Expedient syntheses of neoglycoproteins using phase transfer catalysis and reductive amination as key reactions

Starting from peracetylated chloro- or bromo-glycosyl donors ofN-acetylneurmainic acid,N-acetylglucosamine, glucose and lactose, the correspondingp-formylphenyl glycosides were synthesized stereospecifically under phase transfer catalysed conditions at room temperature in yields of 38–67%. After Zemplén de-O-acetylation, the formyl groups were directly and chemoselectively coupled to the lysine residues of bovine serum albumin by reductive amination using sodium cyanoborohydride. The conjugation reactions were followed as a function of time and under a series of different molar ratios of the reactants to provide glycoconjugates of varying degree of antigenicities. Thus, carbohydrate protein conjugates were made readily available using essentially two key reactions.Presented in part at the 15th International Carbohydrate Symposium, Yokohama, Japan, August 12–17, 1990.     

Structure elucidation of the core regions from Citrobacter O4 and O36 lipopolysaccharides by chemical and enzymatic methods, gas chromatography/mass spectrometry, and NMR spectroscopy at 500 MHz

Novel enterobacterial core oligosaccharides were isolated from Citrobacter O4 and O36 lipopolysaccharides, and their structures were determined by methylation analysis, Smith degradation and enzymatic degradations, gas chromatography/mass spectrometry, and two-dimensional phase-sensitive correlated, relayed coherence transfer, double-quantum, triple-quantum-filtered, and nuclear Overhauser effect (NOE) 1H NMR spectroscopy at 500 MHz. In the formulas, all hexose residues are D-hexopyranoses, and heptoses are L-glycero-D-manno-heptopyranoses; the alternative locations of the side-chain heptose and pyrophosphorylethanolamine (PPEtN) residues are marked by dashed lines; dOclA stands for 3-deoxy-D-manno-octulosonic acid. (formula; see text) Along with these complete cores, incomplete ones, lacking the hexosamine trisaccharides, occur in the lipopolysaccharides of both types. Qualitative NOE data were in good agreement with the minimum energy conformation of the above O36 oligosaccharide, calculated with the aid of the SUGAR program [Sundin, A., Carter, R. E., & Liljefors, T. (1988) J. Mol. Graphics (in press)].     

Structure elucidation of the core octasaccharide from Citrobacter PCM 1487 with the aid of 500-MHz, two-dimensional phase-sensitive correlated, relayed-coherence transfer, double-quantum, triple-quantum filtered, and N.O.E. H-n.m.r. spectra

The main features of the primary structure of the octasaccharide, - -Glcp-(1→2)-- -Glcp-(1→2)-[- -GalpNAc- (1→3)]-- -Galp-(1→3)-- -Glcp-(1→3)-[- -Hepp-(1→7)]-- -Hepp-(1→3)-- -Hep, have been determined in the ab initio manner by ¹H-n.m.r. spectroscopy without resorting to biochemical methods of analysis. Several nontypical interresidue n.O.e. values point to a preferred solution conformation of the molecule.     

Structural studies of the Shigella flexneri variant X, type 5 a and type 5 b O-antigens.

Previous studies of different Shigella flexneri O-antigens indicate that their O-specific region is composed of oligosaccharide repeating units containing a basic tetrasaccharide structure, to which alpha-D-glucopyranosyl groups and O-acetyl groups may be attached to different positions. Structural studies of O-antigens from variant X, type 5a and type 5b lend further support to this assumption. These antigens contain terminal alpha-D-glucopyranosyl groups, one each per repeating unit in X and 5a, two in 5b. The location of these groups in the repeating unit has been determined.     

Core region of Citrobacter O23 lipopolysaccharide. Structure elucidation by chemical methods, gas chromatography/mass spectrometry and NMR spectroscopy at 500 MHz

A novel enterobacterial core region in Citrobacter O23 lipopolysaccharide is described. Its structure was determined by methylation analysis/mass spectrometry, chemical degradation and one- and two-dimensional 1H-NMR spectroscopy: [formula; see text] where PPEtN stands for diphosphorylethanolamine, and dOclA for 3-deoxy-D-manno-octulosonic acid.     

Hafnia alvei lipopolysaccharides: Isolation, sugar composition and SDS-PAGE analysis

Abstract Lipopolysaccharides (LPS) of 33 strains of Hafnia alvei were isolated and purified. LPS content of the dry bacterial mass ranged from 1.2 to 4.5%. All examined lipopolysaccharides contained glucose, glucosamine, heptose, 3-deoxy-octulosonic acid and often galactose. Rhamnose, mannose, galactosamine, mannosamine and unidentified amino sugars were found in some H. alvei strains. Sialic acid was present in LPS of one strain. d -3-Hydroxybutyryl groups also were identified in lipopolysaccharides of 5 strains of this genus.
SDS-PAGE of the lipopolysaccharides was presented in the paper. According to these results two core types exist in H. alvei .     

