Novel electrochemically-mediated peptide dethiylation in processes relevant to native chemical ligation

Here we explore electrochemical dethiylation in processes relevant to Native Chemical Ligation (NCL). NCL's reliance on the redox active amino acid cysteine and β-mercaptamine derivatives suggests a potential role for electrochemistry. We show that the application of a 1 V potential to platinum electrodes in 0.15 M TCEP solution is sufficient to convert Cys to Ala in cyclic peptides, and to cleave the 2-mercapto-2-phenethyl class of acyl transfer auxiliary

Investigation of acyl transfer auxiliary-assisted glycoconjugation for glycoprotein semi-synthesis

Homogeneous glycoprotein syntheses have become possible in the last decade due to advances in chemical ligation strategies, particularly Native Chemical Ligation (NCL). For native glycoproteins this still requires laborious and technically challenging syntheses of glycopeptide components, combined with multi-segment ligation reactions. Here we explore new reactions between sugar-linked acyl transfer auxiliaries and peptide thioesters. We show that native glycoproteins are difficult to produce using this approach but various related analogues are accessible. The results show that site-specific neoglycoconjugation is a viable route to simply glycosylated proteins, which may be extended using well-documented enzymatic processes

Global Continua of Positive Equilibria for some Quasilinear Parabolic Equation with a Nonlocal Initial Condition

This paper is concerned with a quaslinear parabolic equation including a nonlinear nonlocal initial condition. The problem arises as equilibrium equation in population dynamics with nonlinear diffusion. We make use of global bifurcation theory to prove existence of an unbounded continuum of positive solutions

Genome-Wide Studies of Histone Demethylation Catalysed by the Fission Yeast Homologues of Mammalian LSD1

In order to gain a more global view of the activity of histone demethylases, we report here genome-wide studies of the fission yeast SWIRM and polyamine oxidase (PAO) domain homologues of mammalian LSD1. Consistent with previous work we find that the two S. pombe proteins, which we name Swm1 and Swm2 (after SWIRM1 and SWIRM2), associate together in a complex. However, we find that this complex specifically demethylates lysine 9 in histone H3 (H3K9) and both up- and down-regulates expression of different groups of genes. Using chromatin-immunoprecipitation, to isolate fragments of chromatin containing either H3K4me2 or H3K9me2, and DNA microarray analysis (ChIP-chip), we have studied genome-wide changes in patterns of histone methylation, and their correlation with gene expression, upon deletion of the swm1+ gene. Using hyper-geometric probability comparisons we uncover genetic links between lysine-specific demethylases, the histone deacetylase Clr6, and the chromatin remodeller Hrp1. The data presented here demonstrate that in fission yeast the SWIRM/PAO domain proteins Swm1 and Swm2 are associated in complexes that can remove methyl groups from lysine 9 methylated histone H3. In vitro, we show that bacterially expressed Swm1 also possesses lysine 9 demethylase activity. In vivo, loss of Swm1 increases the global levels of both H3K9me2 and H3K4me2. A significant accumulation of H3K4me2 is observed at genes that are up-regulated in a swm1 deletion strain. In addition, H3K9me2 accumulates at some genes known to be direct Swm1/2 targets that are down-regulated in the swm1¿ strain. The in vivo data indicate that Swm1 acts in concert with the HDAC Clr6 and the chromatin remodeller Hrp1 to repress gene expression. In addition, our in vitro analyses suggest that the H3K9 demethylase activity requires an unidentified post-translational modification to allow it to act. Thus, our results highlight complex interactions between histone demethylase, deacetylase and chromatin remodelling activities in the regulation of gene expression

Characterization of Shewanella oneidensis MtrC: a cell-surface decaheme cytochrome involved in respiratory electron transport to extracellular electron acceptors

