Gerhard Lang

Linear and Cyclic Peptides from the Entomopathogenic Bacterium Xenorhabdus nematophilus

Three new peptides, xenortides A and B and xenematide, were isolated from a culture of the nematode-associated entomopathogenic bacterium Xenorhabdus nematophilus. Their structures were elucidated using NMR, MS, and chemical derivatization methods. Xenortides A and B are the N-phenethylamide and tryptamide derivatives, respectively, of the dipeptide (NMe-L-Leu-NMe-L-Phe). The cyclodepsipeptide xenematide has the sequence (Thr-Trp-Trp-Gly), with a 2-phenylacetamide substituent at the threonine residue and one d-tryptophan. The new peptides and the two known compounds xenocoumacin II and nematophin were tested for antibacterial, antifungal, insecticidal, and anti-Artemia salina activities. Xenematide and xenocoumacin II showed moderate antibacterial activities. Xenocoumacin II, nematophin, and the two xenortides were active in the Artemia salina assay, and xenematide acted weakly insecticidal.

Scopularides A and B, Cyclodepsipeptides from a Marine Sponge-Derived Fungus, Scopulariopsis brevicaulis

Two novel cyclodepsipeptides, scopularides A and B, were found in the fungus Scopulariopsis brevicaulis, which was isolated from the marine sponge Tethya aurantium. In addition, the known fungal metabolite paxilline was identified. The structures of the scopularides were elucidated by NMR, MS, and chemical derivatization methods as cyclo-(3-hydroxy-4-methyldecanoyl-Gly-L-Val-D-Leu-L-Ala-L-Phe) and cyclo-(3-hydroxy-4-methyloctanoyl-Gly-L-Val-D-Leu-L-Ala-L-Phe) for scopularide A and B, respectively. Antibiotic activity against Gram-negative bacteria was absent and against Gram-positive bacteria was weak, but activity against several tumor cell lines was significant at 10 microg/mL.

Evariquinone, isoemericellin, and stromemycin from a sponge derived strain of the fungus Emericella variecolor

From a strain of the fungus Emericella variecolor derived from the marine sponge Haliclona valliculata, two new natural products, evariquinone and isoemericellin, were isolated after HPLC-UV, -MS, and -NMR studies of the extract and their structures were elucidated by mass spectrometry and NMR experiments. Evariquinone showed antiproliferative activity towards KB and NCI-H460 cells at a concentration of 3.16 microg/ml. Furthermore, the fungus was found to produce the known metabolites stromemycin, shamixanthone, and 7-hydroxyemodin. Chemical degradation, NMR decoupling experiments, and spin-system simulation provided evidence for the double bonds in stromemycin to be all E-configured. ROESY experiments established the monosaccharide moiety to be glucose.

Subinhibitory concentrations of antibiotics induce phenazine production in a marine Streptomyces sp.

Subinhibitory concentrations of antibiotics were found to enhance and modulate the production of new phenazines, streptophenazines A-H, in a marine Streptomyces isolate. The streptophenazines differ in length and substitution of an alkyl chain. The pattern of metabolites formed depends on the antibiotic used. In the presence of tetracycline, streptophenazines F and G were induced and the production of streptophenazines A-D was increased. When using bacitracin, mainly streptophenazine H was produced. Streptophenazines C and H showed moderate activity against Bacillus subtilis, while streptophenazine C was also active against Staphylococcus lentus.

Matrix-bound 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in tobacco: quantification and evidence for an origin from lignin-incorporated alkaloids.

Substantial quantities of the carcinogenic tobacco-specific N-nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (1; NNK) are still found in the mainstream smoke of tobacco exhaustively extracted with water, indicating the presence of an insoluble, matrix-bound form. Soluble and matrix-bound concentrations of 1 in tobacco were determined by applying a new method using sequential aqueous extraction at room temperature and at 130 °C. On average, 77% and 53% of the total content of 1 were matrix-bound in air-cured (Burley type) and flue-cured tobaccos, respectively. Thermal release of 1 from its matrix-bound form above ca. 200 °C can account for a large fraction of its concentration in cigarette mainstream smoke. An already matrix-bound alkaloid precursor of matrix-bound 1 was identified in vascular tissue of green leaf midribs. The incubation of vascular cell-wall preparations with the lignin precursor coniferyl alcohol and isotopically labeled nicotine or pseudooxynicotine (2) led to the formation of labeled matrix-bound 1 after nitrosation, suggesting that incorporation of nicotine or its oxidized product 2 during lignin polymerization is the origin of the formation of matrix-bound 1.

Identification of CYP82E21 as a functional nicotine N-demethylase in tobacco flowers

In the tobacco plant, nicotine N-demethylase enzymes (NND) belonging to the cytochrome P450 family catalyse the conversion of nicotine to nornicotine, the precursor of the carcinogenic tobacco-specific N-nitrosamine, N-nitrosonornicotine. To date three demethylase genes, namely CYP82E4, CYP82E5 and CYP82E10, have been shown to be involved in this process, while the related CYP82E2 and CYP82E3 genes are not functional. We have identified a further gene named CYP82E21 encoding a putative nicotine N-demethylase closely related to the CYP82E genes. The CYP82E21 gene was found in all Nicotiana tabacum cultivars analysed and originates from the tobacco ancestor Nicotiana tomentosiformis. We show that, in contrast to all other previously characterized NND genes, CYP82E21 is not expressed in green or senescent leaves, but in flowers, more specifically in ovaries. The nicotine N-demethylase activity of CYP82E21 was confirmed by ectopic expression of the coding sequence in a tobacco line lacking functional CYP82E4, CYP82E5 and CYP82E10 genes, resulting in an eightfold increase of nicotine demethylation compared to the control plants. Furthermore, nornicotine formation can be reduced in ovaries by introducing a CYP82E21-specific RNAi construct. Together, our results demonstrate that the CYP82E21 gene encodes a functional ovary-specific nicotine N-demethylase.