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Generation and reactivity of the fragment ion [B12I8 S(CN)]− in the gas phase and on surfaces
No full textGaseous fragment ions generated in mass spectrometers may be employed as “building blocks” for the synthesis of novel molecules on surfaces using ion soft-landing. A fundamental understanding of the reactivity of the fragment ions is required to control bond formation of deposited fragments in surface layers. The fragment ion [B12X11]− (X = halogen) is formed by collision-induced dissociation (CID) from the precursor [B12X12]2− dianion. [B12X11]− is highly reactive and ion soft-landing experiments have shown that this ion binds to the alkyl chains of organic molecules on surfaces. In this work we investigate whether specific modifications of the precursor ion affect the chemical properties of the fragment ions to such an extent that attachment to functional groups of organic molecules on surfaces occurs and binding of alkyl chains is prevented. Therefore, a halogen substituent was replaced by a thiocyanate substituent. CID of the precursor [B12I11(SCN)]2− ion preferentially yields the fragment ion [B12I8S(CN)]−, which shows significantly altered reactivity compared to the fragment ions of [B12I12]2−. [B12I8S(CN)]− has a previously unknown structural element, wherein a sulfur atom bridges three boron atoms. Gas-phase reactions with different neutral reactants (cyclohexane, dimethyl sulfide, and dimethyl amine) accompanied by theoretical studies indicate that [B12I8S(CN)]− binds with higher selectivity to functional groups of organic molecules than fragment ions of [B12I12]2− (e.g., [B12I11]− and [B12I9]−). These findings were further confirmed by ion soft-landing experiments, which showed that [B12I8S(CN)]− ions attacked ester groups of adipates and phthalates, whereas [B12I11]− ions only bound to alkyl chains of the same reagents
Rapid Identification of Bacterial Composition in Wastewater by Combining Reverse Purification Nucleic Acid Extraction and Nanopore Sequencing
No full textWastewater systems represent a threat for the transmission of pathogens in the case of improper treatment. Systematic surveillance based on wastewater is important as an early warning system, e.g., COVID-19. However, current procedures for microorganism identification are often not applicable at the point-of-need. In this study, a rapid and portable DNA extraction protocol was developed and verified to identify the microbial wastewater composition by nanopore sequencing. The study compared different pretreatment and buffer options combined with the reverse purification principle. Extraction efficacy was assessed using Staphylococcus aureus and Escherichia coli spiked wastewater and crude samples from different collection points at the treatment plant. Results were compared to a spin column-based reference method and a real-time polymerase chain reaction. Lysis buffer combined with bead beating showed the best extraction efficacies using an Oxford nanopore. It demonstrated an adequate sensitivity of down to 103 copies. Up to eight different bacterial species could be identified at each collection point by nanopore sequencing. The developed protocol achieved a 3-fold increase of average length, an increase of quality score, and a 15-fold decrease of total reads. The evaluated molecular detection methods are a useful tool for the analysis of bacteria within less than 3.5 h and could contribute to public health monitoring of wastewater
Revascularization Strategy in Myocardial Infarction with Multivessel Disease
No full textThe proportion of patients with multivessel coronary artery disease in individuals experiencing acute coronary syndrome (ACS) varies based on age and ACS subtype. In patients with ST-segment elevation myocardial infarction (STEMI) without cardiogenic shock, the prognostic benefit of complete revascularization has been demonstrated by several randomized trials and meta-analyses, leading to a strong guideline recommendation. However, similar data are lacking for ACS without ST-segment elevation (NSTE-ACS). Non-randomized data suggesting a benefit from complete revascularization in non-ST-segment elevation myocardial infarction (NSTEMI) are prone to selection bias and should be interpreted with caution. A series of large randomized controlled trials have been initiated recently to address these open questions
Similar Binding Modes of cGMP Analogues Limit Selectivity in Modulating Retinal CNG Channels via the Cyclic Nucleotide-Binding Domain
No full textIn treating retinitis pigmentosa, a genetic disorder causing progressive vision loss, selective inhibition of rod cyclic nucleotide-gated (CNG) channels holds promise. Blocking the increased Ca2+-influx in rod photoreceptors through CNG channels can potentially delay disease progression and improve the quality of life for patients. To find inhibitors for rod CNG channels, we investigated the impact of 16 cGMP analogues on both rod and cone CNG channels using the patch-clamp technique. Although modifications at the C8 position of the guanine ring did not change the ligand efficacy, modifications at the N1 and N2 positions rendered cGMP largely ineffective in activating retinal CNG channels. Notably, PET-cGMP displayed selective potential, favoring rod over cone, whereas Rp-cGMPS showed greater efficiency in activating cone over rod CNG channels. Ligand docking and molecular dynamics simulations on cyclic nucleotide-binding domains showed comparable binding energies and binding modes for cGMP and its analogues in both rod and cone CNG channels (CNGA1 vs CNGA3 subunits). Computational experiments on CNGB1a vs CNGB3 subunits showed similar binding modes albeit with fewer amino acid interactions with cGMP due to an inactivated conformation of their C-helix. In addition, no clear correlation could be observed between the computational scores and the CNG channel efficacy values, suggesting additional factors beyond binding strength determining ligand selectivity and potency. This study highlights the importance of looking beyond the cyclic nucleotide-binding domain and toward the gating mechanism when searching for selective modulators. Future efforts in developing selective modulators for CNG channels should prioritize targeting alternative channel domains