MtrC is a decaheme c-type cytochrome associated with the outer cell membrane of Fe(III)-respiring species of the Shewanella genus. It is proposed to play a role in anaerobic respiration by mediating electron transfer to extracellular mineral oxides that can serve as terminal electron acceptors. The present work presents the first spectropotentiometric and voltammetric characterization of MtrC, using protein purified from Shewanella oneidensis MR-1. Potentiometric titrations, monitored by UV–vis absorption and electron paramagnetic resonance (EPR) spectroscopy, reveal that the hemes within MtrC titrate over a broad potential range spanning between approximately +100 and approximately -500 mV (vs. the standard hydrogen electrode). Across this potential window the UV–vis absorption spectra are characteristic of low-spin c-type hemes and the EPR spectra reveal broad, complex features that suggest the presence of magnetically spin-coupled low-spin c-hemes. Non-catalytic protein film voltammetry of MtrC demonstrates reversible electrochemistry over a potential window similar to that disclosed spectroscopically. The voltammetry also allows definition of kinetic properties of MtrC in direct electron exchange with a solid electrode surface and during reduction of a model Fe(III) substrate. Taken together, the data provide quantitative information on the potential domain in which MtrC can operate

Vulnerability of drug‐resistant EML4‐ALK rearranged lung cancer to transcriptional inhibition

A subset of lung adenocarcinomas is driven by the EML4‐ALK translocation. Even though ALK inhibitors in the clinic lead to excellent initial responses, acquired resistance to these inhibitors due to on‐target mutations or parallel pathway alterations is a major clinical challenge. Exploring these mechanisms of resistance, we found that EML4‐ALK cells parental or resistant to crizotinib, ceritinib or alectinib are remarkably sensitive to inhibition of CDK7/12 with THZ1 and CDK9 with alvocidib or dinaciclib. These compounds robustly induce apoptosis through transcriptional inhibition and downregulation of anti‐apoptotic genes. Importantly, alvocidib reduced tumour progression in xenograft mouse models. In summary, our study takes advantage of the transcriptional addiction hypothesis to propose a new treatment strategy for a subset of patients with acquired resistance to first‐, second‐ and third‐generation ALK inhibitors

Development of a LAMP assay for detection of Leishmania infantum infection in dogs using conjunctival swab samples

Background: Leishmania infantum infections in dogs play a crucial role in the transmission of pathogens causing visceral leishmaniasis to humans in the Gansu province, northwest China. To be able to control zoonotic transmission of the parasite to humans, a non-invasive loop-mediated isothermal amplification (LAMP) assay to specifically detect L. infantum infections in dogs was developed. Methods: The primers used in the LAMP assay were designed to target kinetoplast DNA minicircle sequences of the L. infantum isolate MCAN/CN/90/SC and tested using DNA isolated from promastigotes of different Leishmania species. The LAMP assay was evaluated with conjunctional swab samples obtained from 111 and 33 dogs living in an endemic and a non-endemic region of zoonotic visceral leishmaniasis in the Gansu province, respectively. The LAMP assay was also compared with conventional PCR, ELISA and microscopy using conjunctional swab, serum and bone marrow samples from the dogs, respectively. Results: The LAMP assay detected 1 fg of L. infantum DNA purified from cultured promastigotes which was 10-fold more sensitive than a conventional PCR test using Leishmania genus-specific primers. No cross reaction was observed with DNA isolated from promastigotes of L. donovani, L. major, L. tropica, and L. braziliensis, and the L. infantum reference strain MHOM/TN/80/IPT1. The L. infantum-positive rates obtained for field-collected samples were 61.3%, 58.6%, 40.5% and 10.8% by LAMP, PCR, ELISA and microscopy, respectively. As only one out of the 33 samples from control dogs from the non-endemic region of zoonotic visceral leishmaniasis was positive by the LAMP assay and the PCR test, the observed true negative rate (specificity) was 97% for both methods. Conclusion: This study has shown that the non-invasive, conjunctional swab-based LAMP assay developed was more sensitive in the detection of leishmaniasis in dogs than PCR, ELISA and microscopy. The findings indicate that the LAMP assay is a sensitive and specific method for the field surveillance of domestic dogs, particularly of asymptomatic canines, in ZVL-endemic areas in western China

