Violacein is a natural purple pigment of Chromobacterium violaceum with potential medical applications as antimicrobial, antiviral, and anticancer drugs. The initial step of violacein biosynthesis is the oxidative conversion of L-tryptophan into the corresponding ␣-imine catalyzed by the flavoenzyme L-tryptophan oxidase (VioA). A substrate-related (3-(1H-indol-3-yl)-2-methylpropanoic acid) and a product-related (2-(1H-indol-3-ylmethyl)prop-2-enoic acid) competitive VioA inhibitor was synthesized for subsequent kinetic and x-ray crystallographic investigations. Structures of the binary VioA⅐FADH 2 and of the ternary VioA⅐FADH 2 ⅐2-(1H-indol-3-ylmethyl)prop-2-enoic acid complex were resolved. VioA forms a "loosely associated" homodimer as indicated by small-angle x-ray scattering experiments. VioA belongs to the glutathione reductase family 2 of The amino acid tryptophan (Trp) is a versatile metabolite that is shunted into several microbial secondary pathways. The oxidative dimerization of two Trp scaffolds is an essential step for the synthesis of the natural bisindole compounds violacein, rebeccamycin, and staurosporine. The water-insoluble purple pigment violacein is mainly produced by the bacteria Chromobacterium violaceum and Janthinobacterium lividum in tropical habitats where it might be responsible for the shielding against UV radiation (1-3). Various biological activities of violacein, including antibacterial, antiviral, as well as cytotoxic effects against several tumor cell lines, have been demonstrated (4). The related compounds rebeccamycin and staurosporine show strong antitumorigenic activity because of DNA topoisomerase or protein kinase inhibition (5, 6).Synthesis of violacein, rebeccamycin, or staurosporine is based on a closely related initial pathway in which orthologous enzymes catalyze the oxidation of a Trp moiety into the related indole-3-pyruvic acid (IPA) 2 imine by the enzymes VioA, RebO, or StaO ( Fig. 1) (7-9). Subsequently, oxidative coupling of two imines by VioB, RebD, or StaD results in the formation of a short-lived compound that was proposed to be an IPA imine dimer (7, 10). For the synthesis of rebeccamycin and staurosporine, this reactive intermediate is spontaneously converted into chromopyrrolic acid (11-13). By contrast, violacein biosynthesis requires a key intramolecular rearrangement. The postulated IPA imine dimer is the substrate of VioE, which is catalyzing a [1,2]-shift of the indole ring to produce protodeoxyviolaceinic acid (7,14). Fig. 1 gives a schematic overview about the related pathways as follows: common enzymatic reactions and the involved cofactors are highlighted (gray shading), subsequent steps for the synthesis of violacein (VioE, VioD, VioC, and one autocatalytic step) (3, 7), rebeccamycin (RebP, RebC, RebG, and RebM) (15, 16), and staurosporine (StaP, StaC, StaG, StaN, StaMA, and StaMB) (16) are indicated.Synthesis of violacein starts with the FAD-dependent oxidation of L-Trp to IPA imine catalyzed by VioA. The VioA protein from C. violaceum shares a su...

Macrolides are a relatively common structural motif prevalent in Nature. However, the structures of these large ring lactones have been relatively difficult to elucidate via NMR spectroscopy due to the minute amounts of compounds that are sometimes obtainable from natural sources. Thus, GC-MS analysis of individual macrolactones has become the method of choice for the structural identification of these compounds. Here we discuss the mass spectrometric behavior of aliphatic macrolides, evaluating spectra from numerous compounds of various ring size, including derivatives containing methyl branches as well as double bonds. The specific fragmentation of these macrolactones under electron impact conditions allows for the development of a general rule set aimed at the identification of similar compounds by mass spectrometry. In addition, the mass spectra of dimethyl disulfide adducts of unsaturated macrolides are discussed. The mass spectra of almost 50 macrolides are presented.

For a long time, frogs were believed to communicate primarily via the acoustic channel, but during the last decades it became obvious that various lineages also use chemical communication. In this account we will present our research on the identification of volatile lactones from Madagascan Mantellidae and African Hyperoliidae frogs. Both possess male specific glands that can disseminate a range of volatile compounds. Key constituents are macrocyclic lactones. They show high variability in structure and occurrence. We will focus here on the synthetic approaches we have used to clarify constitution and configuration of the glandular compounds. Key synthetic methods are ring closing metathesis and nucleophilic epoxide opening. Often, but not always, the natural compounds occurs in amounts that excludes their investigation by NMR. Instead, we use GC/MS analysis, GC/IR, microreactions and synthesis to identify such components. Several aspects of our work will be described giving some insight in our scientific approach.

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