5-(Sulfamoyl)thien-2-yl 1,3-oxazole inhibitors of carbonic anhydrase II with hydrophilic periphery

Hydrophilic derivatives of an earlier described series of carbonic anhydrase inhibitors have been designed, prepared and profiled against a panel of carbonic anhydrase isoforms, including the glaucoma-related hCA II. For all hydrophilic derivatives, computational prediction of intraocular permeability routes showed the predominance of conjunctival rather than corneal absorption. The potentially reactive primary or secondary amine periphery of these compounds makes them suitable candidates for bioconjugation to polymeric drug carriers. As was shown previously, the most active hCA II inhibitor is efficacious in alleviating intraocular pressure in normotensive rabbits with efficacy matching that of dorzolamide.     

Photodecomposition of Sumithion® [O,O-Dimethyl-O-(3-methyl-4-nitrophenyl)-phosphorothioate]

When Sumithion solutions in various solvents in air or deposits on silica gel chromatoplates were irradiated with UV light or sunlight, the following products were obtained; carboxysumithion, sumioxon, carboxysumioxon, 3-methyl-4-nitrophenol, 3-carboxy-4-nitrophenol and methyl parathion (solutions). A trace amount of sumithion S-isomer was found in aqueous methanol solution irradiated with UV light in nitrogen and water solution exposed to sunlight. The same photoproducts in almost same proportions were formed on both chromatoplates and leaf surfaces of bean plants. Two major products, carboxysumithion and sumioxon, and 3-methyl-4-nitrophenol were further degraded into more polar products, and finally into polymeric hurnic acids. Certain of carbamate and pyrethroid insecticides slightly accelerated the photodecomposition of Sumithion.     

Novel GFP expression using a short N-terminal polypeptide through the defined twin-arginine translocation (Tat) pathway

Escherichia coli is frequently used as a convenient host organism for soluble recombinant protein expression. However, additional strategies are needed for proteins with complex folding characteristics. Here, we suggested that the acidic, neutral, and alkaline isoelectric point (pI) range curves correspond to the channels of the E. coli type-II cytoplasmic membrane translocation (periplasmic translocation) pathways of twin-arginine translocation (Tat), Yid, and general secretory pathway (Sec), respectively, for unfolded and folded target proteins by examining the characteristic pI values of the N-termini of the signal sequences or the leader sequences, matching with the known diameter of the translocation channels, and analyzing the N-terminal pI value of the signal sequences of the Tat substrates. To confirm these proposed translocation pathways, we investigated the soluble expression of the folded green fluorescent protein (GFP) with short N-terminal polypeptides exhibiting pI and hydrophilicity separately or collectively. This, in turn, revealed the existence of an anchor function with a specific directionality based on the N-terminal pI value (termed as N-terminal pI-specific directionality) and distinguished the presence of the E. coli type-II cytoplasmic membrane translocation pathways of Tat, Yid, and Sec for the unfolded and folded target proteins. We concluded that the pI value and hydrophilicity of the short N-terminal polypeptide, and the total translational efficiency of the target proteins based on the ΔGRNA value of the N-terminal coding regions are important factors for promoting more efficient translocation (secretion) through the largest diameter of the Tat channel. These results show that the short N-terminal polypeptide could substitute for the Tat signal sequence with improved efficiency.     

Hydrophilicity effect on CO2/CH4 separation using carbon nanotube membranes: insights from molecular simulation

Abstract

Molecular simulation methods were applied to study the effect of hydrophilicity on CO₂/CH₄ separation using carbon nanotube (CNT) membranes. CNTs with a diameter of ~1 nm were functionalised by varying amounts of carbonyl groups, in order to achieve various hydrophilicity. The presence of –CO groups inside the CNT allow a significant gain in the diffusion selectivity of CO₂, while in contrast the adsorption selectivity is hardly changed. The corresponding permeation selectivity increases as the hydrophilicity of the CNT-based membrane increases. However, the permeability of CO₂ decreases due to a combination of the intermolecular interactions between the gas and functional groups and the steric effects of the added functional groups. Considering both the permeation selectivity and permeability, it was found that the maximum separation performance is achieved in a certain hydrophilic CNT membrane. Moreover, the separation performance of hydrophilic CNTs for CO₂/CH₄ mixtures breaks the Robeson upper bound.     

