Coordination Environment of Cu(II) Ions Bound to N-Terminal Peptide Fragments of Angiogenin Protein

Angiogenin (Ang) is a potent angiogenic factor, strongly overexpressed in patients affected by different types of cancers. The specific Ang cellular receptors have not been identified, but it is known that Ang–actin interaction induces changes both in the cell cytoskeleton and in the extracellular matrix. Most in vitro studies use the recombinant form (r-Ang) instead of the form that is normally present in vivo (“wild-type”, wt-Ang). The first residue of r-Ang is a methionine, with a free amino group, whereas wt-Ang has a glutamic acid, whose amino group spontaneously cyclizes in the pyro-glutamate form. The Ang biological activity is influenced by copper ions. To elucidate the role of such a free amino group on the protein–copper binding, we scrutinized the copper(II) complexes with the peptide fragments Ang(1–17) and AcAng(1–17), which encompass the sequence 1–17 of angiogenin (QDNSRYTHFLTQHYDAK-NH₂), with free amino and acetylated N-terminus, respectively. Potentiometric, ultraviolet-visible (UV-vis), nuclear magnetic resonance (NMR) and circular dichroism (CD) studies demonstrate that the two peptides show a different metal coordination environment. Confocal microscopy imaging of neuroblastoma cells with the actin staining supports the spectroscopic results, with the finding of different responses in the cytoskeleton organization upon the interaction, in the presence or not of copper ions, with the free amino and the acetylated N-terminus peptides.     

Dietary krill oil increases docosahexaenoic acid and reduces 2-arachidonoylglycerol but not N-acylethanolamine levels in the brain of obese Zucker rats

Evidence suggests that dietary long chain polyunsaturated fatty acids (LCPUFAs), and particularly those belonging to the n-3 family, may influence the brain fatty acid profile and, thereby, the biosynthesis of endocannabinoids in rodents. However, the doses used are usually quite high and not comparable with human intake. Recently, we have shown that relatively low doses of dietary n-3 LCPUFAs (4 weeks), in the form of either fish or krill oil, balanced for EPA and DHA content, and against a control diet with no EPA and DHA and similar contents of oleic, linoleic and α-linolenic acids, lower the concentrations of the endocannabinoids, anandamide (AEA) and 2-arachidonoylglycerol (2-AG), in the visceral adipose tissue, and of AEA in the liver and heart, of obese Zucker rats. This, in turn, is associated with lower levels of arachidonic acid in membrane phospholipids and with amelioration of some metabolic syndrome parameters. We investigated here whether in Zucker rats, under the same conditions, fish and krill oil are also able to influence LCPUFA and endocannabinoid profiles in brain. Only krill oil was able to increase significantly DHA levels in brain phospholipids, with no changes in arachidonic acid. DHA increase was associated with lower levels of 2-AG in the brain, whereas AEA and its congeners, N-palmitoylethanolamine and N-oleoylethanolamine, were unchanged. We conclude that, despite the strong impact of dietary n-3 fatty acid on endocannabinoid levels previously observed in peripheral tissues, in the brain only 2-AG is affected by dietary krill oil, suggesting that the beneficial effect of the latter on the metabolic syndrome is mostly exerted by modifying peripheral endocannabinoids. Nevertheless, possible effects of dietary krill oil in the brain through modification of 2-AG levels deserve further investigation.     

Identification of Human Adenovirus in Respiratory Samples with Luminex Respiratory Virus Panel Fast V2 Assay and Real-Time Polymerase Chain Reaction

In order to compare the last version of the Respiratory Virus Panel (RVP) Fast assay for human Adenovirus (hAdv) detection with a specific real-time polymerase chain reaction (qPCR), which is considered the gold standard for hAdv detection, nasopharyngeal samples collected from 309 children (age range, four months to eight years) with respiratory tract infection were tested using the RVP Fast v2 assay (Luminex Molecular Diagnostics, Inc., Toronto, ON, Canada) and a specific TaqMan qPCR to identify hAdv DNA. The RVP Fast v2 assay detected 30/61 (49.2%) hAdv infections that had been identified by real-time qPCR, demonstrating a significantly lower detection rate (p < 0.001). The sensitivity of the RVP Fast v2 assay in comparison to qPCR was lower in younger children (42.9% vs. 57.7%; Cohen’s kappa coefficient, 0.53); in samples with co-infections (40.0% vs. 56.7%; Cohen’s kappa coefficient, 0.52); and in samples with hAdv type C (45.9% vs. 57.1%; Cohen’s kappa coefficient, 0.60). Samples with lower viral loads were associated with a significantly lower sensitivity of the RVP Fast v2 assay (35.1% vs. 68.2%, p = 0.01; Cohen’s kappa coefficients, 0.49). The RVP Fast v2 assay has important limitations for the detection of hAdv and cannot be used to evaluate whether hAdvs are the main etiologic agent responsible for an outbreak or when epidemiological studies are performed.     