Onion routing in deterministic delay tolerant networks

Aquest volum de Lecture notes in computer science, amb el títol Foundations and Practice of Security, recull les actes al 8th International Symposium on Foundations & Practice of Security que va tenir lloc a Clermont-Ferrand (France), del 16 al 28 d'octubre de 2015Deterministic DTNs are networks where the behavior is known in advance or where a repetitive action occurs over time like in public transportation networks. This work proposes the application of an onion routing approach to deterministic DTNs to achieve anonymous communications. We show how the prior stage of path selection in onion routing can be achieved using the information provided by deterministic networks

Lysine-Specific Demethylase 1 (LSD1) Is Required for the Transcriptional Repression of the Telomerase Reverse Transcriptase (hTERT) Gene

BACKGROUND: Lysine-specific demethylase 1 (LSD1), catalysing demethylation of mono- and di-methylated histone H3-K4 or K9, exhibits diverse transcriptional activities by mediating chromatin reconfiguration. The telomerase reverse transcriptase (hTERT) gene, encoding an essential component for telomerase activity that is involved in cellular immortalization and transformation, is silent in most normal human cells while activated in up to 90% of human cancers. It remains to be defined how exactly the transcriptional activation of the hTERT gene occurs during the oncogenic process. METHODOLOGY/PRINCIPAL FINDINGS: In the present study, we determined the effect of LSD1 on hTERT transcription. In normal human fibroblasts with a tight hTERT repression, a pharmacological inhibition of LSD1 led to a weak hTERT expression, and a robust induction of hTERT mRNA was observed when LSD1 and histone deacetylases (HDACs) were both inhibited. Small interference RNA-mediated depletion of both LSD1 and CoREST, a co-repressor in HDAC-containing complexes, synergistically activated hTERT transcription. In cancer cells, inhibition of LSD1 activity or knocking-down of its expression led to significant increases in levels of hTERT mRNA and telomerase activity. Chromatin immunoprecipitation assay showed that LSD1 occupied the hTERT proximal promoter, and its depletion resulted in elevated di-methylation of histone H3-K4 accompanied by increased H3 acetylation locally in cancer cells. Moreover, during the differentiation of leukemic HL60 cells, the decreased hTERT expression was accompanied by the LSD1 recruitment to the hTERT promoter. CONCLUSIONS/SIGNIFICANCE: LSD1 represses hTERT transcription via demethylating H3-K4 in normal and cancerous cells, and together with HDACs, participates in the establishment of a stable repression state of the hTERT gene in normal or differentiated malignant cells. The findings contribute to better understandings of hTERT/telomerase regulation, which may be implicated in the development of therapeutic strategies for telomerase dysregulation-associated human diseases including cancers

Recent changes of water discharge and sediment load in the Yellow River basin, China

The Yellow River basin contributes approximately 6% of the sediment load from all river systems globally, and the annual runoff directly supports 12% of the Chinese population. As a result, describing and understanding recent variations of water discharge and sediment load under global change scenarios are of considerable importance. The present study considers the annual hydrologic series of the water discharge and sediment load of the Yellow River basin obtained from 15 gauging stations (10 mainstream, 5 tributaries). The Mann-Kendall test method was adopted to detect both gradual and abrupt change of hydrological series since the 1950s. With the exception of the area draining to the Upper Tangnaihai station, results indicate that both water discharge and sediment load have decreased significantly (p<0.05). The declining trend is greater with distance downstream, and drainage area has a significant positive effect on the rate of decline. It is suggested that the abrupt change of the water discharge from the late 1980s to the early 1990s arose from human extraction, and that the abrupt change in sediment load was linked to disturbance from reservoir construction.Geography, PhysicalGeosciences, MultidisciplinarySCI(E)43ARTICLE4541-5613