Gas chromatography-mass spectrometry for probing the structure and mechanism of action of enzyme active sites. The role of Glu-270 in carboxypeptidase A.

A new technique for the study of the mechanism of enzymes has been developed. An enzyme, modified by an active-site directed reagent, is digested by one or more proteases. The resulting mixture of oligopeptides is then analyzed directly by gas chromatography-mass spectrometry without the use of separation or isolation procedures. A comparison with unmodified enzyme identifies the modified residue as well as quantifies the reaction. This approach has been applied to the identification of Glu-270 in the active site of carboxypeptidase A using a carbodiimide as modification reagent. Studies on the possible incorporation of 18O (from 18O-enriched water) into Glu-270 or other acidic residues near the active site of carboxypeptidase A show that the oxygens of the carboxyl groups of these residues are not exchangeable.     

Intracellular localization of GDP-l-fucose: Glycoprotein and CMP-sialic acid: Apolipoprotein glycosyltransferases in rat and pork livers

A GDP-l-fucose:glycoprotein fucosyltransferase which transfers l-fucose to terminal β-N-acetyl-d-glucosaminyl residues of sialidase-, β-galactosidase-treated α₁-acid glycoprotein and a CMP-sialic acid:glycoprotein sialyltransferase acting on sialidase-treated apolipoprotein-Ala₁ from human very low density lipoprotein have been shown to be concentrated in rat liver Golgi apparatus preparations at enrichments of 40- and 45-fold, respectively, and in pork liver Golgi-rich fractions at enrichments of 35- and 20-fold, respectively. A second fucosyltransferase acting on sialidase-treated α₁-acid glycopretein was absent from rat liver and was enriched only 13-fold in a pork liver Golgi-rich fraction. The smooth-surfaced microsome fraction was the only other rat liver subcellular fraction with appreciable levels of the GDP-l-fucose: β-N-acetyl-d-glucosaminide fucosyltransferase and the lipoprotein sialyltransferase (enrichments of 2.6- and 5.2-fold, respectivley). This enrichment could not be attributed to the plasma membrane content of the smooth microsome fraction since plasma membrane fractions from rat liver were shown to have relatively low concentrations of these two transferases (enrichments of 0.3 or less). Rat liver plasma membrane was also shown to have similarly low relative specific activities for three other glycosyltransferases (sialyl-, galactosyl-, and N-acetylglucosaminyl-). The accurate determination of the glycosyltransferase activities of the plasma membrane fraction required the use of relatively low concentrations of plasma membrane and relatively high concentrations of nucleotide-sugars in order to avoid interference by the high nucleotide-sugar pyrophosphatase and hydrolase activities of this fraction.     

Meta‐replication,sampling bias,and multi‐scale model selection: A case study on snow leopard (Panthera uncia) in western China

Replicated multiple scale species distribution models (SDMs) have become increasingly important to identify the correct variables determining species distribution and their influences on ecological responses. This study explores multi‐scale habitat relationships of the snow leopard (Panthera uncia) in two study areas on the Qinghai–Tibetan Plateau of western China. Our primary objectives were to evaluate the degree to which snow leopard habitat relationships, expressed by predictors, scales of response, and magnitude of effects, were consistent across study areas or locally landcape‐specific. We coupled univariate scale optimization and the maximum entropy algorithm to produce multivariate SDMs, inferring the relative suitability for the species by ensembling top performing models. We optimized the SDMs based on average omission rate across the top models and ensembles’ overlap with a simulated reference model. Comparison of SDMs in the two study areas highlighted landscape‐specific responses to limiting factors. These were dependent on the effects of the hydrological network, anthropogenic features, topographic complexity, and the heterogeneity of the landcover patch mosaic. Overall, even accounting for specific local differences, we found general landscape attributes associated with snow leopard ecological requirements, consisting of a positive association with uplands and ridges, aggregated low‐contrast landscapes, and large extents of grassy and herbaceous vegetation. As a means to evaluate the performance of two bias correction methods, we explored their effects on three datasets showing a range of bias intensities. The performance of corrections depends on the bias intensity; however, density kernels offered a reliable correction strategy under all circumstances. This study reveals the multi‐scale response of snow leopards to environmental attributes and confirms the role of meta‐replicated study designs for the identification of spatially varying limiting factors. Furthermore, this study makes important contributions to the ongoing discussion about the best approaches for sampling bias correction.     

