic and biocatalytic methods are available for constructing these macrocycles, versatile (bio)catalysts able to incorporate a variety of amino acids that compose the macrocycle would be useful for the creation of diverse peptide cyclophanes. In this report, we synergized the use of bioinformatic tools to map the biosynthetic landscape of radical SAM enzymes (3-CyFEs) that catalyze three-residue cyclophane formation in the biosynthesis of a new family of RiPP natural products, the triceptides.

Retrograde sulfur flow from glucosinolates to cysteine in .

Specialized (secondary) metabolic pathways in plants have long been considered one-way routes of leading primary metabolite precursors to bioactive end products. Conversely, endogenous degradation of such "end" products in plant tissues has been observed following environmental stimuli, including nutrition stress. Therefore, it is of general interest whether specialized metabolites can be reintegrated into primary metabolism to recover the invested resources, especially in the case of nitrogen- or sulfur-rich compounds.

Amphiol, an Antifungal Fungal Pigment from sp. PF1464.

An antifungal metabolite, designated amphiol (), was isolated from the culture broth of the fungus sp. PF1464. It exists as a mixture of inseparable tautomers, an acetal form and a keto form.

Post-translational formation of strained cyclophanes in bacteria.

Cyclic peptide natural products have served as important drug molecules, with several examples used clinically. Enzymatic or chemical macrocyclization is the key transformation for constructing these chemotypes. Methods to generate new and diverse cyclic peptide scaffolds enabling the modular and predictable synthesis of peptide libraries are desirable in drug discovery platforms.

Atypical Myrosinase as a Mediator of Glucosinolate Functions in Plants.

Glucosinolates (GLSs) are a well-known class of specialized plant metabolites, distributed mostly in the order Brassicales. A vast research field in basic and applied sciences has grown up around GLSs owing to their presence in important agricultural crops and the model plant , and their broad range of bioactivities beneficial to human health. The major purpose of GLSs in plants has been considered their function as a chemical defense against predators.

Chemical Interactions of Cryptic Actinomycete Metabolite 5-Alkyl-1,2,3,4-tetrahydroquinolines through Aggregate Formation.

Organisms often produce secondary metabolites as a mixture of biosynthetically related congeners. However, why are metabolites with minor chemical variations produced simultaneously? 5-Alkyl-1,2,3,4-tetrahydroquinolines (5aTHQs) are small, lipophilic metabolites produced by Streptomyces nigrescens HEK616 when cultured with Tsukamurella pulmonis TP-B0596. A mixture of 5aTHQs forms aggregates that show enhanced membrane affinity and biological activity.

Publisher Correction: A cheminformatics approach to characterize metabolomes in stable-isotope-labeled organisms.

In the originally published Supplementary Information for this paper, the files presented as Supplementary Tables 3, 4, and 7 were duplicates of Supplementary Tables 5, 6, and 9, respectively. All Supplementary Table files are now correct online.

A cheminformatics approach to characterize metabolomes in stable-isotope-labeled organisms.

We report a computational approach (implemented in MS-DIAL 3.0; http://prime.psc.

Leaf Endoplasmic Reticulum Bodies Identified in Arabidopsis Rosette Leaves Are Involved in Defense against Herbivory.

ER bodies are endoplasmic reticulum (ER)-derived organelles specific to the order Brassicales and are thought to function in plant defense against insects and pathogens. ER bodies are generally classified into two types: constitutive ER bodies in the epidermal cells of seedlings, and wound-inducible ER bodies in rosette leaves. Herein, we reveal a third type of ER body found in Arabidopsis () rosette leaves and designate them "leaf ERbodies" (L-ER bodies).

Identification of the common biosynthetic gene cluster for both antimicrobial streptoaminals and antifungal 5-alkyl-1,2,3,4-tetrahydroquinolines.

5-Alkyl-1,2,3,4-tetrahydroquinolines (5aTHQs) and streptoaminals (STAMs) are natural products isolated from the combined-culture of Streptomyces nigrescens HEK616 and Tsukamurella pulmonis TP-B0596. Despite their unique structures, their biosynthetic pathway has yet to be elucidated. In the present study, we conducted a feeding experiment using 13C-labeled acetates and demonstrated that 5aTHQs are likely synthesized by the action of polyketide synthase (PKS).

Discovery and Total Synthesis of Streptoaminals: Antimicrobial [5,5]-Spirohemiaminals from the Combined-Culture of Streptomyces nigrescens and Tsukamurella pulmonis.

