Receptors for glucocorticoids in lung tissue

Gas chromatography-mass spectrometric identification of partially methylated aminosugars has been developed: (a) various kinds of O-methylated 2-deoxy-2-(N-methyl)-acetamidohexitols were prepared from partially O-(1-methoxy)-ethylated 2-deoxy-2-acetamidohexoses, and their gas chromatography-mass spectrometric patterns were determined; (b) permethylated glycolipids gave a satisfactory yield of 2-deoxy-2-N-methyl-amidohexoses by acetolysis with 0.5 n sulfuric acid in 95% acetic acid, followed by aqueous hydrolysis; (c) the resulting partially methylated aminosugars and neutral sugars were analyzed after borohydride reduction and acetylation according to the procedure of Lindberg and associates (Björndal, Lindberg and Svennson, 1967; Björndal, Hellerqvist, Lindberg and Svensson, 1970).This method was successfully applied to analysis of aminosugar linkages in blood group B-active ceramide pentasaccharide from rabbit erythrocytes and in Forssman antigen of equine spleen. The structure of blood group B-active glycolipid of rabbit erythrocyte was found to be Galα1 → 3Galβ1 → 4G1cNAcβ1 → 3Ga11 → 4Glc → Cer, and that of Forssman antigen to be GaNAcα1 → 3GalNAcβ1 → 3Galα1 → 4Ga11 → 4Glc → Cer.     

小刺猴头菌丝体多糖的结构研究

对小刺猴头过滤掉发酵液的发酵菌丝体,水提和碱提后获得的均一组分多糖HMP-w1.1和HMP-a1.1进行结构性质的研究。结果表明:HMP-w1.1是分子量为36.3 kD的α型吡喃糖,单糖组成为甘露糖(Man),葡萄糖(Glc),半乳糖(Gal),岩藻糖(Fuc);HMP-a1.1是分子量为42.8 kD的β型吡喃糖,单糖组成为甘露糖(Man),半乳糖醛酸(GalUA),葡萄糖(Glc),半乳糖(Gal),岩藻糖(Fuc)。综合高碘酸氧化和Smith降解的试验结果,推断HMP-w1.1的糖苷键构型可能为1→、1→4、1→4,6、1→6、1→2、1→2,6;HMP-a1.1的糖苷键构型可能为1→6、1→2、1→2,6。     

Structure of the water-insoluble α-d-glucan of streptococcus salivarius HHT

Water-insoluble, non-adherent α-d-glucans have been obtained from Streptococcus salivarius HHT under two sets of conditions: from a growing culture, or synthesized enzymically by using a glucosyltransferase. In the former case, the glucan ([α]_d + 197°) was shown by methylation analysis to have a slightly branched structure containing a relatively high proportion (80 %) of (1→3)-α-d-glucosidic linkages, together with small proportions of (1→6)- and (1→4)-α-d-glucosidic linkages. The enzymically synthesized glucan had a much less-branched structure, containing 88 % of (1→3)-α-d-glucosidic linkages. Both glucans, on Smith degradation (sequential periodate oxidation, borohydride reduction, and mild acid hydrolysis), gave linear, (1→3)-α-d-glucosidic polysaccharides (yields, 82-90%) that constitute the backbone chains. The presence of small proportions of glycerol, erythritol, 1-O-α-d-glucosyl-d-glycerol, and also 2-O-α-d-glucosyl-d-erythritol in the products of Smith degradation suggests that the short side-chains are attached to the backbone chain by (1→4)-, (1→6)-, and (1→3)-α-d-glucosidic linkages     

Depolymerization of heparin with diazomethane. Structure of N,O-methylated,even-numbered oligosaccharides produced by β-eliminative cleavage of the 2-amino-2-deoxyglycosylic linkage

The long-period reaction of heparin with excess diazomethane at 20° resulted in cleavage at the β-position of the uronic acid carboxyl group to give a mixture of methyl α- and β-glycosides of N,O-methylated di-, tetra-, and hexa-saccharides having a 4,5-unsaturated uronic acid, nonreducing end-group. The major disaccharides obtained were methyl O-(4-deoxy-3-O-methyl-α-l-threo-hex-4-enopyranosyluronic acid 2-sulfate)-(1→4)-2-deoxy-3-O-methyl-2-(N-methylsulfoamino)-α- and -β-d-glucopyranoside. The reaction of heparin at 4° yielded a mixture of methylated, higher-molecular-weight oligosaccharides, which retained some affinity for antithrombin III-Sepharose.     

