Sebastian Schieferdecker

Structure and Biosynthetic Assembly of Gulmirecins, Macrolide Antibiotics from the Predatory Bacterium Pyxidicoccus fallax

The gulmirecins constitute a new class of glycosylated macrolides that were isolated from the predatory bacterium Pyxidicoccus fallax HKI 727. Their structures were solved by a combination of NMR spectroscopic experiments and chemical derivatization. Analysis of the annotated gulmirecin gene cluster complemented the configurational assignment and provided insights into the stereochemical course of the biosynthetic assembly. The gulmirecins exhibit strong activity against staphylococci, including methicillin-resistant Staphylococcus aureus, but no cytotoxic effects on human cells.

Myxochelins target human 5-lipoxygenase.

Extracts of the predatory myxobacterium Pyxidicoccus fallax HKI 727 showed antiproliferative effects on leukemic K-562 cells. Bioactivity-guided fractionation led to the isolation of the bis-catechol myxochelin A and two new congeners. The biosynthetic origin of myxochelins C and D was confirmed by feeding studies with isotopically labeled precursors. Pharmacological testing revealed human 5-lipoxygenase (5-LO) as a molecular target of the myxochelins. In particular, myxochelin A efficiently inhibited 5-LO activity with an IC50 of 1.9 μM and reduced the proliferation of K-562 cells at similar concentrations.

The 5-lipoxygenase inhibitor RF-22c potently suppresses leukotriene biosynthesis in cellulo and blocks bronchoconstriction and inflammation in vivo.

5-Lipoxygenase (5-LO) catalyzes the first two steps in leukotriene (LT) biosynthesis. Because LTs play pivotal roles in allergy and inflammation, 5-LO represents a valuable target for anti-inflammatory drugs. Here, we investigated the molecular mechanism, the pharmacological profile, and the in vivo effectiveness of the novel 1,2-benzoquinone-featured 5-LO inhibitor RF-22c. Compound RF-22c potently inhibited 5-LO product synthesis in neutrophils and monocytes (IC50⩾22nM) and in cell-free assays (IC50⩾140nM) without affecting 12/15-LOs, cyclooxygenase (COX)-1/2, or arachidonic acid release, in a specific and reversible manner, supported by molecular docking data. Antioxidant or iron-chelating properties were not evident for RF-22c and 5-LO-regulatory cofactors like Ca(2+) mobilization, ERK-1/2 activation, and 5-LO nuclear membrane translocation and interaction with 5-LO-activating protein (FLAP) were unaffected. RF-22c (0.1mg/kg; i.p.) impaired (I) bronchoconstriction in ovalbumin-sensitized mice challenged with acetylcholine, (II) exudate formation in carrageenan-induced paw edema, and (III) zymosan-induced leukocyte infiltration in air pouches. Taken together, RF-22c is a highly selective and potent 5-LO inhibitor in intact human leukocytes with pronounced effectiveness in different models of inflammation that warrants further preclinical analysis of this agent as anti-inflammatory drug.

Harnessing Enzymatic Promiscuity in Myxochelin Biosynthesis for the Production of 5‐Lipoxygenase Inhibitors

The siderophore myxochelin A is a potent inhibitor of human 5‐lipoxygenase (5‐LO). To clarify whether the iron‐chelating properties of myxochelin A are responsible for this activity, several analogues of this compound were generated in the native producer Pyxidicoccus fallax by precursor‐directed biosynthesis. Testing in a cell‐free assay unveiled three derivatives with bioactivity comparable with that of myxochelin A. Furthermore, it became evident that inhibition of 5‐LO by myxochelins does not correlate with their iron affinities.