Light-enhanced dark respiration in leaves and mesophyll protoplasts of pea in relation to photorespiration,respiration and some metabolites content

The respiration rate of leaves and mesophyll protoplasts of pea (Pisum sativum L.), from plants which were previously kept in darkness for 24 h was doubled following a period of photosynthesis at ambient level of O₂ (21 %), whereas the low level of O₂ (1 % and 4 % for leaves and protoplasts, respectively) reduced this light-enhanced dark respiration (LEDR) to the rate as noted before the illumination. Similarly to respiration rate, the oxygen at used concentrations had no effect on the ATP/ADP ratio in the dark-treated leaves. However, the ATP/ADP ratio in leaves photosynthesizing at 21 % O₂ was higher (up to 40 %, dependence on CO₂ concentration in the range 40–1600 1 dm⁻³) than in those photosynthesizing at 1 % O₂ or darkened at air (21 % O₂). Also, at 1 % O₂ the accumulation of malate was suppressed (by about 40 %), to a value noted for leaves darkened at 21 % O₂. The dark-treatment of leaves reduced the ability of isolated mitochondria to oxidize glycine (by about twofold) and succinate, but not malate. Mitochondria from both the light- and dark-treated leaves did not differ in qualitative composition of free amino acids, however, there were significant quantitative differences especially with respect to aspartate, alanine, glutamate and major intermediates of the photorespiratory pathway (glycine, serine). Our results suggest that accumulation of photorespiratory and respiratory metabolites in pea leaves during photosynthesis at 1 % O₂ is reduced, hence the suppression of postillumination respiration rate.     

Essential Oils,Silver Nanoparticles and Propolis as Alternative Agents Against Fluconazole Resistant Candida albicans,Candida glabrata and Candida krusei Clinical Isolates

Development of effective and safe therapeutic treatment of fungal infections remains one of the major challenge for modern medicine. The aim of presented investigation was to analyze the in vitro antifungal activity of selected essential oils, ethanolic extracts of propolis and silver nanoparticles dropped on TiO₂ against azole-resistant C. albicans (n = 20), C. glabrata (n = 14) and C. krusei (n = 10) clinical isolates. Among tested essential oils, the highest activity has definitely been found in the case of the oil isolated from the bark of Cinnamomum cassia, with MIC and MFC values for all tested strains in the range of 0.0006–0.0097 % (v/v) and 0.0012–0.019 % (v/v), respectively. High activity was also observed for the Lemon, Basil, Thyme, Geranium and Clove (from buds) essential oils. Significant differences in fungicidal activity have been observed in the case of four tested propolis samples. Only one of them revealed high activity, with MFC values in the range from 0.156 to 1.25 % (v/v). Satisfactory fungicidal activity, against C. albicans and C. glabrata isolates, was also observed in the case of silver nanoparticles, however C. krusei isolates were mostly resistant. We also revealed that constituents of most of essential oils and propolis as well as silver nanoparticles are not substrates for drug transporters, which belong to the most important factors affecting resistance of Candida spp. clinical isolates to many of conventional antimycotics. To conclude, the results of our investigation revealed that essential oils, propolis and silver nanoparticles represent high potential for controlling and prevention candidiasis.     

Understanding the origins of bacterial resistance to aminoglycosides through molecular dynamics mutational study of the ribosomal A-site

Paromomycin is an aminoglycosidic antibiotic that targets the RNA of the bacterial small ribosomal subunit. It binds in the A-site, which is one of the three tRNA binding sites, and affects translational fidelity by stabilizing two adenines (A1492 and A1493) in the flipped-out state. Experiments have shown that various mutations in the A-site result in bacterial resistance to aminoglycosides. In this study, we performed multiple molecular dynamics simulations of the mutated A-site RNA fragment in explicit solvent to analyze changes in the physicochemical features of the A-site that were introduced by substitutions of specific bases. The simulations were conducted for free RNA and in complex with paromomycin. We found that the specific mutations affect the shape and dynamics of the binding cleft as well as significantly alter its electrostatic properties. The most pronounced changes were observed in the U1406C∶U1495A mutant, where important hydrogen bonds between the RNA and paromomycin were disrupted. The present study aims to clarify the underlying physicochemical mechanisms of bacterial resistance to aminoglycosides due to target mutations.