Purification and methylation analysis of cell wall material from Solanum tuberosum

Cell wall material (CWM) of potatoes was prepared by sequentially extracting the wet ball-milled tissue with 1 % aq. Na deoxycholate, PhOHHOAcH₂O and 90 % (v/v) aq. DMSO. The purity of the CWM (e.g. absence of residual starch) was established by carbohydrate analysis using different acid hydrolysis conditions and by methylation studies. The partially methylated alditol acetates from the CHCl₃MeOH soluble fraction (S) of the methylated CWM were separated into 15 main peaks by GLC. Fourteen of these peaks were carbohydrate derivatives and the identity of most of these was established by MS. Reduction of the hydrolysate of S with NaBD₄ was used to identify the carbohydrate derivatives present in peaks 7 and 11 above. The occurrence of 4-linked galacturonosyl residues in the methylated polymers was established after reduction of S with LiAlH₄ and LiAlD₄. The main glycosidic linkages present in the non-cellulosic polysaccharides of the wall in descending order of concentration are: 4-linked galactose, 4-linked galacturonic acid, 5-linked arabinose and 4,6-linked glucose. The major branch points are those through 0–6 of glucose and 0–4 of rhamnose. Arabinose, galactose and xylose residues constituted the non-reducing ends. Graded acid hydrolysis of the CWM made it possible to assess the relative strengths of some of the glycosidic linkages. The general structural features of the CWM are discussed in the light of these results.     

Determination of the linkages in some methylated,sialic acid-containing,meningococcal polysaccharides by mass spectrometry

The application of gas-liquid chromatography-mass spectrometric (g.l.c.-m.s.) analysis to a number of sialic acid-containing polysaccharides of meningococcal origin has been studied. Methylation of these polysaccharides by the Hakomori conditions resulted in both O- and N-methylation. Methanolysis of the methylated polysaccharides from serogroup C [(2→9)-linked], colominic acid [(2→8)-linked], and serogroups Y and W-135 [both (1→4)-linked], yielded the respective 4,7,8,4,7,9-, and 7,8,9-tri-O-methyl derivatives of methyl N-acetyl-N-methyl-β-D-neuraminate methyl glycoside. As model compounds, methyl N-acetyl-4,7,8,9-tetra-O-methyl-α-D-neuraminate methyl glycoside and its N-methyl derivative were also synthesized. All of the methylated derivatives could be identified on the basis of their typical fragmentation-patterns, indicating that this method is applicable to the determination of the position of linkages to sialic acid residues in biopolymers.     

Structural characterization and biological activities of two α-glucans from <Emphasis Type="Italic">Radix Paeoniae Alba</Emphasis>

Radix Paeoniae Alba is widely used in Chinese traditional medicine to treat various diseases such as gastrointestinal disorders, cancer, and other diseases. In this study, two polysaccharides RPAPW1 and RPAPW2 were isolated from Radix Paeoniae Alba by DEAE-52 cellulose chromatography and G-25 sephadex. According to physicochemical methods, NMR and methylation analysis, RPAPW1 and RPAPW2 were established to be α-glucans consisting of predominant 4-linked α- Glc residues branched at O-6 and contained trace amount of protein and uronic acid. Immunological tests indicated that RPAPW1, RPAPW2 and could promote splenocyte proliferation and RAW264.7 phagocytic activity. In vitro, RPAPW1 and RPAPW2 elicited a week reducing power, DPPH scavenging activity and could not protect the PC12 cells from H₂O₂ damage. These data implied polysaccharides RPAPW1 and RPAPW2 had the potential to be a natural immunopotentiating and antioxidant supplement for preparing functional foods and nutraceuticals.     

