A Uniform Benchmark for Testing SsrA-Derived Degrons in the Escherichia coli ClpXP Degradation Pathway

The ssrA degron is commonly used in fusion proteins to control protein stability in bacteria or as an interaction module. These applications often rely on the modular activities of the ssrA tag in binding to the SspB adaptor and in engaging the ClpXP protease. However, a comparison of these activities for a substantial standard set of degron variants has not been conducted previously, which may hinder the development of new variants optimized exclusively for one application. Here, we strive to establish a benchmark that will facilitate the comparison of ssrA variants under uniform conditions. In our workflow, we included methods for expression and purification of ClpX, ClpP, SspB and eGFP-degrons, assays of ClpX ATPase activity, of eGFP-degron binding to SspB and for measuring eGFP-degron degradation in vitro and in vivo. Using uniform, precise and sensitive methods under the same conditions on a range of eGFP-degrons allowed us to determine subtle differences in their properties that can affect their potential applications. Our findings can serve as a reference and a resource for developing targeted protein degradation approaches.     

Diketopiperazine-Based,Flexible Tadalafil Analogues: Synthesis,Crystal Structures and Biological Activity Profile

Phosphodiesterase 5 (PDE5) is one of the most extensively studied phosphodiesterases that is highly specific for cyclic-GMP hydrolysis. PDE5 became a target for drug development based on its efficacy for treatment of erectile dysfunction. In the present study, we synthesized four novel analogues of the phosphodiesterase type 5 (PDE5) inhibitor—tadalafil, which differs in (i) ligand flexibility (rigid structure of tadalafil vs. conformational flexibility of newly synthesized compounds), (ii) stereochemistry associated with applied amino acid building blocks, and (iii) substitution with bromine atom in the piperonyl moiety. For both the intermediate and final compounds as well as for the parent molecule, we have established the crystal structures and performed a detailed analysis of their structural features. The initial screening of the cytotoxic effect on 16 different human cancer and non-cancer derived cell lines revealed that in most cases, the parent compound exhibited a stronger cytotoxic effect than new derivatives, except for two cell lines: HEK 293T (derived from a normal embryonic kidney, that expresses a mutant version of SV40 large T antigen) and MCF7 (breast adenocarcinoma). Two independent studies on the inhibition of PDE5 activity, based on both pure enzyme assay and modulation of the release of nitric oxide from platelets under the influence of tadalafil and its analogues revealed that, unlike a reference compound that showed strong PDE5 inhibitory activity, the newly obtained compounds did not have a noticeable effect on PDE5 activity in the range of concentrations tested. Finally, we performed an investigation of the toxicological effect of synthesized compounds on Caenorhabditis elegans in the highest applied concentration of 6a,b and 7a,b (160 μM) and did not find any effect that would suggest disturbance to the life cycle of Caenorhabditis elegans. The lack of toxicity observed in Caenorhabditis elegans and enhanced, strengthened selectivity and activity toward the MCF7 cell line made 7a,b good leading structures for further structure activity optimization and makes 7a,b a reasonable starting point for the search of new, selective cytotoxic agents.     

Vibrational Circular Dichroism Unravels Supramolecular Chirality and Hydration Polymorphism of Nucleoside Crystals

Vibrational circular dichroism (VCD) spectroscopy has been widely used to study (bio)molecules in solution. However, its solid-state applications have been restricted due to experimental limitations and artifacts. Having overcome some of them, the first VCD study of nucleoside crystals is now presented. A two-orders-of-magnitude enhancement of VCD signal was observed due to high molecular order in the crystals and resulting supramolecular chirality. This allowed to obtain high-quality VCD spectra within minutes using minute amounts of samples. The VCD technique is extremely sensitive in detecting changes in a crystal order and is able to distinguish different hydration states of crystals. This elevates it to a new level, as a fast and efficient tool to study chiral crystalline samples. This study demonstrates that VCD is capable of near-instantaneous detection of hydration polymorphs and crystal degradation, which is of substantial interest in pharmaceutical industry (quality and stability control).     

Novel Mesoporous Organosilicas with Task Ionic Liquids: Properties and High Adsorption Performance for Pb(II)

Removal of toxic contaminants such as Pb(II) from waste solutions is environmentally requested. Therefore, in this paper, for potential novel sorbents, mesoporous ionic liquid-functionalized silicas were synthesized and tested for the removal of Pb(II) from aqueous solutions. The successful synthesis of the adsorbents was proved by nuclear magnetic resonance (²⁹Si and ¹³C NMR), Fourier transform infrared spectroscopy (FTIR), and elemental analysis. The structural and textural properties were determined using scanning electron microscopy (SEM), X-ray diffraction (XRD), high-resolution transmission electron microscopy (TEM), and low-temperature N₂ sorption, and the result showed that the applied procedure made it possible to obtain highly ordered particles with a two-dimensional mesostructure. The effects of several parameters including initial pH, contact time, adsorption temperature, and Pb(II) concentration were studied in detail and were discussed to evaluate the adsorption properties of the fabricated materials towards Pb(II). The obtained results confirmed a very high potential of the sorbents; however, the adsorption properties depend on the structure and amounts of the functional group onto fabricated materials. The sample ILS-Ox3-40 showed fast kinetics (equilibrium reached within 10 min) and capacity of 172 mg/g, and that makes it a promising sorbent for the cleanup of water contaminated by lead. It was also indicated that, regardless on structure of the tested materials, the Pb(II) removal was spontaneous and exothermic. The fabricated mesoporous silicas exhibited that they were easy to regenerate and had excellent reusability.     

