X-ray crystal structure of galabiose, O-alpha-D-galactopyranosyl-(1---4)-D-galactopyranose

O-alpha-D-Galactopyranosyl-(1---4)-D-galactopyranose, C12H22O11, Mr = 342.30, crystallises in the orthorhombic space group P2(1)2(1)2(1), and has alpha = 5.826(1), b = 13.904(3), c = 17.772(4) A, Z = 4, and Dx = 1.579 g.cm-3. Intensity data were collected with a CAD4 diffractometer. The structure was solved by direct methods and refined to R = 0.063 and Rw = 0.084 for 2758 independent reflections. The glycosidic linkage is of the type 1-axial-4-axial with torsion angles phi O-5' (O-5'-C-1'-O-1'-C-4) = 98.1(2) degrees, psi C-3 (C-3-C-4-O-1'-C-1') = -81.9(3) degrees, phi H (H-1'-C-1'-O-1'-C-4) = -18 degrees, and psi H (H-4-C-4-O-1'-C-1') = 35 degrees. The conformation is stabilised by an O-3 . . . O-5' intramolecular hydrogen-bond with length 2.787(3) A and O-3-H . . . O-5' = 162 degrees. The glycosidic linkage causes a folding of the molecule with an angle of 117 degrees between the least-square planes through the pyranosidic rings. The crystal investigated contained 56(1)% of alpha- and 44(1)% of beta-galabiose as well as approximately 70% of the gauche-trans and approximately 30% of the trans-gauche conformers about the exocyclic C-5'-C-6' and C-5-C-6 bonds. The crystal packing is governed by hydrogen bonding that engages all oxygen atoms except the intramolecular acceptor O-5' and the glycosidic O-1' oxygen atoms.     

Complete lipopolysaccharide of Plesiomonas shigelloides O74:H5 (strain CNCTC 144/92). 2. Lipid A, its structural variability, the linkage to the core oligosaccharide, and the biological activity of the lipopolysaccharide

Plesiomonas shigelloides is a Gram-negative bacterium associated with waterborne infections, which is common in tropical and subtropical habitats. Contrary to the unified antigenic classification of P. shigelloides, data concerning the structure and activity of their lipopolysaccharides (LPS and endotoxin) are limited. This study completes the structural investigation of phenol- and water-soluble fractions of P. shigelloides O74 (strain CNCTC 144/92) LPS with the emphasis on lipid A heterogeneity, describing the entire molecule and some of its biological in vitro activities. Structures of the lipid A and the affinity-purified decasaccharide obtained by de-N,O-acylation of P. shigelloides O74 LPS were elucidated by chemical analysis combined with electrospray ionization multiple-stage mass spectrometry (ESI-MS(n)), MALDI-TOF MS, and NMR spectroscopy. Lipid A of P. shigelloides O74 is heterogeneous, and three major forms have been identified. They all were asymmetric, phosphorylated, and hexaacylated, showing different acylation patterns. The beta-GlcpN4P-(1-->6)-alpha-GlcpN1P disaccharide was substituted with the primary fatty acids: (R)-3-hydroxytetradecanoic acid [14:0(3-OH)] at N-2 and N-2' and (R)-3-hydroxydodecanoic acid [12:0(3-OH)] at O-3 and O-3'. The heterogeneity among the three forms (I-III) of P. shigelloides O74 lipid A was attributed to the substitution of the acyl residues at N-2' and O-3' with the secondary acyls: (I) cis-9-hexadecenoic acid (9c-16:1) at N-2' and 12:0 at O-3', (II) 14:0 at N-2' and 12:0 at O-3', and (III) 12:0 at N-2' and 12:0 at O-3'. The pro-inflammatory cytokine-inducing activities of P. shigelloides O74 LPS were similar to those of Escherichia coli O55 LPS.     