A series of lipidic spirohemiaminals, designated streptoaminals, is reported. These were discovered by surveying the unique molecular signatures identified in the mass spectrometry data of the combined-culture broth of Streptomyces nigrescens HEK616 and Tsukamurella pulmonis TP-B0596. Mass spectrometry analysis showed that streptoaminals appeared as a cluster of ion peaks, which were separated by 14 mass unit intervals, implying the presence of alkyl chains of different lengths.

Stereochemical Assignment and Biological Evaluation of BE-14106 Unveils the Importance of One Acetate Unit for the Antifungal Activity of Polyene Macrolactams.

Heronamides are a class of potent antifungal metabolites produced by marine-derived actinomycetes. The number of hydroxy groups and the stereochemistry of the two hydroxylated methine carbons are important for the activity of heronamide C, whereas the effect of the hydrocarbon chains is not known. In this study, the stereochemistry and the biological activity of BE-14106, another member of the heronamide class of antibiotics, isolated from an actinomycete Actinoalloteichus cyanogriseus IFM 11549 was investigated.

A novel resting form of the trinuclear copper center in the double mutant of a multicopper oxidase, CueO, Cys500Ser/Glu506Ala.

A multicopper oxidase, CueO was doubly mutated at its type I copper ligand, Cys500 and an acidic amino acid residue located in the proton transfer pathway, Glu506, to Ser and Ala, respectively. Cys500Ser/Glu506Ala was mainly in a novel resting form to afford the absorption band at ca. 400 nm and an EPR signal with a highly anisotropic character derived from type III copper.

5-Alkyl-1,2,3,4-tetrahydroquinolines, new membrane-interacting lipophilic metabolites produced by combined culture of Streptomyces nigrescens and Tsukamurella pulmonis.

Eight novel 5-alkyl-1,2,3,4-tetrahydroquinolines (5aTHQs) bearing different side chains have been isolated from a combined culture of Streptomyces nigrescens HEK616 and Tsukamurella pulmonis TP-B0596. The chemical structures including the absolute configuration were elucidated by spectroscopic analysis and total synthesis. 5aTHQs inhibited the growth of wild-type fission yeast while only weakly inhibiting the growth of several mutant strains synthesizing premature ergosterol.

Structure and biological activity of 8-deoxyheronamide C from a marine-derived Streptomyces sp.: heronamides target saturated hydrocarbon chains in lipid membranes.

Polyene macrolactams are a class of microbial metabolites, many of which show potent biological activities with unidentified modes of action. Here we report that 8-deoxyheronamide C, a new 20-membered polyene macrolactam from a marine-derived actinomycete Streptomyces sp., is a unique membrane binder.

New insights into the catalytic active-site structure of multicopper oxidases.

Structural models determined by X-ray crystallography play a central role in understanding the catalytic mechanism of enzymes. However, X-ray radiation generates hydrated electrons that can cause significant damage to the active sites of metalloenzymes. In the present study, crystal structures of the multicopper oxidases (MCOs) CueO from Escherichia coli and laccase from a metagenome were determined.

ATR-FTIR study of the protonation states of the Glu residue in the multicopper oxidases, CueO and bilirubin oxidase.

Redox-induced protonation state changes of the Glu residue in the multicopper oxidases, CueO and bilirubin oxidase (BO), were studied by attenuated total reflectance-Fourier transform infrared spectroscopy. By monitoring IR bands of the carboxylic acid C=O stretch in the wild-type and Glu-to-Gln mutant enzymes the Glu506 of CueO (Glu463 of BO) was found to be unprotonated in the oxidised and protonated in the reduced forms. The results provided direct evidence for proton uptake by the Glu, suggesting it plays a key role in the proton donation to the activated oxygen species in the catalytic cycle.

Four-electron reduction of dioxygen by a multicopper oxidase, CueO, and roles of Asp112 and Glu506 located adjacent to the trinuclear copper center.

The mechanism of the four-electron reduction of dioxygen by a multicopper oxidase, CueO, was studied based on reactions of single and double mutants with Cys(500), a type I copper ligand, and the noncoordinating Asp(112) and Glu(506), which form hydrogen bonds with the trinuclear copper center directly and indirectly via a water molecule. The reaction of C500S containing a vacant type I copper center produced intermediate I in an EPR-silent peroxide-bound form. The formation of intermediate I from C500S/D112N was restricted due to a reduction in the affinity of the trinuclear copper center for dioxygen.