Solvent interactions and conformational choice in a core N-glycan segment: gas phase conformation of the central, branching trimannose unit and its singly hydrated complex

The intrinsic conformational preferences and structures of the branched trimannoside, alpha-phenyl 3,6-di-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranoside (which contains the same carbohydrates found in a key subunit of the core pentasaccharide in N-glycans) and its singly hydrated complex, have been investigated in the gas phase isolated at low temperature in a molecular beam expansion. Conformational assignments of their infrared ion dip spectra, based on comparisons between experiment and ONIOM (B3LYP/6-31+G(d):HF/6-31G(d)) and single-point MP2 calculations have identified their preferred structures and relative energies. The unhydrated trimannoside populates a unique structure supported by two strong, central hydrogen bonds linking the central mannose unit (CM), and its two branches (3M and 6M) closely together, through a cooperative hydrogen-bonding network: OH4(CM)-->OH6(3M)-->OH6(6M). A closely bound structure is also retained in the singly hydrated oligosaccharide, with the water molecule bridging across the 3M and 6M branches to provide additional bonding. This structure contrasts sharply with the more open, entropically favored trimannoside structure determined in aqueous solution at 298 K. In principle this structure can be accessed from the isolated trimannoside structure by a simple conformational change, a twist about the alpha(1,3) glycosidic linkage, increasing the dihedral angle psi[C1(3M)-O3(3M)-C3(CM)-C2(CM)] from approximately 74 degrees to approximately 146 degrees to enable accommodation of a water molecule at the centrally bound site occupied by the hydroxymethyl group on the 3M ring and mediation of the water-linked hydrogen-bonded network: OH4(CM) -->OH(W)-->OH6(6M). The creation of a "water pocket" motif localized at the bisecting axis of the trimannoside is strikingly similar to the structure of more complex N-glycans in water, suggesting perhaps a general role for the "bisecting" OH4 group in the central (CM) mannose unit.     

Mixed-phase PdRu bimetallic structures with high activity and stability for formic acid electrooxidation

Aiming at investigating the effect of structure on electrocatalytic properties, Pd(50)Ru(50) nanoparticles (NPs) with three different structures were carefully designed in a one-pot polyol process for application in formic acid electrooxidation. The three structures are: (1) single-phase PdRu nanodendrites (denoted as PR-1), (2) a mixed-phase mixture of PdRu nanodendrites and monometallic Ru NPs (denoted as PR-2), and (3) a mixed-phase mixture of monometallic Pd and Ru NPs (denoted as PR-3). From PR-1 to PR-3, the structure was varied from single-phase to mixed-phase. The relative position of Ru was altered from completely Pd-connected (PR-1), to a mixture of Pd-connected and monometallic (PR-2), and completely monometallic (PR-3). All PdRu NPs outperform the commercial Pd/C. PR-2 exhibits the highest peak current density, but its stability is slightly lower than that of PR-3. When both the current density and the durability are taken into consideration, PR-2 is the best choice of catalyst for formic acid oxidation. It indicates that both the Pd-connected Ru NPs and monometallic Ru NPs in the mixed-phase PR-2 are essential to improve the electrocatalytic properties. Our study also illustrates that the electrochemical active surface area (ECSA) and hydrogen storage capacity of the as-prepared PdRu NPs are greatly enhanced after several hundred scans in formic acid, indicating the possibility for highly restorable catalysts in direct formic acid fuel cells.     