Prebiotic diet changes neural correlates of food decision-making in overweight adults: a randomised controlled within-subject cross-over trial
No full textAnimal studies suggest that prebiotic, plant-derived nutrients could improve homoeostatic and hedonic brain functions through improvements in microbiome–gut–brain communication. However, little is known if these results are applicable to humans. Therefore, we tested the effects of high-dosed prebiotic fibre on reward-related food decision-making in a randomised controlled within-subject cross-over study and assayed potential microbial and metabolic markers
Systematic distributions of interaction strengths across tree interaction networks yield positive diversity–productivity relationships
No full textUnderstanding the mechanisms underlying diversity–productivity relationships (DPRs) is crucial to mitigating the effects of forest biodiversity loss. Tree–tree interactions in diverse communities are fundamental in driving growth rates, potentially shaping the emergent DPRs, yet remain poorly explored. Here, using data from a large-scale forest biodiversity experiment in subtropical China, we demonstrated that changes in individual tree productivity were driven by species-specific pairwise interactions, with higher positive net pairwise interaction effects on trees in more diverse neighbourhoods. By perturbing the interactions strength from empirical data in simulations, we revealed that the positive differences between inter- and intra-specific interactions were the critical determinant for the emergence of positive DPRs. Surprisingly, the condition for positive DPRs corresponded to the condition for coexistence. Our results thus provide a novel insight into how pairwise tree interactions regulate DPRs, with implications for identifying the tree mixtures with maximized productivity to guide forest restoration and reforestation efforts
The Challenge to Stabilize, Extract and Analyze Urinary Cell-Free DNA (ucfDNA) during Clinical Routine
No full textBackground: The “Liquid Biopsy” has become a powerful tool for cancer research during the last decade. Circulating cell-free DNA (cfDNA) that originates from tumors has emerged as one of the most promising analytes. In contrast to plasma-derived cfDNA, only a few studies have investigated urinary cfDNA. One reason might be rapid degradation and hence inadequate concentrations for downstream analysis. In this study, we examined the stability of cfDNA in urine using different methods of preservation under various storage conditions. Methodology: To mimic patient samples, a pool of healthy male and female urine donors was spiked with a synthetic cfDNA reference standard (fragment size 170 bp) containing the T790M mutation in the EGFR gene. Spiked samples were preserved with three different buffers and with no buffer over four different storage periods (0 h; 4 h; 12 h; 24 h) at room temperature vs. 4 °C. The preservatives used were Urinary Analyte Stabilizer (UAS, Novosanis, Wijnegem, Belgium), Urine Conditioning Buffer (UCB, Zymo, Freiburg, Germany) and a self-prepared buffer called “AlloU”. CfDNA was extracted using the QIAamp MinElute ccfDNA Mini Kit (Qiagen, Hilden, Germany). CfDNA concentration was measured using the Qubit™ 4 fluorometer (Thermo Fisher Scientific, Waltham, MA, USA). Droplet digital PCR (ddPCR) was used for detection and quantification of the T790M mutation. Results: Almost no spiked cfDNA was recoverable from samples with no preservation buffer and the T790M variant was not detectable in these samples. These findings indicate that cfDNA was degraded below the detection limit by urinary nucleases. Stabilizing buffers showed varying efficiency in preventing this degradation. The most effective stabilizing buffer under all storage conditions was the UAS, enabling adequate recovery of the T790M variant using ddPCR. Conclusion: From a technical point of view, stabilizing buffers and adequate storage conditions are a prerequisite for translation of urinary cfDNA diagnostics into clinical routine
Targeting ion channels with ultra-large library screening for hit discovery
No full textIon channels play a crucial role in a variety of physiological and pathological
processes, making them attractive targets for drug development in diseases
such as diabetes, epilepsy, hypertension, cancer, and chronic pain. Despite the
importance of ion channels in drug discovery, the vastness of chemical space
and the complexity of ion channels pose significant challenges for identifying
drug candidates. The use of in silico methods in drug discovery has dramatically
reduced the time and cost of drug development and has the potential to
revolutionize the field of medicine. Recent advances in computer hardware
and software have enabled the screening of ultra-large compound libraries.