Solvent‐Driven Conformational Exchange for Amide‐Linked Bichromophoric BODIPY Derivatives

The fluorescence lifetime and quantum yield are seen to depend in an unexpected manner on the nature of the solvent for a pair of tripartite molecules composed of two identical boron dipyrromethene (BODIPY) residues attached to a 1,10‐phenanthroline core. A key feature of these molecular architectures concerns the presence of an amide linkage that connects the BODIPY dye to the heterocyclic platform. The secondary amide derivative is more sensitive to environmental change than is the corresponding tertiary amide. In general, increasing solvent polarity, as measured by the static dielectric constant, above a critical threshold tends to reduce fluorescence but certain hydrogen bond accepting solvents exhibit anomolous behaviour. Fluorescence quenching is believed to arise from light‐induced charge transfer between the two BODIPY dyes, but thermodynamic arguments alone do not explain the experimental findings. Molecular modelling is used to argue that the conformation changes in strongly polar media in such a way as to facilitate improved rates of light‐induced charge transfer. These solvent‐induced changes, however, differ remarkably for the two types of amide.     

Explosive hazards in polyaniline chemistry

Polyaniline, a conducting polymer, is prepared by the oxidation of aniline with various oxidants. When using ammonium peroxydisulfate or hydrogen peroxide at high concentrations, the temperature may exceed boiling point of reaction mixture, which then explodes. This represents a safety hazard in the synthesis of polyaniline. Also polyaniline samples prepared in the solutions of nitric acid may spontaneously ignite or decompose during the characterization. The synthesis using the recommended protocol is safe.     

Influence of the Regression Technique on the Extrapolated Retention Values of Single-Point Adsorption Compounds on Silica Gel

JPC – Journal of Planar Chromatography – Modern TLC - The retention (RM) values of nine one-point adsorption model compounds: diphenylamine, indol, 2-naphtol, 1-naphtol, 1-naphtylamine,...     

Nano-titanium oxide doped with gold,silver, and palladium — synthesis and structural characterization

Nano-titania doped with noble metals (Au/TiO₂, Ag/TiO₂, Pd/TiO₂) has been synthesized by mild hydrolysis of the mixture of metal salts or complexes and titanium isopropoxide ((iPr-O)₄Ti). After thermal decomposition of the obtained precursors, nanomaterials were formed. Morphological characterization of the nanomaterials was provided by scanning electron microscopy (SEM) and stereological analysis, determining the BET specific surface area, and BJH nanoporosity (pore volume, pore size). It has been found that the structure of nanomaterials (size of nanoparticles and agglomerates) depended strongly on the method of the (iPr-O)₄Ti hydrolysis. A minor dependence on the kind of solvents and precursors of noble metals was observed. The presence of doping metal nanoparticles was confirmed by transmission electron microscopy (TEM) and energy dispersive X-ray spectroscopy (EDX). Nanomaterial phases were identified by X-ray diffraction (XRD). According to the XRD patterns, Ag/TiO₂ and Pd/TiO₂ products with doping metals in their oxidized form contain Ag-Ti and Pd-Ti phases. Peaks of the metal oxides Ag₂O and PdO are absent in the XRD patterns. The average size of TiO₂ nanoparticles is situated in the region of 20–60 nm, whereas metals are present as about 10–15 nm sized particles and fine nanoparticles.     

In vivo evaluation of small-molecule thermoresponsive anticancer drugs potentiated by hyperthermia

Hyperthermia used as an adjuvant with chemotherapy is highly promising in the treatment of certain cancers. Currently, the small molecule drugs used in combination with hyperthermia were not designed for this application. Herein, we report the evaluation of a chlorambucil and a ruthenium compound modified with a long fluorous chain, which exhibit thermoresponsive activity in colorectal adenocarcinoma xenografts in athymic mice in combination with mild hyperthermia (42 °C). Intraperitoneal injection of the derivatives followed by local hyperthermia showed a synergistic tumor growth reduction by 79% and 90% for the chlorambucil and ruthenium-based derivatives, respectively, with the latter exhibiting a higher synergy in combination with hyperthermia compared to the monotherapies. Histological analysis shows that both derivatives in combination with hyperthermia significantly decrease the number of proliferating tumor cells.     

New procedure for the control of the treatment of industrial effluents to remove volatile organosulfur compounds

We present a new procedure for the determination of volatile organosulfur compounds in samples of industrial effluents using dispersive liquid–liquid microextraction and gas chromatography with flame photometric detection. Initially, the extraction parameters were optimized. These included: type and volume of extraction solvent, volume of disperser solvent, salting out effect, pH, time and speed of centrifugation as well as extraction time. The procedure was validated for 30 compounds. The developed procedure has low detection limits of 0.0071–0.49 μg/L and a good precision (relative standard deviation values of 1.2–5.0 and 0.6–4.1% at concentrations of 1 and 10 μg/L, respectively). The procedure was used to determine the content of volatile organosulfur compounds in samples of effluents from the production of bitumens before and after chemical treatment, in which six compounds were identified, including 2‐mercaptoethanol, thiophenol, thioanisole, dipropyl disulfide, 1‐decanethiol, and phenyl isothiocyanate at concentrations ranging from 0.47 to 8.89 μg/L. Problems in the determination of organosulfur compounds related to considerable changes in composition of the effluents, increase in concentration of individual compounds and appearance of secondary pollutants during effluent treatment processes are also discussed.