Synthesis of oligosaccharides corresponding to Vibrio cholerae O139 polysaccharide structures containing dideoxy sugars and a cyclic phosphate

A spacer-equipped tetrasaccharide, p-aminocyclohexylethyl alpha-l-Colp-(1-->2)-beta-d-Galp-(1-->3)-[alpha-l-Colp-(1-->4)]-beta-D-GlcpNAc, containing a 4,6-cyclic phosphate in the galactose residue, has been synthesised. The structure corresponds to a part of the repeating unit of the capsular (and lipo-) polysaccharide of the endemic bacteria Vibrio cholerae type O139 synonym Bengal. The synthetic strategy allows continuous syntheses of the complete O139 hexasaccharide repeating unit as well as of the structurally related repeating unit of serotype O22. Starting from ethyl 2-azido-4,6-O-benzylidene-2-deoxy-1-thio-beta-D-glucopyranoside, a thioglycoside tetrasaccharide donor block was constructed through two orthogonal glycosylations with glycosyl bromide donors. First, a properly protected galactose moiety was introduced using silver triflate as promoter and subsequently the two colitose residues, carrying electron-withdrawing protecting groups for stability reasons, under halide-assisted conditions. The tetrasaccharide block was then linked to the spacer in a NIS-TMSOTf-promoted coupling. Transformation of the azido group into an acetamido group using H2S followed by removal of temporary protecting acetyl groups gave a 4',6'-diol, which was next phosphorylated with methyl dichlorophosphate and deprotected to yield the 4,6-cyclic phosphate tetrasaccharide target structure.     

A new bioactive triterpenoid saponin from the seeds of Lactuca scariola Linn

A new triterpenoid saponin, 3-O-[beta-D-galactopyranosyl-(1 --> 3)-O-beta-D-xylopyranosyl-(1 --> 4)-O-alpha-L-rhamnopyranosyl]-oleanolic acid; has been isolated from the seeds of Lactuca scariola. Its structure was determined by various spectral analysis and chemical degradations. This compound shows antimicrobial activity against various bacteria and fungi.     

Spectroscopic, structural, and conformational properties of (Z)-4,4,4-trifluoro-3-(2-hydroxyethylamino)-1-(2-hydroxyphenyl)-2-buten-1-one, C12H12F3NO3: a trifluoromethyl-substituted β-aminoenone

The (Z)-4,4,4-trifluoro-3-(2-hydroxyethylamino)-1-(2-hydroxyphenyl)-2-buten-1-one (C(12)H(12)F(3)NO(3)) compound was thoroughly studied by IR, Raman, UV-visible, and (13)C and (19)F NMR spectroscopies. The solid-state molecular structure was determined by X-ray diffraction methods. It crystallizes in the P2(1)/c space group with a = 12.1420(4) ?, b = 7.8210(3) ?, c = 13.8970(5) ?, β = 116.162(2)°, and Z = 4 molecules per unit cell. The molecule shows a nearly planar molecular skeleton, favored by intramolecular OH···O and NH···O bonds, which are arranged in the lattice as an OH···O bonded polymer coiled around crystallographic 2-fold screw-axes. The three postulated tautomers were evaluated using quantum chemical calculations. The lowest energy tautomer (I) calculated with density functional theory methods agrees with the observed crystal structure. The structural and conformational properties are discussed considering the effect of the intra- and intermolecular hydrogen bond interactions.     

Novel bioactive constituents from Enhydra fluctuans LOUR

A novel bioactive isoflavone glycoside 4',5,6,7-tetrahydroxy-8-methoxyisoflavone-7-O-beta-D-galactopyranosyl-(1 --> 3)-O-beta-D-xylopyranosyl-(1 --> 4)-O-alpha-l-rhamnopyranoside from the methanolic extract of the leaves of Enhydra fluctuans Lour. Its structure A was elucidated by various spectral analysis and chemical degradations. This compound exhibited microbial activity against various bacteria and fungi.     

