Status of research in stock enhancement and sea ranching

Reviews in Fish Biology and Fisheries -     

Validation of Drug-Like Inhibitors against Mycobacterium Tuberculosis L-Aspartate α-Decarboxylase Using Nuclear Magnetic Resonance (1H NMR)

The catalytic activity of L-aspartate α-decarboxylase (ADC) is essential for the growth of several micro-organisms, including Mycobacterium tuberculosis (Mtb), and has triggered efforts for the development of pharmaceutically active compounds against tuberculosis. The present study is a continuation of our recent chemoinformatics-based design approach for identifying potential drug-like inhibitors against MtbADC. We report an NMR-based protocol that allows label-free and direct monitoring of enzymatic conversion, which we have combined with a systematic testing of reported and newly identified potential inhibitors against MtbADC. Quantification of enzymatic conversion in the absence and presence of inhibitors allowed for a relative measure of the inhibitory effect (k _rel). Among the newly identified compounds, D-tartrate, L-tartrate, and 2,4-dihydroxypyrimidine-5-carboxylate were found to inhibit the enzyme with k _rel values of 0.36, 0.38, and 0.54, respectively. In addition to the identification of potential building blocks for the development of therapeutic agents, the current study highlights the importance of electrostatic interactions governing enzyme-inhibitor binding.     

Membrane lysis by gramicidin S visualized in red blood cells and giant vesicles

The cationic amphiphilic antimicrobial peptide gramicidin S (GS) is an effective antibiotic. Its applicability is however restricted to topical infections due to its hemolytic activity. In this study, the process of GS induced hemolysis was investigated in detail for the first time. The morphological changes of red blood cells (RBCs) inflicted by GS were visualized and explained in terms of a physical model. The observed fast rupture events were further investigated with giant unilamellar vesicles (GUVs) as model systems for RBCs. Measurements of membrane fluctuations in GUVs revealed that the membrane surface tension was increased after incubation with GS. These findings are in agreement with the hypothesis that amphiphilic peptides induce membrane rupture by an increase in membrane tension.     

Synthesis, mesomorphism and luminescence properties of palladium(II) and platinum(II) complexes with dimeric Schiff base liquid crystals

A series of ortho-metallated Pd and Pt complexes containing dimeric liquid crystals Schiff base as cyclometallated ligands and N-benzoyl thiourea derivatives as co-ligands were prepared and investigated for their liquid crystalline properties. Their structures were assigned based on elemental analysis, IR and ¹H NMR spectroscopy while the mesogenic properties were investigated by DSC and polarising optical microscopy. The complexes show either monotropic or enantiotropic transitions with nematic and smectic A phases being displayed, with the mesomorphic behaviour strongly related to the type of N-benzoyl thiourea as well as the metal center used. The structure of a palladium(II) complex has been solved by X-ray diffraction.The platinum(II) complexes show photoluminescence properties both in solution and in solid state at room temperature, with the emission band centered around 600 nm. These are the first examples of metallomesogens based on Schiff base cyclometallated ligands that display luminescence properties.     

Ultrasensitive in situ visualization of active glucocerebrosidase molecules

Deficiency of glucocerebrosidase (GBA) underlies Gaucher disease, a common lysosomal storage disorder. Carriership for Gaucher disease has recently been identified as major risk for parkinsonism. Presently, no method exists to visualize active GBA molecules in situ. We here report the design, synthesis and application of two fluorescent activity-based probes allowing highly specific labeling of active GBA molecules in vitro and in cultured cells and mice in vivo. Detection of in vitro labeled recombinant GBA on slab gels after electrophoresis is in the low attomolar range. Using cell or tissue lysates, we obtained exclusive labeling of GBA molecules. We present evidence from fluorescence-activated cell sorting analysis, fluorescence microscopy and pulse-chase experiments of highly efficient labeling of GBA molecules in intact cells as well as tissues of mice. In addition, we illustrate the use of the fluorescent probes to study inhibitors and tentative chaperones in living cells.     