Synthetic receptor analogues: the conformation of methyl 4-O-alpha-D-galactopyranosyl-beta-D-galactopyranoside (methyl beta-D-galabioside) and related derivatives, determined by n.m.r. and computational methods

The conformations of galabiose and its methyl and ethyl beta-glycosides as well as the 3-deoxy, 3-O-methyl, 3-deoxy-3-C-methyl, 3-deoxy-3-C-ethyl, and 6-deoxy analogues were investigated by n.m.r. (1H, 13C, n.O.e.) and computational (HSEA) methods. A good correlation was found between the computational data and the n.m.r. data for aqueous solutions. The conformations in aqueous solution were similar, whereas crystalline galabiose or methyl beta-D-galabioside in solution in methyl sulfoxide adopted different conformations that showed intramolecular hydrogen bonds (O-5'. . . O-3 and O-2'. . . O-6, respectively).     

A simple procedure for synthesis of diastereoisomers of thymidine cyclic 3',5'-phosphate derivatives

Diastereoisomeric thymidine cyclic (3',5')-methanephosphonates (3a), cyclic (3',5')-phosphoranilidates (3b) and cyclic (3',5')-phosphoranilidothioates (3c) were prepared by treatment of diastereoisomerically pure thymidine 3'-O-[O-(4-nitrophenyl)methanephosphonates] (2a), 3'-O-[O-(4-nitrophenyl)phosphoranilidates] (2b) or 3'-O-[O-(4-nitrophenyl)phosphoranilidothioates] (2c), respectively, with sodium hydroxide in dioxane-water solution.     

Conformational analysis of levanbiose by molecular mechanics.

A relaxed conformational energy map for levanbiose, O-beta-D-fructofuranosyl-(2----6)-beta-D-fructofuranoside, was computed with the molecular mechanics program MM2(87). All torsion angles of the three linkage bonds were driven by 30 degrees increments while two primary alcohol groups were held at three staggered forms. The steric energy of all other parameters was optimized. The side groups were retained at the same relative positions on the two rings in this first part of the study so our results are directly applicable to the study of polymeric levan with identical repeating units. The low-energy dimers did not lead to viable polymers. The interresidue linkage torsion angles defined by C-6-O-2'-C-2'-C-1' (phi) and O-5-C-5-C-6-O-2' (omega) have minima at +60 degrees and -60 degrees, respectively, with accessible minima at other staggered forms. As observed in inulobiose, the preferred torsion angle at central linkage bond defined by C-5-C-6-O-2'-C-2' (psi) was antiperiplanar. An analysis of all conformations of staggered side groups showed that the C-1 and C-1' groups had little effect but the C-6' group showed a preference for chi-6'(O-5'-C-5'-C-6'-O-6') = -60 degrees. The fructofuranose rings were started at the low-energy 4(3)T conformation (angle of pseudorotation, phi 2 = 265 degrees) that was retained except when the linkage conformations created severe inter-residue conflict.     

A crystallographic determination of a chemical structure: 6-amino-10-(beta-D-ribofuranosylamino)pyrimido-[5,4-d]pyrimidine, an example of an unusual D-ribose conformation.

The crystal and molecular structure of 6-amino-10-(beta-D-ribofuranosylamino)-pyrimido[5,4-d]pyrimidine has been determined by single crystal X-ray diffraction methods. The crystals are triclinic, of noncentric space group Pl, with cell dimensions a equals 5.434 (5), b equals 12.269 (19), c equals 4.574 (4) A, alpha equals 92.3 (1), beta equals 94.0 (1), gamma equals 95.3 degrees (1) and Z equals 1. The structure has been refined to an R value of 0.049 (Rw equals 0.063), by use of counter measured intensity data for 1063 observed reflections. The pyrimidopyrimidine ring is planar. The sugar moiety is in the envelope conformation with O-1'-endo (0E), and there is an intramolecular hydrogen bond (2.58 A) (O-3'-H-O3'...O-2'). All oxygen atoms except O-1' ring oxygen-atom are involved in hydrogen bonding. The pyrimidopyrimidine rings lie in planes 3.4 A apart.     

Structural studies of the polysaccharide antigen of Eubacterium saburreum, strain 49.