Molecular dynamics simulation of the PEO sulfonic acid anion in water

Atomistic molecular modeling has been used to study the sulfonic acid anion of poly(ethylene oxide) (PEO sulfonic acid anion) in vacuum and a polymer electrolyte system consisting of the PEO sulfonic acid anion in water. The vibrational spectra of the molecules were simulated by the local mode method and found to be in good agreement with the experimental IR and Raman spectra. The structure of PEO sulfonic acid anion was studied in vacuum and water and compared to the structure of an isolated PEO sulfonic acid in vacuum. The simulated value for the root mean square end-to-end distance for the PEO sulfonic acid anion was 22 Å in vacuum and 12 Å in water. The root mean square radius of gyration of the PEO sulfonic acid anion was 8.4 Å in vacuum and 5.6 Å in water. The PEO sulfonic acid anion was randomly coiled in water and in an extended shape in vacuum.     

Bipartite network analysis reveals metabolic gene expression profiles that are highly associated with the clinical outcomes of acute myeloid leukemia

Dysregulated and reprogrammed metabolism are one of the most important characteristics of cancer, and exploiting cancer cell metabolism can aid in understanding the diverse clinical outcomes for patients. To investigate the differences in metabolic pathways among patients with acute myeloid leukemia (AML) and differential survival outcomes, we systematically conducted microarray data analysis of the metabolic gene expression profiles from 384 patients available from the Gene Expression Omnibus and Cancer Genome Atlas databases. Pathway enrichment analysis of differentially expressed genes (DEGs) showed that the metabolic differences between low-risk and high-risk patients mainly existed in two pathways: biosynthesis of unsaturated fatty acids and oxidative phosphorylation. Using the gene-pathway bipartite network, 62 metabolic genes were identified from 272 DEGs involved in 88 metabolic pathways. Based on the expression patterns of the 62 genes, patients with shorter overall survival (OS) durations in the training set (hazard ratio (HR) = 1.58, p = 0.038) and in two test sets (HR = 1.69 and 1.56 and p = 0.089 and 0.029, respectively) were well discriminated by hierarchical clustering analysis. Notably, the expression profiles of ALAS2, BCAT1, BLVRB, and HK3 showed distinct differences between the low-risk and high-risk patients. In addition, models for predicting the OS outcome of AML from the 62 gene signatures achieved improved performance compared with previous studies. In conclusion, our findings reveal significant differences in metabolic processes of patients with AML with diverse survival durations and provide valuable information for clinical translation.     

A 3D-impedimetric immunosensor based on foam Ni for detection of sulfate-reducing bacteria

A 3D-immunosensor based on simple and efficient trapping platform (foam Ni) combining with adsorption of gold nanoparticles and specific recognition of biological/chemical molecular has been reported for detection of sulfate-reducing bacteria (SRB) using electrochemical impedance spectroscopy (EIS). The impedance spectra were also used to characterize the successful construct and stepwise modification of the impedimetric immunosensors. This results show that a linear relationship between electron-transfer resistance (R_ct) values and the logarithm of the SRB concentrations was obtained for the SRB concentration range of 2.1 × 10¹–2.1 × 10⁷ cfu/ml. Additionally, the fabricated immunosensor shows a high selectivity against other bacteria.     

Facile method for the preparation of lyso-GM1 and lyso-GM2

This paper reports a facile method for the preparation of lyso-GM1 [Gal beta1-->3GalNAc beta1--> 4(Neu5Ac alpha2-->3)Galbeta1-->4Glc beta1-->1'-sphingosine] and lyso-GM2 [GalNAc beta1-->4(Neu5Ac alpha2-->3)Gal beta1-->4Glc beta1-->sphingosine], respectively, from GM1 [Galbeta1-->3GalNAc beta1-->4(Neu5Ac alpha2-->3)Galbeta1-->4Glc beta1-->1'-Cer] and GM2[GalNAc beta1-->4(Neu5Ac alpha2-->3)Galbeta1-->4Glc beta1-->1'-Cer], using sphingolipid ceramide deacylase and high performance anion-exchange chromatography (HPAEC). The enzymatically released lyso-GM1 and/or lyso-GM2 was effectively separated from its parent ganglioside by HPAEC using a Mono Q HR 5/5 column with an Amersham Biosciences fast protein liquid chromatography system. The yield was almost quantitative and the separation completed in approximately 3 h. This method is more convenient and effective than the conventional method using alkaline hydrolysis and silicic acid chromatography to generate and purify lyso-gangliosides.     