Development of a Hypersensitive Periodate‐Cleavable Amino Acid that is Methionine‐ and Disulfide‐Compatible and its Application in MHC Exchange Reagents for T Cell Characterisation

Incorporation of cleavable linkers into peptides and proteins is of particular value in the study of biological processes. Here we describe the synthesis of a cleavable linker that is hypersensitive to oxidative cleavage as the result of the periodate reactivity of a vicinal amino alcohol moiety. Two strategies directed towards the synthesis of a building block suitable for solid‐phase peptide synthesis were developed: a chemoenzymatic route, involving L ‐threonine aldolase, and an enantioselective chemical route; these led to α,γ‐diamino‐β‐hydroxybutanoic acids in diastereoisomerically mixed and enantiopure forms, respectively. Incorporation of the 1,2‐amino alcohol linker into the backbone of a peptide generated a conditional peptide that was rapidly cleaved at very low concentrations of sodium periodate. This cleavable peptide ligand was applied in the generation of MHC exchange reagents for the detection of antigen‐specific T cells in peripheral blood cells. The extremely low concentration of periodate required to trigger MHC peptide exchange allowed the co‐oxidation of methionine and disulfide residues to be avoided. Conditional MHC reagents hypersensitive to periodate can now be applied without limitations when UV irradiation is undesired or less practical.     

Niche partitioning,shape of species response,and diversity in the phytobenthos across the rocky shoreline of a large peri-Alpine lake

By colonizing a particular depth zone across the transition from aquatic to terrestrial habitats, littoral species may exhibit specific physiological and ecological adaptations, as well as characteristic responses to the gradient of conditions across the ecotone. The objectives of the study were: (i) to identify the depth zone (location in relation to average water level) where the replacement of species occurs most rapidly; (ii) to test whether and to what extent the occurrence of species, their abundance and their response to the gradient evolves over time; and (iii) to assess the shape of the species' response to the gradient, calculating the niche overlap of the dominant species. Results showed that the diversity of species peaked at a depth between 18 and 48 cm, in a zone thought to be of intermediate disturbance (the transect depth was, on average, 103 cm). The main macroalgal species (the red alga, Bangia atropurpurea and the green algae, Jaoa bullata, and Cladophora glomerata), showed a variety of response shapes to the gradient: monotonic, symmetrical and skewed, depending on the stage of seasonal growth. The efficient regulation of growth of B. atropurpurea along the fluctuating gradient was interpreted as an adaptive trait giving it an advantage over more slowly reacting species. The spatial and temporal niches of B. atropurpurea and Jaoa bullata overlapped widely only in early spring, whereas later their optimal habitats were clearly differentiated. This suggested partial niche segregation between these two species, and a potential seasonal interaction.     

Use of native species and biodegradable chelating agents in the phytoremediation of abandoned mining areas

BACKGROUND: The application of phytostabilization and assisted phytoextraction to the remediation of abandoned mining areas can be a valuable method to reclaim these areas without modifying soil and landscape characteristics. An in situ application of a continuous phytoextraction technique was carried out in the area of Campo Pisano (Sardinia, Italy), followed by a laboratory assisted phytoextraction test using the biodegradable chelating agents methylglycine diacetic acid (MGDA) and iminodissuccinic acid (IDSA). The plants used were Scrophularia canina subsp. bicolor, Cistus salviifolius and Teucrium flavum subsp. glaucum. RESULTS: The plant that accumulated more Pb was T. glaucum (353 mg kg^?1) while C. salviifolius demonstrated better ability to accumulate Zn (1560 mg kg^?1). S. bicolor showed a better tolerance to metals but accumulated 119 mg kg^?1 of Pb. Accumulation of metals immediately after chelant application was up to 300 mg kg^?1 of Pb and 3000 mg kg^?1 of Zn which did not further increase during the assisted phytoextraction experiment. CONCLUSION: The plant that demonstrated to be most suitable for phytoremediation application was S. bicolor due to its higher biomass production and tolerance to metals. The low cation exchange capacity and the high concentration of Ca and Mg in soil determined a low chelant effectiveness. Copyright © 2009 Society of Chemical Industry     