Specific conversion of d-galactose into d-galacturonic acid residues in glycoproteins: a facile method for carbohydrate linkage-analysis

The terminal d-galactopyranosyl residues of asialoglycopeptides isolated from human α₁-acid glycoprotein were oxidized in nearly quantitative yield to the corresponding uronic acid residues by a two-step sequence employing d-galactose oxidase followed by treatment with Tollens reagent, Ag(NH₃)₂⁺. Mild acid hydrolysis of the oxidized glycopeptides led to the isolation of the corresponding aldobiuronic acid(s). Structural and colorimetric analysis revealed that only one aldobiuronic acid, 2-amino-2-deoxy-4-O-(β-d-galactopyranosyluronic acid)-d-glucose, was isolated from the oxidized glycopeptides of α₁-acid glycoprotein. This method can readily distinguish between the (1→3), (1→4), and (1→6) isomers of the corresponding aldobiuronic acids.     

Isolation and characterisation of cell wall polysaccharides from cocoa (Theobroma cacao L.) beans

 Cell wall material (CWM) was prepared from sun-dried cocoa (Theobroma cacao L.) bean cotyledons before and after fermentation. The monosaccharide composition of the CWM was identical for unfermented and fermented beans. Polysaccharides of the CWM were solubilised by sequential extraction with 0.05 M trans-1,2-diaminocyclohexane-N,N,N′,N′-tetraacetic acid (CDTA), 0.05 M Na₂CO₃, and 1 M, 4 M and 8 M KOH. The non-cellulosic sugar composition for each fraction was similar for unfermented and fermented samples, indicating that fermentation caused no significant modification of the structural features of individual cell wall polysaccharides. Pectic polysaccharides accounted for 60% of the cell wall polysaccharides but only small amounts could be solubilised in solutions of CDTA, Na₂CO₃, and 1 M and 4 M KOH. The bulk of the pectic polysaccharides were solubilised in 8 M KOH and were characterised by a rhamnogalacturonan backbone heavily substituted with side-chains of 5-linked arabinose and 4-linked galactose. Linkage analysis indicated the presence of additional acidic polysaccharides, including a xylogalacturonan and a glucuronoxylan. Cellulose, xyloglucan and a galactoglucomannan accounted for 28%, 8% and 3% of the cell wall polysaccharides, respectively. It is concluded that the types and structural features of cell wall polysaccharides in cocoa beans resemble those found in the parenchymatous tissue of many fruits and vegetables rather than those reported for many seed storage polysaccharides. Received: 29 May 1999 / Accepted: 19 October 1999     

Structural studies on the extracellular polysaccharide of the red alga, Porphyridium cruentum

Partial acid hydrolyzates of the extracellular polysaccharide from Porphyridiunm cruentum yield three disaccharides and two uronic acids. These constitute all of the uronic acid in the polymer. The novel disaccharides are 3-O-(α-$\text{D}$-glucopyranosyl- uronic acid)-$\text{L}$-galactose, 3-O-(2-O-methyl-ca-glucopyranosyluronic acid)-$\text{D}$- galactose, and 3-0-(2-0-methyl-a-$\text{D}$-glucopyranosyluronic acid)-$\text{D}$-glucose. The polyanion of high molecular weight contains $\text{D}$- and $\text{L}$-galactose, xylose, $\text{D}$-glucose, $\text{D}$-glucuronic acid and 2-O-methyl-D-glucuronic acid, and sulfate in molar ratio (relative to $\text{D}$-glucose) of 2.12:2.42:1.00:1.22:2.61. Preliminary periodate-oxidation studies suggest that the hexose and uronic acids are joined to other residues by ( 1→3) glycosidic linkages. About one-half of the xylose residues are (1→3)-linked.     