Comparison of aqueous molecular dynamics with NMR relaxation and residual dipolar couplings favors internal motion in a mannose oligosaccharide.

An investigation has been performed to assess how aqueous dynamical simulations of flexible molecules can be compared against NMR data. The methodology compares state-of-the-art NMR data (residual dipolar coupling, NOESY, and (13)C relaxation) to molecular dynamics simulations in water over several nanoseconds. In contrast to many previous applications of residual dipolar coupling in structure investigations of biomolecules, the approach described here uses molecular dynamics simulations to provide a dynamic representation of the molecule. A mannose pentasaccharide, alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->2)-D-Manp, was chosen as the model compound for this study. The presence of alpha-linked mannan is common to many glycopeptides, and therefore an understanding of the structure and the dynamics of this molecule is of both chemical and biological importance. This paper sets out to address the following questions. (1) Are the single structures which have been used to interpret residual dipolar couplings a useful representation of this molecule? (2) If dynamic flexibility is included in a representation of the molecule, can relaxation and residual dipolar coupling data then be simultaneously satisfied? (3) Do aqueous molecular dynamics simulations provide a reasonable representation of the dynamics present in the molecule and its interaction with water? In summary, two aqueous molecular dynamics simulations, each of 20 ns, were computed. They were started from two distant conformations and both converged to one flexible ensemble. The measured residual dipolar couplings were in agreement with predictions made by averaging the whole ensemble and from a specific single structure selected from the ensemble. However, the inclusion of internal motion was necessary to rationalize the relaxation data. Therefore, it is proposed that although residual dipolar couplings can be interpreted as a single-structure, this may not be a correct interpretation of molecular conformation in light of other experimental data. Second, the methodology described here shows that the ensembles from aqueous molecular dynamics can be effectively tested against experimental data sets. In the simulation, significant conformational motion was observed at each of the linkages, and no evidence for intramolecular hydrogen bonds at either alpha(1-->2) or alpha(1-->3) linkages was found. This is in contrast to simulations of other linkages, such as beta(1-->4), which are often predicted to maintain intramolecular hydrogen bonds and are coincidentally predicted to have less conformational freedom in solution.     

First synthesis of an α-d-Fucp3NAc containing oligosaccharide: a study on d-Fucp3NAc glycosylation

3-Acetamido-3,6-dideoxy-d-galactopyranose (d-Fucp3NAc) is an aminosugar almost exclusively found in phytopathogenic O-antigens. The glycosylation reaction involving d-Fucp3NAc donors was studied with several rhamnosyl acceptors, revealing that the best yields and highest α-stereoselectivity were obtainable by coupling a N-phenyl trifluoroacetimidate glycosyl donor in a ternary mixture (dioxane/DME/toluene 4:1:1) as solvent. For the first time a synthetic access to α-d-Fucp3NAc containing oligorhamnans, that are interesting molecules for studying the effects of O-antigen model oligosaccharides on the modulation of plant response to bacteria, was reported. An example is the pentasaccharide repeating unit of the major O-antigen component from Pseudomonas syringae pv. holci IMV 8300, which was synthesized as its methyl glycoside.     