Macrophage-specific up-regulation of apolipoprotein E gene expression by STAT1 is achieved via long range genomic interactions

In atherogenesis, macrophage-derived apolipoprotein E (apoE) has an athero-protective role by a mechanism that is not fully understood. We investigated the regulatory mechanisms involved in the modulation of apoE expression in macrophages. The experiments showed that the promoters of all genes of the apoE/apoCI/apoCIV/apoCII gene cluster are enhanced by multienhancer 2 (ME.2), a regulatory region that is located 15.9 kb downstream of the apoE gene. ME.2 interacts with the apoE promoter in a macrophage-specific manner. Transient transfections in RAW 264.7 macrophages showed that the activity of ME.2 was strongly decreased by deletion of either 87 bp from the 5' end or 131 bp from the 3' end. We determined that the minimal fragment of this promoter that can be activated by ME.2 is the proximal -100/+73 region. The analysis of the deletion mutants of ME.2 revealed the importance of the 5' end of ME.2 in apoE promoter transactivation. Chromatin conformational capture assays demonstrated that both ME.2 and ME.1 physically interacted with the apoE promoter in macrophages. Our data showed that phorbol 12-myristate 13-acetate-induced differentiation of macrophages is accompanied by a robust induction of apoE and STAT1 expression. In macrophages (but not in hepatocytes), STAT1 up-regulated apoE gene expression via ME.2. The STAT1 binding site was located in the 174-182 region of ME.2. In conclusion, the specificity of the interactions between the two multienhancers (ME.1 and ME.2) and the apoE promoter indicates that these distal regulatory elements play an important role in the modulation of apoE gene expression in a cell-specific manner.     

Neural mechanisms of context-dependent processing of CO2 avoidance behavior in fruit flies

The fruit fly, Drosophila melanogaster, innately avoids even low levels of CO₂. CO₂ is part of the so-called Drosophila stress odor produced by stressed flies, but also a byproduct of fermenting fruit, a main food source, making the strong avoidance behavior somewhat surprising. Therefore, we addressed whether feeding states might influence the fly’s behavior and processing of CO₂. In a recent report, we showed that this innate behavior is differentially processed and modified according to the feeding state of the fly. Interestingly, we found that hungry flies require the function of the mushroom body, a higher brain center required for olfactory learning and memory, but thought to be dispensable for innate olfactory behaviors. In addition, we anatomically and functionally characterized a novel bilateral projection neuron connecting the CO₂ sensory input to the mushroom body. This neuron was essential for processing of CO₂ in the starved fly but not in the fed fly. In this Extra View article, we provide evidence for the potential involvement of the neuromodulator dopamine in state-dependent CO₂ avoidance behavior. Taken together, our work demonstrates that CO₂ avoidance behavior is mediated by alternative neural pathways in a context-dependent manner. Furthermore, it shows that the mushroom body is not only involved in processing of learned olfactory behavior, as previously suggested, but also in context-dependent innate olfaction.     

Identification of glucose kinase‐dependent and ‐independent pathways for carbon control of primary metabolism,development and antibiotic production in Streptomyces coelicolor by quantitative proteomics

Members of the soil‐dwelling prokaryotic genus Streptomyces are indispensable for the recycling of complex polysaccharides, and produce a wide range of natural products. Nutrient availability is a major determinant for the switch to development and antibiotic production in streptomycetes. Carbon catabolite repression (CCR), a main signalling pathway underlying this phenomenon, was so far considered fully dependent on the glycolytic enzyme glucose kinase (Glk). Here we provide evidence of a novel Glk‐independent pathway in Streptomyces coelicolor, using advanced proteomics that allowed the comparison of the expression of some 2000 proteins, including virtually all enzymes for central metabolism. While CCR and inducer exclusion of enzymes for primary and secondary metabolism and precursor supply for natural products is mostly mediated via Glk, enzymes for the urea cycle, as well as for biosynthesis of the γ‐butyrolactone Scb1 and the responsive cryptic polyketide Cpk are subject to Glk‐independent CCR. Deletion of glkA led to strong downregulation of biosynthetic proteins for prodigionins and calcium‐dependent antibiotic (CDA) in mannitol‐grown cultures. Repression of bldB, bldN, and its target bldM may explain the poor development of S. coelicolor on solid‐grown cultures containing glucose. A new model for carbon catabolite repression in streptomycetes is presented.