Identification of Aminoimidazole and Aminothiazole Derivatives as Src Family Kinase Inhibitors

Src family kinases (SFKs) are a family of non‐receptor tyrosine kinases (TKs) implicated in the regulation of many cellular processes. The aberrant activity of these TKs has been associated with the growth and progression of cancer. In particular, c‐Src is overexpressed or hyperactivated in a variety of solid tumors and is most likely a strong promoting factor for the development of metastasis. Herein, the synthesis of new 4‐aminoimidazole and 2‐aminothiazole derivatives and their in vitro biological evaluation are described for their potential use as SFK inhibitors. Initially, 2‐aminothiazole analogues of dasatinib and 4‐aminoimidazole derivatives were synthesized and tested against the SFKs Src, Fyn, Lyn, and Yes. Five hits were identified as the most promising compounds, with K_i values in the range of 90–480 nm . A combination of molecular docking, homology modeling, and molecular dynamics were then used to investigate the possible binding mode of such compounds within the ATP binding site of the SFKs. Finally, the antiproliferative activities of the best candidates were evaluated against SH‐SY5Y and K562 cell lines. Compound 3 b [2‐(4‐{2‐methyl‐6‐[(5‐phenylthiazol‐2‐yl)amino]pyrimidin‐4‐yl}piperazin‐1‐yl)ethanol] was found to be the most active inhibitor.     

Yeast vectors for the integration/expression of any sequence at the TYR1 locus

We have constructed new yeast vectors for targeted integration and conditional expression of any sequence at the Saccharomyces cerevisiae TYR1 locus which becomes disrupted. We show that vector integration is not neutral, causing prototrophy for tyrosine and auxotrophy for the vector's selectable marker (uracil or leucine, depending on the vector used). This feature allows a double screening of transformed yeast cells, improving the identification of colonies with the desired chromosomal structure. The GAL10 gene promoter has been added to drive conditional expression of cloned sequences. Using these vectors, chromosomal structure verification of recombinant clones is no longer necessary, since the noise of non-homologous recombination, as well as spontaneous reversion of the selected phenotype, can easily be identified. The ability of the vector to conditionally control gene expression has been confirmed using the gene for the green fluorescent protein (GFP) as a reporter.     

Optimal input sets for steering quantized systems

Limited capacity of communication channels has strongly pushed the analysis of control systems subject to a quantized input set. Quantized control system of type x ⁺ = x + u, where the u takes values in a set of 2m + 1 integer numbers, symmetric with respect to 0 arise in some fundamental situations, e.g., flat, nilpotent, and linear systems with quantized feedback. In this paper we consider this special type of systems and analyze the reachable set after K steps. We find explicit expressions, for each K and for each m, of m input values such that the reachable set after K steps is as large as possible.     

Influence of thermal ageing on the stability of polymer bulk heterojunction solar cells

A new approach is presented in order to improve the thermal stability of polymer: [6-6]-phenyl C₆₁ butyric acid methyl ester (PCBM) bulk heterojunction solar cells. The central idea in this approach is the use of a polymer with high glass transition temperature (T_g), well above the normal operating temperatures of the devices. In this paper, a PPV-derivative with a T_g of 150 °C was used as an electron donor and the thermal stability of the obtained solar cells was compared with solar cells based on the reference material poly[2-methoxy-5-(3′,7′-dimethyloctyloxy)-1,4-phenylene vinylene] (MDMO-PPV) with a T_g of 45 °C. The use of the material with higher glass transition temperature resulted in a significant improvement of the thermal stability of the photovoltaic parameters. Furthermore, a systematic transmission electron microscope (TEM) study demonstrates that the better thermal stability of performance coincides with a more stable active layer morphology. Both improvements are attributed to the reduced free movement of the electron donor material (PCBM) within the active layer of the solar cell.