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American foulbrood ( AFB) caused by the bee pathogenic bacterium P aenibacillus larvae is the most devastating bacterial disease of honey bees worldwide. From AFB-dead larvae, pure cultures of P . larvae can normally be cultivated indicating that P . larvae is able to defend its niche against all other bacteria present. Recently, comparative genome analysis within the species P . larvae suggested the presence of gene clusters coding for multi-enzyme complexes, such as non-ribosomal peptide synthetases ( NRPSs). The products of these enzyme complexes are known to have a wide range of biological activities including antibacterial activities. We here present our results on antibacterial activity exhibited by vegetative P . larvae and the identification and analysis of a novel antibacterially active P . larvae tripeptide (called sevadicin; Sev) produced by a NRPS encoded by a gene cluster found in the genome of P . larvae. Identification of Sev was ultimately achieved by comparing the secretome of wild-type P . larvae with knockout mutants of P . larvae lacking production of Sev. Subsequent mass spectrometric studies, enantiomer analytics and chemical synthesis revealed the sequence and configuration of the tripeptide, D-Phe- D- ALa-Trp, which was shown to have antibacterial activity. The relevance of our findings is discussed in respect to host-pathogen interactions. [ABSTRACT FROM AUTHOR]

Nonribosomal peptides (NRPs), a large family of natural products, possess numerous pharmaceutically significant bioactivities. However, many native microbial producers of NRPs are not cultivable or have low production yields making mass production infeasible. The recombinant production of natural products in a surrogate host has emerged as a strategy to overcome these limitations. De novo recombinant production of the NRP antibiotic valinomycin in an engineered Escherichia coli host strain was established with the necessary biosynthetic pathway constituents from Streptomyces tsusimaensis. In the present study, the initially modest valinomycin yields could be significantly increased from 0.3 up to 2.4 mg L by switching from a batch to an enzyme-based fed-batch mode in shake flasks. A subsequent design of experiment-driven optimization of parallel fed-batch cultivations in 24-well plates with online monitoring of dissolved oxygen and pH led to valinomycin yields up to 6.4 mg L. Finally, repeated glucose polymer feeding to enzyme-based high cell density cultivations in shake flasks resulted in cell densities of OD >50 and a valinomycin titer of appr. 10 mg L. This represents a 33-fold improvement compared to the initial batch cultivations and is the highest concentration of a nonribosomal peptide which has been produced in E. coli without feeding of specific precursors so far to our knowledge. Also, such a small-scale optimization under fed-batch conditions may be generally applicable for the development and scale-up of natural product production processes in E. coli. [ABSTRACT FROM AUTHOR]

The E. coli siderophore enterobactin, one of the strongest FeIII chelators known to date, is also capable of binding SiIV under physiological conditions. We report on the synthesis and structural characterization of the tris(catecholate) SiIV-enterobactin complex and its GeIV and TiIV analogues. Comparative structural analysis, supported by quantum-chemical calculations, reveals the correlation between the ionic radius and the structural changes in enterobactin upon complexation. [ABSTRACT FROM AUTHOR]

Over the past decade, drug discovery programs have started to address the optimization of key ADME properties already at an early stage of the process. Hence, analytical chemists have been confronted with tremendously rising sample numbers and have had to develop methodologies accelerating quantitative liquid chromatography/tandem mass spectrometry (LC/MS/ MS). This article focuses on the application of a generic and fully automated LC/MS/MS, named Rapid and Integrated Analysis System (RIAS), as a high-throughput platform for the rapid quantification of drug-like compounds in various in vitro ADME assays. Previous efforts were dedicated to the setup and feasibility study of a workflow-integrated platform combining a modified high-throughput liquid handling LC/MS/MS system controlled by a customized software interface and a customized data-processing and reporting tool. Herein the authors present an extension of this previously developed basic application to a broad set of ADME screening campaigns, covering CYP inhibition, Caco-2, and PAMPA assays. The platform is capable of switching automatically between various ADME assays, performs MS compound optimization if required, and provides a speed of 8 s from sample to sample, independently of the type of ADME assay. Quantification and peak review are adopted to the high-throughput environment and tested against a standard HPLC-ESI technology. (Journal of Biomolecular Screening 2011;16:370-377) [ABSTRACT FROM PUBLISHER]

Abstract: The glycopeptide antibiotics vancomycin and teicoplanin are used in the hospital as drugs of last resort to combat resistant Gram‐positive pathogens, in particular methicillin‐resistant Staphylococcus aureus. All glycopeptides consist of a heptapeptide backbone in which the aromatic residues are connected to form a rigid cup‐shaped structure required to stably interact with the d‐Ala‐d‐Ala terminus of bacterial cell wall precursors. Structural diversity is generated by variations in the composition of the backbone, preferably at amino acid positions 1 and 3, and by different glycosylation, methylation, and chlorination patterns. The identification of several glycopeptide biosynthesis gene clusters, the development of genetic techniques to manipulate at least some of the producing actinomycetes, and subsequent molecular analysis enabled the elucidation of their biosynthetic pathways. This led to biochemical methods being combined with molecular genetic techniques and analytical chemistry. Knowledge of the biosynthesis made it possible to apply different approaches for the generation of novel glycopeptide derivatives by mutasynthesis, precursor‐directed biosynthesis, and genetic engineering. [Copyright &y& Elsevier]