Engineering an EGFR-binding Gp2 domain for increased hydrophilicity

The Gp2 domain is a 45 amino-acid scaffold that has been evolved for specific, high-affinity binding towards multiple targets and was proven useful in molecular imaging and biological antagonism. It was hypothesized that Gp2 may benefit from increased hydrophilicity for improved physiological distribution as well as for physicochemical robustness. We identified seven exposed hydrophobic sites for hydrophilic mutations and experimentally evaluated single mutants, which yielded six mutations that do not substantially hinder expression, binding affinity or specificity (to epidermal growth factor receptor), and thermal stability. Eight combinations of these mutations improved hydrophilicity relative to the parental Gp2 clone as assessed by reverse-phase high-performance liquid chromatography (p < 0.05). Secondary structures and refolding abilities of the selected single mutants and all multimutants were unchanged relative to the parental ligand. A variant with five hydrophobic-to-hydrophilic mutations was identified with enhanced solubility as well as reasonable binding affinity ( K _d = 53–63 nM), recombinant yield (1.3 ± 0.8 mg/L), and thermal stability ( T _m = 53 ± 3°C). An alternative variant with a cluster of three leucine-to-hydrophilic mutations was identified with increased solubility, nominally increased binding affinity ( K _d = 13–28 nM) and reasonable thermal stability ( T _m = 54.0 ± 0.6°C) but reduced yield (0.4 ± 0.3 mg/L). In addition, a ≥7°C increase in the midpoint of thermal denaturation was observed in one of the single mutants (T21N). These mutants highlight the physicochemical tradeoffs associated with hydrophobic-to-hydrophilic mutation within a small protein, improve the solubility and hydrophilicity of an existent molecular imaging probe, and provide a more hydrophilic starting point for discovery of new Gp2 ligands towards additional targets.     

Surface Wettability and Chemistry of Ozone Perfusion Processed Porous Collagen Scaffold

Crosslinking treatment of collagen has often been used to improve the biological stability and mechanical properties of 3D porous collagen scaffolds. However, accompanying these improvements, the collagen fibril surface becomes hydrophobic nature resulting in a reduced surface wettability. The wetting of the collagen fibril by culture medium is reduced and it is difficult for the medium to diffuse into the 3D structure of a porous collagen scaffold. This paper reports a “perfusion processing” strategy using ozone to improve the surface wettability of chemical crosslinked collagen scaffolds. Surface wettability, surface composition and biological stability were analyzed to evaluate the effectiveness of this surface processing strategy. It was observed that ozone perfusion processing improved surface wettability for both exterior and interior surfaces of the porous 3D collagen scaffold. The improvement in wettability is attributed to the incorporation of oxygen-containing functional groups onto the surface of the collagen fibrils, as confirmed by X-ray Photoelectron Spectroscopy (XPS) analysis. This leads to a significant improvement in water taking capability without compromising the bulk biological stability and mechanical properties, and confirms that ozone perfusion processing is an effective tool to modify the wettability both for interior and exterior surfaces throughout the scaffold.     

Triglyceride Release and Fatty Acid Oxidation by the Perfused Liver of Rats Fed a Low Protein Diet

Livers of growing rats fed a 5 or 20 protein calories percent (PC %) diet containing purified whole egg protein for three weeks were perfused in situ and the release of triglycerides (TG) and the oxidation of fatty acid by the liver alone were estimated by infusing palmitic acid-l-¹⁴C to the perfusion medium.

The release of TG from the liver of the 5 PC% group was significantly lower in both unfractionated perfusate plasma and perfusate plasma very low density lipoprotein (VLDL) than that of the 20 PC% group, whereas the content of liver TG of the 5 PC% group was higher than that of the 20 PC% group. Significantly lower radioactivity appeared in TG of both unfractionated perfusate plasma and perfusate plasma VLDL of the 5 PC% group than that of the 20 PC% group, while total radioactivity of liver TG was higher in the 5 PC% group than in the 20 PC% group. The ¹⁴CO₂ production in the perfused liver of the 5 PC% group increased gradually with time rather than decreased in comparison with that of the 20 PC% group.

These findings suggest that a major factor responsible for the liver lipid accumulation in rats fed the low protein diet is not an impaired fatty acid oxidation in the liver but an impaired secretion of TG from the liver.