Biotransformation pathway and kinetics of the hydrolysis of the 3-O- and 20-O-multi-glucosides of PPD-type ginsenosides by ginsenosidase type I

Ginsenosidase type I from Aspergillus niger g.48 can hydrolyze the 3-O- and 20-O-multi-glycosides of PPD-type ginsenosides. The enzyme molecular weight is approximately 74 kDa. When hydrolyzing the glycosides of Rb1, Rb3, Rb2 and Rc, the structures of which only differ in their terminal 20-O-glycosides, ginsenosidase type I hydrolyzes both the 3-O- and 20-O-glycosides of Rb1 and Rb3 using two pathways, but the enzyme first hydrolyzes the 3-O-glucosides of Rb2 and Rc using one pathway. One pathway of Rb1 hydrolyzes the 20-O-Glc of Rb1 to Rd→F2→C-K; another pathway hydrolyzes the 3-O-Glc of Rb1 to Gyp17→Gyp75→C-K. Two hydrolysis pathways are used to hydrolyze the 20-O-Xyl and the 3-O-Glc of Rb3. According to the enzyme reaction parameters K_m, V_max and V₀ at a 10 mM substrate concentration, the enzyme hydrolysis velocity values decrease in the following order: the 20-O-Xyl of Rb3→Rd> the 20-O-Glc of Rb1→Rd> the 3-O-Glc of Rc> the 3-O-Glc of Rb2> the 3-O-Glc of Rd> the 3-O-Glc of Rb3→C-Mx1> the 3-O-Glc of Rb1→Gyp17> the 3-O-Glc of F2> the 3-O-Glc of 20(S)-Rg3.     

Isolation from a softwood xylan of oligosaccharides containing two 4-O-methyl-d-glucuronic acid residues

Partial hydrolysis of a larch arabino(4-O-methylglucurono)xylan afforded two series of oligouronides composed of 4-O-methyl- d-glucuronic acid and d-xylose residues. The first series included aldouronic acids up to the aldopentaouronic acid. Methylation analysis indicated that the aldopentao- and aldotetrao-uronic acids were mixtures of isomers. One aldotetraouronic acid was isolated and identified as O-β-d-Xylp-(1 → 4)-O-β-d-Xylp-(1 → 4)-O-(4-O-Me-α-d-GlcAp)-(1 → 2)-d-Xyl. The two isomeric aldotriouronic acids were separated from each other. The acids of the second series, which were composed of two uronic acids and 2-4 d-xylose residues, were identified as follows: O-β-d-Xylp-(1 → 4)-O-(4-O-Me-α-d-GlcAp)-(1 → 2)-O-β-d-Xylp-(1 → 4)-O(4-O-Me-α-d-GlcAp)-(1 → 2)-O-β-d-Xylp-(1 → 4)-d-Xyl, O-(4-O-Me-α-d-GlcAp)-(1 → 2)-O-β-d-Xylp-(1 → 4)-O-(4-O-Me-α-d-GlcAp)-(1 → 2)-O-β-d-Xylp-(1 → 4)-O-β-d -Xylp-(1 → 4)-D-Xyl, O-(4-O-Me-α-d-GlcAp)-(1 → 2)-O-β-d-Xylp-(1 → 4)-O-(4-O-Mec-α-d-GlcAp)-(1 → 2)-O-β-d-Xylp-(1 → 4)-D-Xyl, and O-(4-O-Me-α-d-GlcAp)-(1 → 2)-O-β-d-Xylp-(1 → 4)-O-(4-O-Me-α-d-GlcAp)-(1 → 2)-D-Xyl. The first three compounds were new acidic oligosaccharides. The 4-O-methyl-d-glucuronic acid in the second series was present in a larger proportion than in the first series, indicating that a large proportion of the uronic acid side-chains were located on two contiguous D-xylose residues in the backbone of the softwood xylan.     