The polysaccharide antigen produced by Eubacterium saburreum, strain L 49, is composed of D-glycero-D-galacto-heptose and a new sugar, tentatively identified as 6-deoxy-D-altro-heptose. It contains chains of alternating (1 leads to 3)- and (1 leads to 6)- linked beta-D-glycero-D-galacto-heptopyranosyl residues, the latter being substituted with 6-deoxy-alpha-heptofuranosyl groups at O-3. The polysaccharide further contains 0-acetyl groups, linked to O-7 of part of the heptosyl residues and to O-2 of part of the 6-deoxyheptosyl groups.     

Crystal structure and n.m.r. analysis of lactulose trihydrate.

The 13C CPMAS n.m.r. spectrum of 4-O-beta-D-galactopyranosyl-D-fructose (lactulose) trihydrate, C12H22O11.3 H2O, identifies the isomer in the crystals as the beta-furanose. This is confirmed by a crystal structure analysis, using CuK alpha X-ray data at room temperature. The space group is P212121, with Z = 4 and cell dimensions a = 9.6251(3), b = 12.8096(3), c = 17.7563(4) A. The structure was refined to R = 0.031 and Rw 0.025 for 1929 observed structure amplitudes. All the hydrogen atoms were unambigously located on difference syntheses. The conformation of the pyranose ring is the normal 4C1 chair and that of the furanose ring is 4T3. The 1----4 linkage torsion angles are O-5'-C-1'-O-1'-C-4 = 79.9(2) degrees and C-1'-O-1'-C-4-C-5 = -170.3(2) degrees. All hydroxyls, ring and glycosidic oxygens, and water molecules are involved in the hydrogen bonding, which consists of infinite chains linked together by water molecules to form a three-dimensional network. There is a three-centered intramolecular, interresidue hydrogen bond from O-3-H to O-5' and O-6'. The n.m.r. spectrum of the amorphous, dehydrated trihydrate suggests the occurrence of a solid-state reaction forming the same isomeric mixture as was observed in crystalline anhydrous lactulose, although the mutarotation of the trihydrate when dissolved in Me2SO is very slow.     

Template controlled coupling and recombination of oligonucleotide blocks containing thiophosphoryl groups.

Oxidation of a pair of 3'- and 5'-thiophosphoryloligonucleotides in the presence of a complementary oligonucleotide template is shown to provide an effective means for selectively linking oligonucleotide blocks. Coupling proceeds rapidly and efficiently under mild conditions in dilute aqueous solutions (microM range for oligomers, 2-15 min at 0-4 degrees C with K3Fe(CN)6 or KI3 as oxidant). This chemistry was demonstrated by polymerization of a thymidylate decamer derivative (sTTTTTTTTTTs) in the presence of poly(dA) and by coupling oligomers possessing terminal thiophosphoryl groups (ACACCCAATTs + sCTGAAAATGG and ACACCCAATs + sCTGAAAATGG) in the presence of a template (CCATTTTCAGAATTGGGTGT). Efficient linking of 5' to 3' phosphoryl groups can be achieved under conditions where virtually no coupling takes place in absence of a template. A novel feature of the chemistry is that catalyzed recombinations of oligomers containing internal -OP(O)(O-)SSP(O)(O-)O- linkages can be directed by hydrogen bonding to a complementary oligonucleotide. Convenient procedures are reported for solid phase synthesis of the requisite oligonucleotide 3'- and 5'-phosphorothioates.     

A 1H-NMR and MD study of intramolecular hydrogen bonds in methyl beta-cellobioside.

The existence of an HO-3...O-5' intramolecular hydrogen bond in methyl beta-cellobioside in solution in Me2SO-d6 and H2O-CD3OD (4:1 w/w) was studied by 500-MHz 1H-NMR spectroscopy and MD simulations. Temperature coefficients for the chemical shift of the hydroxyl resonances in these solvents were determined and the rates of proton exchange in the latter solvent were obtained from NOE data. With H2O-CD3OD as the solvent, the HO-3...O-5' hydrogen bond was insignificant, but its presence in Me2SO-d6 was confirmed.     