Neural networks in intestinal immunoregulation

Key physiological functions of the intestine are governed by nerves and neurotransmitters. This complex control relies on two neuronal systems: an extrinsic innervation supplied by the two branches of the autonomic nervous system and an intrinsic innervation provided by the enteric nervous system. As a result of constant exposure to commensal and pathogenic microflora, the intestine developed a tightly regulated immune system. In this review, we cover the current knowledge on the interactions between the gut innervation and the intestinal immune system. The relations between extrinsic and intrinsic neuronal inputs are highlighted with regards to the intestinal immune response. Moreover, we discuss the latest findings on mechanisms underlying inflammatory neural reflexes and examine their relevance in the context of the intestinal inflammation. Finally, we discuss some of the recent data on the identification of the gut microbiota as an emerging player influencing the brain function.     

Lung tissue extracellular matrix-derived hydrogels protect against radiation-induced lung injury by suppressing epithelial–mesenchymal transition

This study aimed to examine whether lung tissue extracellular matrix (ECM) hydrogels have protective effects on radiation-induced lung injury (RILI). The cytocompatibility and histocompatibility were tested for the obtained ECM-derived hydrogel. Sprague–Dawley rats were randomly divided into three groups (n = 18): control group (control); rats receiving irradiation and intratracheal injection of normal saline (IR + NS); and rats receiving irradiation and intratracheal injection of lung ECM-derived hydrogel (IR + ECM). The wet/dry weight ratio was used to evaluate the congestion and edema of the lungs. Histopathological analysis of lung tissues was performed using hemotoxylin and eosin staining and Masson's trichrome staining. Immunohistochemical staining and western blot analyses were carried out to determine the expression of epithelial–mesenchymal transition (EMT)-related proteins in lung tissues (E-cadherin, α-smooth muscle actin [α-SMA], and vimentin). In addition, tumor necrosis factor-α (TNF-α), transforming growth factor-β1 (TGF-β1) and interleukin-6 (IL-6), hydroxyproline, malondialdehyde (MDA), and superoxide dismutase (SOD) levels were also evaluated. The ECM-derived hydrogels had good cytocompatibility and histocompatibility. ECM-derived hydrogel treatment improved lung histopathology injury and pulmonary edema. Higher expression of E-cadherin and lower expression of vimentin and α-SMA were found in the IR + ECM group compared with those in the IR + NS group. Hydroxyproline levels were reduced by ECM-derived hydrogel treatment compared with those in the IR + NS group. Obvious increases of TNF-α, IL-6, and TGF-β1 were identified following irradiation. Marked reductions in MDA content and increases in SOD were induced by ECM-derived hydrogel treatment in rats after radiation. ECM-derived hydrogels were shown to protect against RILI, potentially by reducing EMT, inflammation, and oxidative damage.     

Biocides formulation of essential oils having antimicrobial activity

Essential oils of fennel, peppermint, caraway, eucalyptus, geranium and lemon were tested for their antimicrobial activities against some plant pathogenic micro-organisms (Fusarium oxysporum, Alternaria alternate, Penicilium italicum Penicilium digitatum and Botyritus cinerea). Essential oils of fennel, peppermint, caraway were selected as an active ingredient for the formulation of biocides due to their efficiency in controlling the tested micro-organisms. Successful emulsifiable concentrates (biocides) were prepared from these oils using different emulsifiers (Emulgator B.L.M. Tween20 and Tween80) and different fixed oils (sesame, olive, cotton and soybean oils). Physico-chemical properties of the formulated biocide (spontaneous emulsification, emulsion stability test, cold stability and heat stability tests as well as viscosity, surface tension and pH) were measured. The prepared biocides were ready to be tested for application in a future work as a safe pesticide against different pathogens.     

Studies on Bleomycin-DNA and Bleomycin-Iron Interactions

Abstract

The bleomycins, a group of antitumor antibiotics (Figure 1), cause the degradation of DNA by a process requiring iron(II) and dioxygen (1,2). DNA degradation appears to involve two steps: association of the drug with the nucleic acid and degradation of the DNA. As part of studies directed toward achieving an understanding of how the bleomycins degrade DNA, we have examined various properties of the drug using a variety of chemical and physico- chemical techniques, including NMR and Mössbauer spectroscopy. We have studied both the interaction of the antibiotic with its target (DNA) as well as its association with its metal ion cofactor. This work has been performed on the intact drug and its derivatives as well as on synthetic models of the parent drug. This paper reviews and updates the recent work from this laboratory on the bleomycins.