In addition to the isolation and complete characterisation of a new lignan, hyoscyamal, three other compounds, balanophonin, pongamoside C and pongamoside D have been isolated from the seeds of Hyoscyamus niger. The structures of the compounds were settled on the basis of spectroscopic analysis. This is the first report on the isolation of these compounds from a solanaceous plant. [ABSTRACT FROM AUTHOR]

The cAMP-dependent aquaporin-2 (AQP2) redistribution from intracellular vesicles into the plasma membrane of renal collecting duct principal cells induces water reabsorption and fine-tunes body water homeostasis. However, the mechanisms controlling the localization of AQP2 are not understood in detail. Using immortalized mouse medullary collecting duct (MCD4) and primary rat inner medullary collecting duct (IMCD) cells as model systems, we here discovered a key regulatory role of Aurora kinase A (AURKA) in the control of AQP2. The AURKA-selective inhibitor Aurora-A inhibitor I and novel derivatives as well as a structurally different inhibitor, Alisertib, prevented the cAMP-induced redistribution of AQP2. Aurora-A inhibitor I led to a depolymerization of actin stress fibers, which serve as tracks for the translocation of AQP2-bearing vesicles to the plasma membrane. The phosphorylation of cofilin-1 (CFL1) inactivates the actin-depolymerizing function of CFL1. Aurora-A inhibitor I decreased the CFL1 phosphorylation, accounting for the removal of the actin stress fibers and the inhibition of the redistribution of AQP2. Surprisingly, Alisertib caused an increase in actin stress fibers and did not affect CFL1 phosphorylation, indicating that AURKA exerts its control over AQP2 through different mechanisms. An involvement of AURKA and CFL1 in the control of the localization of AQP2 was hitherto unknown. [ABSTRACT FROM AUTHOR]

Bacillus amyloliquefaciens FZB42 is a Gram-positive, plant-associated bacterium, which stimulates plant growth and produces secondary metabolites that suppress soil-borne plant pathogens. Its 3,918-kb genome, containing an estimated 3,693 protein-coding sequences, lacks extended phage insertions, which occur ubiquitously in the closely related Bacillus subtilis 168 genome. The B. amyloliquefaciens FZB42 genome reveals an unexpected potential to produce secondary metabolites, including the polyketides bacillaene and difficidin. More than 8.5% of the genome is devoted to synthesizing antibiotics and siderophores by pathways not involving ribosomes. Besides five gene clusters, known from B. subtilis to mediate nonribosomal synthesis of secondary metabolites, we identified four giant gene clusters absent in B. subtilis 168. The pks2 gene cluster encodes the components to synthesize the macrolactin core skeleton. [ABSTRACT FROM AUTHOR]

A wide variety of enzymatic pathways that produce specialized metabolites in bacteria, fungi and plants are known to be encoded in biosynthetic gene clusters. Information about these clusters, pathways and metabolites is currently dispersed throughout the literature, making it difficult to exploit. To facilitate consistent and systematic deposition and retrieval of data on biosynthetic gene clusters, we propose the Minimum Information about a Biosynthetic Gene cluster (MIBiG) data standard., Chemistry and Chemical Biology

The article discusses a study regarding the development of antibiotic teixobactin.. Topics discussed include the blocking of biosynthetic precursor Lipid II by teixobactin, the bacterial cell wall biosynthesis, and the activity-based isolation of Gram-negative proteobacterium Eleftheria terrae that leads to the discovery of teixobactin.

Albicidin is a potent DNA gyrase inhibitor produced by the sugarcane pathogenic bacterium Xanthomonas albilineans. Here we report the elucidation of the hitherto unknown structure of albicidin, revealing a unique polyaromatic oligopeptide mainly composed of p-aminobenzoic acids. In vitro studies provide further insights into the biosynthetic machinery of albicidin. These findings will enable structural investigations on the inhibition mechanism of albicidin and its assessment as a highly effective antibacterial drug. [ABSTRACT FROM AUTHOR]

Assassin bug venoms are potent and exert diverse biological functions, making them potential biomedical goldmines. Besides feeding functions on arthropods, assassin bugs also use their venom for defense purposes causing localized and systemic reactions in vertebrates. However, assassin bug venoms remain poorly characterized. We collected the venom from the assassin bug Rhynocoris iracundus and investigated its composition and bioactivity in vitro and in vivo. It caused lysis of murine neuroblastoma, hepatoma cells, and healthy murine myoblasts. We demonstrated, for the first time, that assassin bug venom induces neurolysis and suggest that it counteracts paralysis locally via the destruction of neural networks, contributing to tissue digestion. Furthermore, the venom caused paralysis and melanization of Galleria mellonella larvae and pupae, whilst also possessing specific antibacterial activity against Escherichia coli, but not Listeria grayi and Pseudomonas aeruginosa. A combinatorial proteo-transcriptomic approach was performed to identify potential toxins responsible for the observed effects. We identified neurotoxic Ptu1, an inhibitory cystin knot (ICK) toxin homologous to ω-conotoxins from cone snails, cytolytic redulysins homologous to trialysins from hematophagous kissing bugs, and pore-forming hemolysins. Additionally, chitinases and kininogens were found and may be responsible for insecticidal and cytolytic activities. We demonstrate the multifunctionality and complexity of assassin bug venom, which renders its molecular components interesting for potential biomedical applications. [ABSTRACT FROM AUTHOR]