A study on size effect of carboxymethyl starch nanogel crosslinked by electron beam radiation

The formation of carboxymethyl starch (CMS) nanogel with 50 nm less particle size was carried out through a radiation crosslinked process on the electron beam (EB) linear accelerator. Changes of intrinsic viscosities and weight averaged molecular weight in the CMS concentration, which ranged from 3 to 10 mg ml^?1 in absorbed doses were investigated. There were some new peaks in the ¹H NMR spectra of CMS nanogel compared with those of CMS polymer. These results were anticipated that the predominant intramolecular crosslinking of dilute CMS aqueous solution occurred while being exposed to a short intense pulse of ionizing radiation. Hydrodynamic radius (often called particle size, R_h) and distribution of particle size were measured by a dynamic light scattering technique. The radiation yield of intermolecular crosslinking of CMS solution was calculated from the expression of G_x (Charlesby, 1960, Jung-Chul, 2010). The influence of the “size effect” was demonstrated by testing culture of Lactobacillus bacteria on MRS agar culture medium containing CMS nanogel and polymer. Results showed that the number of Lactobacillus bacteria growing on nanogel containing culture medium is about 170 cfu/ml and on polymer containing culture medium is only 6 cfu/ml.     

Synthesis of the glycopeptidolipid of Mycobacterium avium Serovar 4: first example of a fully synthetic C-mycoside GPL

The preparation of the glycopeptidolipid (GPL) present in the cell wall of Mycobacterium aviumSerovar 4, namely 3,4-di-O-Me-alpha-L-Rhap-(1-->1)[R-C(21)H(43)CH(OH)CH(2)CO-D-Phe-[4-O-Me-alpha-L-Rhap-(1-->4)-2-O-Me-alpha-L-Fucp-(1-->3)-alpha-L-Rhap-(1-->2)-6-deoxy-alpha-L-Talp-(1-->3)]-D-allo-Thr-D-Ala-L-Alaol] (1), is described. The synthesis was based on the disconnection of the final structure into four building blocks, an L-rhamnosyl pseudodipeptide, a 6-deoxy-L-talosyl dipeptide, a trisaccharide donor, and a 3-hydroxyalkanoic acid. The key steps are the creation of the glycosidic linkage between the trisaccharide donor, used as a pentenyl glycoside, and the 6-deoxy-L-talose unit of an appropriate D-Phe-O-(6-deoxy-L-talosyl)-D-allo-Thr derivative and the final coupling of the two glycodipeptide fragments. Pentenyl glycosides were shown to provide useful donors in several glycosylation steps. This work constitutes the first synthesis of the full structure of a so-called "polar mycoside C" GPL.     

Conformation analysis and molecular mobility of ethylene and tetrafluoroethylene copolymer using solid-state 19F MAS and 1H --> 19F CP/MAS NMR spectroscopy

The changes in the conformation and molecular mobility accompanied by a phase transition in the crystalline domain were analyzed for ethylene (E) and tetrafluoroethylene (TFE) copolymer, ETFE, using variable-temperature (VT) solid-state 19F magic angle spinning (MAS) and 1H --> 19F cross-polarization (CP)/MAS NMR spectroscopy. The shifts of the signals for fluorines in TFE units to higher frequency and the continuing decrease and increase in the T1rho(F) values suggest that conformational exchange motions exist in the crystalline domain between 42 and 145 degrees C. Quantum chemical calculations of magnetic shielding constants showed that the high-frequency shift of TFE units should be induced by trans to gauche conformational changes at the CH2-CF2 linkage in the E-TFE unit. Although the 19F signals of the crystalline domain are substantially overlapped with those of the amorphous domain at ambient probe temperature (68 degrees C), they were successfully distinguished by using the dipolar filter and spin-lock pulse sequences at 145 degrees C. The dipolar coupling constants for the crystalline domain, which can be estimated by fitting the dipolar oscillation behaviors in the 1H --> 19F CP curve, showed a significant decrease with increasing temperature from 42 to 145 degrees C. This is due to the averaging of 1H-19F dipolar interactions originating from the molecular motion in the crystalline domain. The increase in molecular mobility in the crystalline domain was clearly shown by VT T1rho(F) and 1H --> 19F CP measurements in the phase transition temperature range.     