Integration of different methods at various scales and dimensions is becoming
an inevitable trend in drug development. In this review, we provide an overview
of current state-of-the-art computational chemistry methodologies for ultra-
large compound library screening and their application to ion channel drug
discovery research. We discuss the advantages and limitations of various in
silico techniques, including virtual screening, molecular mechanics/dynamics
simulations, and machine learning-based approaches. We also highlight several
successful applications of computational chemistry methodologies in ion channel
drug discovery and provide insights into future directions and challenges in
this field
Structure of Staphylococcus aureus ClpP Bound to the Covalent Active-Site Inhibitor Cystargolide A
No full textThe caseinolytic protease is a highly conserved serine protease, crucial to prokaryotic and eukaryotic protein homeostasis, and a promising antibacterial and anticancer drug target. Herein, we describe the potent cystargolides as the first natural β-lactone inhibitors of the proteolytic core ClpP. Based on the discovery of two clpP genes next to the cystargolide biosynthetic gene cluster in Kitasatospora cystarginea, we explored ClpP as a potential cystargolide target. We show the inhibition of Staphylococcus aureus ClpP by cystargolide A and B by different biochemical methods in vitro. Synthesis of semisynthetic derivatives and probes with improved cell penetration allowed us to confirm ClpP as a specific target in S. aureus cells and to demonstrate the anti-virulence activity of this natural product class. Crystal structures show cystargolide A covalently bound to all 14 active sites of ClpP from S. aureus, Aquifex aeolicus, and Photorhabdus laumondii, and reveal the molecular mechanism of ClpP inhibition by β-lactones, the predominant class of ClpP inhibitors
Temporal and spatial mapping of theoretical biomass potential across the European Union
No full textWith the increasing challenge to shift our economic system from carbon to renewable energy carriers, the demand for biogenic resources is growing. Biogenic municipal waste, agricultural by-products and industrial residues are under-utilised but are increasingly gaining in value. To date, there is no continuous database for these resources in the EU-27 countries. Existing datasets that estimate resource potentials for a single point in time often lack validation. A reliable and continuous database is thus needed to support the growing bioeconomy.
Spatial and temporal high-resolution data of biogenic residues serve as an invaluable resource for identifying areas with significant theoretical biomass potential and allows an in-depth understanding of dynamic patterns over time. This study elucidates the theoretical biomass potentials of 13 distinct biomasses from municipal waste, agricultural by-products and industrial residues quantified annually from 2010–2020. The spatial scope of the research covers the EU-27 Member States incorporating all entities represented at various levels within the Nomenclature of Territorial Units for Statistics (NUTS) as delineated by Eurostat, where possible. The regionalised data are subsequently validated against regional statistics from different countries. The findings demonstrate the feasibility of creating a time series of theoretical biomass potentials for the 13 selected waste types, by-products, and residues, and underscore the critical role of data validation when regionalising national or sub-national data to smaller NUTS entities. It could be shown that the values of small regions (NUTS 3) correlated well on average. When looking at individual regions in detail, regional characteristics such as the location of cultivation, waste management or reporting methods could lead to over- or underestimates of up to 100 %. Therefore, data at the regional level provide only limited reliability. In the case of industrial residues, regionalisation gave good results localising preference regions of high theoretical biomass potential, but more data on industrial production are needed to also estimate residual quantities at sub-national and local levels.
The biomass potentials modelled in this study have been published in an open-access database, which is designed as an extensible tool, enabling the understanding of national and regional trends of theoretical biomass potentials in the European Union and of the reliability of the regionalised data.
The estimated theoretical potential dataset can be downloaded free of charge from: https://doi.org/10.48480/g53t-ks72 [Titel anhand dieser DOI in Citavi-Projekt übernehmen] (Günther et al., 2023)