刺五加果水溶性多糖的研究——AS-2的分离纯化与结构探讨

从刺五加果中抽提出水溶性粗多糖。经酸性乙醇分级及反复冻融得到多糖AS-2。AS-2经Sepharose CL-4B柱层析为单一对称峰,经醋酸纤维素膜电泳为一条带,冻融后高速离心无沉淀可证明其为均一级分。G.C分析表明,AS-2由Ara、Xyl、Rha、Gal、Glc组成,其单糖摩尔比为1.6:1.2:1.8:1.0:3.6。AS-2的分子量约为78kD,比旋光度[α]_D~(25)=+17°,特性粘度[η]=0.068。红外光谱分析含β型糖苷键。部分酸水解、酶解、高碘酸酸化、Smith降解、完全甲基化、G.C,G.C-M.S的分析结果表明,以β(1→3)Glc及β(1→4)Glc构成分子的主链。Glc的C_3上带有分支,约每4个己糖残基带有1个侧链。侧链上,Rha多以1→4苷键相连,部分残基C_2上有分支。Gal存在(1→6)及(1→3)连接方式,多数Glc以(1→6)苷键连结,少数Glc出现在分子非还原末端。位于分子末端的还有Ara与Xyl。     

Chemical synthesis of a (1→2)-d-glucopyranan

1,2-Anhydro-3,4,6-tri-O-benzyl-α-d-glucopyranose was polymerized with a number of Lewis acids. Phosphorus pentafluoride at ?60° caused polymerization to a product rich in β linkages. Other Lewis acids at higher temperatures gave perbenzylated polysaccharides of lower molecular weight with less stereoselectivity. Debenzylation of the most-regular derivative gave a polysaccharide whose specific rotation was +14.7° and whose ¹³C-n.m.r. spectrum had six absorptions corresponding to those of natural (1→2)-β-d-glucopyranans and additional minor peaks presumably due to some α-anomeric configurations. It was estimated to have ～90% of β linkages.     

Analytical and structural features of the gum exudate from Combretum hartmannianum schweinf.

The gum exudate from Combretum hartmannianum is water-soluble, forms very viscous solutions, and contains galactose (22%), arabinose (43%), mannose (10%), xylose (6%), rhamnose (4%), glucuronic acid (6%), 4-O-methylglucuronic acid (2%), and galacturonic acid (7%). The acidic components produced on hydrolysis of the gum were 6-O-(β-D-glucopyranosyluronic acid)-D-galactose, and two saccharides that had the same chromatographic mobility, and contained mannose and galacturonic acid, and galactose and 4-O-methylglucuronic acid, respectively. Methylation and methanolysis of the gum indicated the presence of terminal uronic acid, rhamnose, xylose, galactose, arabinofuranose, and arabinopyranose. Controlled, acid hydrolysis indicated the presence of (1→3)-linked arabinopyranose side-chains and (1→6)-linked galactose residues. C. hartmannianum gum, when subjected to two Smith-degradations, yielded Polysaccharides I and II, both of which contained galactose, arabinose, and mannose. Insufficient crude gum was available for a complete structural study, but the molecule was shown to contain long, sparsely branched chains of (1→6)-linked galactose residues, to which are attached (1→3)-linked arabinose and (1→3)-linked mannose side-chains.     

Total hemicelluloses from Hordeum vulgare plants at different stages of maturity

The changes in the composition of the total hemicelluloses of leaf and stem tissues of field-grown barley plants have been examined at different stages of maturation. In each plant the proportion of xylose residues in the total hemicellulose increases with tissue maturity, that of galactose varies little, and the proportions of arabinose, glucose and uronic acid residues decrease. The ratio of β(1 → 3) to β(1 → 4) linkages in the β-glucans decreases with tissue maturity and there is a decrease in the $\text{DP}$_n of these β-glucans.     

The Conformational Properties of the Glc3Man Unit Suggest Conformational Biasing within the Chaperone-assisted Glycoprotein Folding Pathway