Determining the crystal structure of cellulose III(I) by modeling

Recently, a one-chain monoclinic unit cell for cellulose III(I) having P2(1) symmetry and a single glucose in the asymmetric unit was proposed, based on high-resolution diffraction patterns. The new work challenged a two-chain structure that was published 25 years earlier, although it did not provide new three-dimensional coordinates. Our goals were to solve the structure by modeling, find whether modeling would reject the previously determined two-chain unit cell, and compare the model with the anticipated experimental structure. Combinations of three rotamers of the O-2, O-3, and O-6 hydroxyl groups produced 27 'up' and 27 'down' starting structures. Clusters ('minicrystals') of 13 cellotetraose chains terminated by methyl groups for each of the 54 starting structures were optimized with MM3(96). Hydroxyl groups on 16 of these 54 structures reoriented to give very similar hydrogen-bonding schemes in the interiors, along with the lowest energies. Hydrogen bonds included the usual intramolecular O-3H...O-5' linkage, with O-6' also accepting from O-3H. Interchain hydrogen bonds form an infinite, cooperative O-6H...O-2H...O-6 network. Direct comparison of total minicrystal energies for the one- and two-chain unit cell was inappropriate because the two-chain cell's alternate chains are shifted 0.9 A along the z-axis. To get comparable energy values, models were built with both cellotetraose and cellohexaose chains. The differences in their energies represent the energies for the central layers of cellobiose units. The one-chain cell models had much lower energy. The eight best 'up' one-chain models agree reasonably well with the structure newly determined by experiment.     

The crystal structure of the alpha-cellobiose.2 NaI.2 H(2)O complex in the context of related structures and conformational analysis

The crystal structure of beta-D-glucopyranosyl-(1-->4)-alpha-D-glucopyranose (alpha-cellobiose) in a complex with water and NaI was determined with Mo K(alpha) radiation at 150 K to R=0.027. The space group is P2(1) and unit cell dimensions are a=9.0188, b=12.2536, c=10.9016 A, beta=97.162 degrees. There are no direct hydrogen bonds among cellobiose molecules, and the usual intramolecular hydrogen bond between O-3 and O-5' is replaced by a bridge involving Na+, O-3, O-5', and O-6'. Both Na+ have sixfold coordination. One I(-) accepts six donor hydroxyl groups and three C-H***I(-) hydrogen bonds. The other accepts three hydroxyls, one Na+, and five C-H***I(-) hydrogen bonds. Linkage torsion angles phi(O-5) and psi(C-5) are -73.6 and -105.3 degrees, respectively (phi(H)=47.1 degrees and psi(H)=14.6 degrees ), probably induced by the Na+ bridge. This conformation is in a separate cluster in phi,psi space from most similar linkages. Both C-6-O-H and C-6'-O-H are gg, while the C-6'-O-H groups from molecules not in the cluster have gt conformations. Hybrid molecular mechanics/quantum mechanics calculations show <1.2 kcal/mol strain for any of the small-molecule structures. Extrapolation of the NaI cellobiose geometry to a cellulose molecule gives a left-handed helix with 2.9 residues per turn. The energy map and small-molecule crystal structures imply that cellulose helices having 2.5 and 3.0 residues per turn are left-handed.     

Synthesis of 2-(4-trifluoroacetamidophenyl)ethyl O-(L-glycero-alpha-D-manno-heptopyranosyl)-(1----7)-O-(L-glycero-alpha- D-manno- heptopyranosyl)-(1----3)-L-glycero-alpha-D-manno-heptopyranoside, corresponding to the heptose region of the Salmonella Ra core structure.

The title trisaccharide was synthesized from methyl 2,3,4-tri-O-benzyl-L-glycero-alpha-D-manno-heptopyranoside by acetolysis, followed by conversion into ethyl thioglycosides and also glycosyl bromides, which were both used in glycosylation reactions. In glycosylations using thioglycosides as glycosyl donors, N-iodosuccinimide-silver triflate and dimethyl(methylthio)sulfonium triflate were used as promoters, and in glycosylations with glycosyl bromides silver triflate was used. The protecting groups introduced into intermediates during the synthesis of the title trisaccharide were designed to allow later glycosylation at O-3' to give larger oligosaccharide fragments of the Salmonella LPS core region, and also to allow the introduction of phosphate groups at O-4 and O-4', a structural element that is suggested to be present in the Ra core.     