Characterization of Tryptophanase from Vibrio cholerae O1

Tryptophanase (Trpase) encoded by the tnaA gene catalyzes the conversion of tryptophan to indole, which is an extracellular signaling molecule detected in various bacteria including Vibrio cholerae. Indole has been demonstrated to regulate biofilm formation, drug resistance, plasmid maintenance and spore formation of bacteria. In the present study, the tnaA gene from V. cholerae O1 (VcTrpase) was cloned and expressed in E. coli BL21(DE3) tn5:tnaA (a Trpase-deficient competent). VcTrpase was purified by Ni²⁺-NTA chromatography. The obtained VcTrpase had a molecular mass of approximately 49 kDa, a specific activity of 3 U/mg protein, and absorption peaks at 330 and 435 nm. Using a site-directed mutagenesis technique, replacement of Arg419 by Val resulted in a VcTrpase completely devoid of activity. Thus, this site can be a target for drug design for controlling V. cholerae.     

Isolation and structure of a GD3-Type ganglioside molecular species possessing neuritogenic activity from the starfish Luidia maculata

A GD3-type ganglioside molecular species, LMG-4 (1), has been obtained from the polar lipid fraction of the chloroform/methanol extract of the starfish Luidia maculata. The structure of this ganglioside has been determined on the basis of chemical and spectroscopic evidence to be 1-O-[(N-acetyl-alpha-D-neuraminosyl)-(2-->8)-(N-acetyl-alpha-D-neuraminosyl)-(2-->3)-beta-D-galactopyranosyl-(1-->4)-beta-D-glucopyranosyl]-ceramide. The ceramide moiety was composed of heterogeneous 2-hydroxy fatty acid and phytosphingosine units. This is the first report on the isolation and structure elucidation of GD3-type ganglioside from echinoderms. Moreover, 1 exhibited neuritogenic activity toward the rat pheochromocytoma PC12 cells in the presence of nerve growth factor.     

An Antibody with a Variable‐Region Coiled‐Coil “Knob” Domain

The X‐ray crystal structure of a bovine antibody (BLV1H12) revealed a unique structure in its ultralong heavy chain complementarity determining region 3 (CDR3H) that folds into a solvent‐exposed β‐strand “stalk” fused to a disulfide crosslinked “knob” domain. We have substituted an antiparallel heterodimeric coiled‐coil motif for the β‐strand stalk in this antibody. The resulting antibody (Ab‐coil) expresses in mammalian cells and has a stability similar to that of the parent bovine antibody. MS analysis of H–D exchange supports the coiled‐coil structure of the substituted peptides. Substitution of the knob‐domain of Ab‐coil with bovine granulocyte colony‐stimulating factor (bGCSF) results in a stably expressed chimeric antibody, which proliferates mouse NFS‐60 cells with a potency comparable to that of bGCSF. This work demonstrates the utility of this novel coiled‐coil CDR3 motif as a means for generating stable, potent antibody fusion proteins with useful pharmacological properties.     

Saturation transfer difference 1D-TOCSY experiments to map the topography of oligosaccharides recognized by a monoclonal antibody directed against the cell-wall polysaccharide of group A streptococcus

A new saturation transfer difference 1D-TOCSY NMR experiment that allows the investigation of complex ligands interacting with proteins and its application in the mapping of which portions of oligosaccharide ligands (epitope) interact with a complementary antibody are described. The interaction between trisaccharide and hexasaccharide ligands, corresponding to fragments of the cell-wall polysaccharide of Streptococcus Group A, and a monoclonal antibody directed against the polysaccharide is investigated at the molecular level. The polysaccharide consists of alternating alpha-(1-->2) and alpha-(1-->3) linked L-rhamnopyranose (Rha) residues with branching N-acetyl-D-glucopyranosylamine (GlcNAc) residues linked beta-(1-->3) to alternate rhamnopyranose rings. The epitope is proven to consist not only of the immunodominant GlcNAc sugar but also of an entire branched trisaccharide repeating unit. The experimental NMR data serve to check and validate the computed models of the oligosaccharide-antibody complexes.