Structure of Ralsolamycin, the Interkingdom Morphogen from the Crop Plant Pathogen Ralstonia solanacearum

Ralsolamycin, an inducer of chlamydospore formation in fungi, was recently reported from the plant pathogenic bacterium Ralstonia solanacearum. Although interpretation of tandem mass data and bioinformatics enabled a preliminary chemical characterization, the full structure of ralsolamycin was not resolved. We now report the recovery of this secondary metabolite from an engineered R. solanacearum strain. The structure of ralsolamycin was elucidated by extensive spectroscopic analyses. Chemical derivatization as well as bioinformatics were used to assign the absolute stereochemistry. Our results identified an erroneous genome sequence, thereby emphasizing the value of chemical methods to complement bioinformatics-based procedures in natural product research.

In Vivo Dearomatization of the Potent Antituberculosis Agent BTZ043 via Meisenheimer Complex Formation

Nitrobenzothiazinones are among the most potent antituberculosis agents. Herein, we disclose an unprecedented in vivo reduction process that affords Meisenheimer complexes of the clinical candidates BTZ043 and PBTZ169. The reduction is reversible, occurs in all mammalian species investigated, has a profound influence on the in vivo ADME characteristics, and has considerable implications for the design and implementation of clinical studies. The reduction was confirmed by chemical studies that enabled the complete characterization of the Meisenheimer complex and its subsequent chemistry. Combination of the in vivo and chemical studies with LC-MS characterization and assay development also provides a basis for rational lead optimization of this very promising class of antituberculosis agents.

Variochelins, Lipopeptide Siderophores from Variovorax boronicumulans Discovered by Genome Mining

Photoreactive siderophores have a major impact on the growth of planktonic organisms. To date, these molecules have mainly been reported from marine bacteria, although evidence is now accumulating that some terrestrial bacteria also harbor the biosynthetic potential for their production. In this paper, we describe the genomics-driven discovery and characterization of variochelins, lipopeptide siderophores from the bacterium Variovorax boronicumulans, which thrives in soil and freshwater habitats. Variochelins are different from most other lipopeptide siderophores in that their biosynthesis involves a polyketide synthase. We demonstrate that the ferric iron complex of variochelin A possesses photoreactive properties and present the MS-derived structures of two degradation products that emerge upon light exposure.

New myxothiazols from the predatory bacterium Myxococcus fulvus.

Extracts of the predatory bacterium Myxococcus fulvus HKI 722 showed promising antimicrobial activities in the agar diffusion assay. A combined chemical and computational analysis led to the identification of five thiazole-containing antibiotics. Two of the isolated compounds represent previously unrecognized members of the myxothiazol family of natural products. Their antibiotic properties were determined in comparison with those of the known myxothiazols A and Z.

Myxochelin-Inspired 5-Lipoxygenase Inhibitors - Synthesis and Biological Evaluation

A total of 48 analogues of the natural product myxochelin A were prepared and evaluated for their inhibitory effects on human 5‐lipoxygenase in both cell‐free and cell‐based assays. Structure–activity relationship analysis revealed that the secondary alcohol function and only chiral center of myxochelin A is not required for biological activity. By expanding the diaminoalkane linker of the two aromatic residues it was possible to generate a myxochelin derivative with superior activity against 5‐lipoxygenase in intact cells.

Herpetopanone, a diterpene from Herpetosiphon aurantiacus discovered by isotope labeling

The genome of the predatory bacterium Herpetosiphon aurantiacus 114-95T harbors a number of biosynthesis genes, including four terpene cyclase genes. To identify the terpenes biosynthesized from H. aurantiacus 114-95T, we fed the strain with 13C-labeled glucose and, subsequently, searched for characteristic mass shifts in its metabolome. This approach led to the discovery of a new natural product, of which the isotope pattern is indicative for a diterpene originating from the methylerythritol phosphate pathway. After large-scale fermentation of H. aurantiacus 114-95T, the putative diterpene was isolated in sufficient quantity to enable NMR-based structure elucidation. The compound, for which the name herpetopanone is proposed, features a rare octahydro-1H-indenyl skeleton. Herpetopanone bears resemblance to cadinane-type sesquiterpenes from plants, but is structurally entirely unprecedented in bacteria. Based on its molecular architecture, a possible biosynthetic pathway is postulated.