Interaction of D-glucose with alkaline-earth metal ions. Synthesis, spectroscopic, and structural characterization of Mg(II)- and Ca(II)-D-glucose adducts and the effect of metal-ion binding on anomeric configuration of the sugar

The interaction of D-glucose with the hydrated alkaline-earth metal halides has been studied in solution, and adducts of the type Mg(D-glucose)X2.4 H2O, Ca(D-glucose)X2.4 H2O, and Ca(D-glucose)2X2.4 H2O, where X = Cl- and Br-, have been isolated, and characterized by means of F.t.-i.r. and 1H-n.m.r. spectroscopy, X-ray powder diffraction, and molar conductivity measurements. Spectroscopic and other evidence suggested that the Mg(II) ion in the Mg(D-glucose)X2.4 H2O adducts six-coordinate, binding to a D-glucose molecule (possibly via O-1 and O-2 atoms) and to four H2O molecules, whereas, in the corresponding 1:1 Ca-D-glucose adduct, the Ca(II) ion is possibly seven-coordinate, binding to a sugar moiety (through the O-1, O-2, and other sugar donor atoms) and to four H2O molecules. In 1:2 Ca(D-glucose)2X2.4 H2O, the calcium ion may be eight-coordinate, binding to two D-glucose molecules (possibly via the O-1 and O-2 atoms of each sugar moiety) and to four H2O molecules. The strong, sugar H-bonding network is rearranged upon D-glucose adduct-formation, and the alpha-anomeric configuration is favored by these metal cation coordinations.     

Non-alkaloid Constituents of Buxus sinica (Buxaceae)

From the aerial parts of Buxus sinica, ten non- alkaloid compounds were isolated and identified as cleomiscosin A ( 1 ), 3, 5-dihydroxyl-4', 6, 7-trimethyl flavone-3′- O-β- D-glucopyranoside ( 2 ), 3, 5, 3', 4′tetrahydroxyl-3, 6, 7-trimethyl flavone (3 ) , cleomiscosin A-4′- O-β- D-gluco-pyranoside (4 ), 3, 5-dimethoxybenzoicacid-4- O-β- D-glucopyranoside (5 ), 4′, 5-dihydroxyl-3, 6, 7-trimethyl flavone (6), lupine (7), ( + )-pinoresinol- O-β- D-glucopyranoside (8 ), β-sitosterol (9), and daucosterol (10). Their structures were identified by spectral evidence.     

Prenylated flavonoids, monoterpenoid furanocoumarins and other constituents from the twigs of Dorstenia elliptica (Moraceae)

A monoprenylated flavan and two monoterpenoid substituted furanocoumarins were isolated from the twigs of Dorstenia elliptica along with 3-(3,3-dimethylallyl)-4,2',4'-trihydroxylchalcone, psoralen, bergapten, O-[3-(2,2-dimethyl-3-oxo-2H-furan-5-yl)butyl]bergaptol, beta-sitosterol and its beta-D-glucopyranoside. The structure of the flavan was determined as 6(1,1-dimethylallyl)-7,4'-dihydroxylflavan and the monoterpenoid substituted furanocoumarins were assigned as O-[3-(2,2-dimethyl-3-oxo-2H-furan-5-yl)-3-hydroxybutyl]-bergaptol and O-[2-(5-hydroxy-2,6,6-trimethyl-3-oxo-2H-pyran-2-yl)ethyl]bergaptol, respectively, using spectroscopic analysis, especially, 2D NMR spectra.     

Strain specificity of outer membrane proteins in Escherichia coli.

Outer membrane proteins of various strains of Escherichia coli were compared using three different systems of sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. The outer membranes of E. coli K-12, E. coli B, and E. coli J-5 had distinctive protein compositions. As regards proteins which interact with peptidoglycan, E. coli K-12 contained O-8 and O-9, while E. coli B possessed one protein which migrated to the position of O-9. Although E. coli J-5 possessed two such proteins, O-8' and O-9', their positions on polyacrylamide gel were different from those of O-8 and O-9. Protein O-7, which migrates slightly more slowly than O-8, was found specifically in E. coli K-12. Proteins O-10 and O-11 were found in all strains tested, although the relative amounts were different depending on the strain. Strains of E. coli K-12 and E. coli J-5 gave three major bands, O-2a, O-2b, and O-3, in the region of high molecular weight. These proteins were repressed by iron in the cultivation media. Strains of E. coli B, on the other hand, gave only O-2b and O-3. E. coli J-5 gave two other major bands in this region, but the amounts were not controlled by iron in the cultivation media.     

Reinvestigation of deoxyribonucleoside phosphorothioites: synthesis and properties of deoxyribonucleoside-3'' dimethyl phosphites.

Further investigation of the synthesis of deoxyribonucleoside-3' (t-butyl) O-(2-cyanoethyl) phosphorothioites as monomer building units for the phosphorothioite approach has led us to conclude that internucleotidic bond formation proceeded via bis(deoxyribonucleoside-3') (2-cyanoethyl) phosphite intermediates, which proved to be activated by iodine, rather than the mechanism previously reported. In connection with this study, deoxyribonucleoside-3' dimethyl phosphites were synthesized and detailed properties of them are also described.     

Structural characterization of the lipid A region of Aeromonas salmonicida subsp. salmonicida lipopolysaccharide

The lipid A components of Aeromonas salmonicida subsp. salmonicida from strains A449, 80204-1 and an in vivo rough isolate were isolated by mild acid hydrolysis of the lipopolysaccharide. Structural studies carried out by a combination of fatty acid, electrospray ionization-mass spectrometry and nuclear magnetic resonance analyses confirmed that the structure of lipid A was conserved among different isolates of A. salmonicida subsp. salmonicida. All analyzed strains contained three major lipid A molecules differing in acylation patterns corresponding to tetra-, penta- and hexaacylated lipid A species and comprising 4'-monophosphorylated beta-2-amino-2-deoxy-d-glucopyranose-(1-->6)-2-amino-2-deoxy-d-glucopyranose disaccharide, where the reducing end 2-amino-2-deoxy-d-glucose was present primarily in the alpha-pyranose form. Electrospray ionization-tandem mass spectrometry fragment pattern analysis, including investigation of the inner-ring fragmentation, allowed the localization of fatty acyl residues on the disaccharide backbone of lipid A. The tetraacylated lipid A structure containing 3-(dodecanoyloxy)tetradecanoic acid at N-2',3-hydroxytetradecanoic acid at N-2 and 3-hydroxytetradecanoic acid at O-3, respectively, was found. The pentaacyl lipid A molecule had a similar fatty acid distribution pattern and, additionally, carried 3-hydroxytetradecanoic acid at O-3'. In the hexaacylated lipid A structure, 3-hydroxytetradecanoic acid at O-3' was esterified with a secondary 9-hexadecenoic acid. Interestingly, lipid A of the in vivo rough isolate contained predominantly tetra- and pentaacylated lipid A species suggesting that the presence of the hexaacyl lipid A was associated with the smooth-form lipopolysaccharide.     

Taxines from the needles of Taxus wallichiana.

A taxine, 5 alpha O-(3'-dimethylamino-3'-phenylpropionyl) taxinine M (1) together with two known compounds 7-O-acetyltaxine A (2) and 2 alpha-acetoxy-2' beta-deacetylaustrospicatine (3) were isolated from the needles of the Himalayan yew, Taxus wallichiana Zucc. Their structures were elucidated on the basis of the NMR spectral data, ESI-MS/MS analysis and chemical methods. Compounds 1 and 3 showed moderate cytotoxic activity against the lung cancer cell line A549 in vitro.