A major puzzle is: are all glycoproteins routed through the ER calnexin pathway irrespective of whether this is required for their correct folding? Calnexin recognizes the terminal Glcα1-3Manα linkage, formed by trimming of the Glcα1-2Glcα1-3Glcα1-3Manα (Glc₃Man) unit in Glc₃Man₉GlcNAc₂. Different conformations of this unit have been reported. We have addressed this problem by studying the conformation of a series of N-glycans; i.e. Glc₃ManOMe, Glc₃Man₄,₅,₇GlcNAc₂ and Glc₁Man₉GlcNAc₂ using 2D NMR NOESY, ROESY, T-ROESY and residual dipolar coupling experiments in a range of solvents, along with solution molecular dynamics simulations of Glc₃ManOMe. Our results show a single conformation for the Glcα1-2Glcα and Glcα1-3Glcα linkages, and a major (65%) and a minor (30%) conformer for the Glcα1-3Manα linkage. Modeling of the binding of Glc₁Man₉GlcNAc₂ to calnexin suggests that it is the minor conformer that is recognized by calnexin. This may be one of the mechanisms for controlling the rate of recruitment of proteins into the calnexin/calreticulin chaperone system and enabling proteins that do not require such assistance for folding to bypass the system. This is the first time evidence has been presented on glycoprotein folding that suggests the process may be optimized to balance the chaperone-assisted and chaperone-independent pathways.     

Synthesis of two isomeric methyl β-D-xylotriosides containing a (1→2)-β-linkage

Partial hydrolysis with acid, methylation analysis (including uronic acid degradation), Smith degradation, and p.m.r. spectroscopy have been used to determine the primary structure of the capsular polysaccharide of Klebsiella serotype k64. The hexasaccharide repeating-unit, which also contains one O-acetyl substituent, comprises a 4)-α-d-GlcpA-(1 → 3)-α-d-Manp-(1 → 3)-β-d-Glcp-(1 → 4)-α-d-Manp-(1 → chain with a 4,6-O-(l-carboxyethylidene)-β-d-glucopyranosyl and an l-rhamnosyl group attached to the 4-linked d-mannosyl residue at O-2 and O-3, respectively.     

Structure of a heteroxylan of gum exudate of the palm Scheelea phalerata (uricuri)

The polysaccharide isolated from the gum exudate of palm Scheelea phalerata (SPN) was water-insoluble and composed of Fuc, Ara, Xyl, and uronic acid moieties in a 5:34:54:7 molar ratio: 12% of phenolics were also present. A soluble polysaccharide (SPNa) was obtained after alkaline treatment, which contained Fuc, Ara, Xyl and uronic acid in a 7:44:42:7 molar ratio, with only 2% phenolics. SPNa had an M(W) approximately 1.04 x 10(5) g mol(-1) and was almost monodisperse (M(W)/M(N) : 1.25 +/-0.22). It had a branched structure with side chains of 2-O-substituted Xylp (approximately 8%) and 3-O-substituted Araf (12%) units, and a large proportion of nonreducing end-units of Araf (15%), Fucp (10%), Xylp (4%), and Arap (6%). The (1 --> 4)-linked beta-Xylp main-chain units were 3-O- (9%), 2-O- (13%), and 2,3-di-O- (13%) substituted. Its (13)C NMR spectrum contained at least 9 C-1 signals, those at delta 108.6 and 107.7 arising from alpha-Araf units. Others were present at delta 175.4 from C-6 of alpha-GlcpA and delta 15.6 from C-6 of Fucp units. The main chain of SPNa was confirmed by analysis of a Smith-degraded polysaccharide (SPDS): methylation analysis provided a 2,3-Me(2)-Xyl (65%) derivative and its (13)C NMR spectrum showed five main signals typical of a (1 --> 4)-linked beta-Xylp units. Methylation analysis of a carboxy-reduced polysaccharide (SPN-CR) revealed a 2,3,4,6-Me(4)-Glc derivative (4%) arising from nonreducing end-units of GlcpA. Alpha-GlcpA-(1 --> 2)-alphabeta-Xy1p and alpha-GlcpA-(1 --> 2)-beta-Xylp-(1 --> 4)-alphabeta-Xylp were obtained via partial acid hydrolysis of SPN, showing the structure of side-chain substituents on O-2 of the main-chain units.     

Structural studies of sapote (Sapota achras) gum

Sapote gum contains residues of L-arabinose (pyranose and furanose), D-xylose, D-glucuronic acid, and 4-O-methyl-D-glucuronic acid in the ratio 1.0:2.8:0.48:0.52. The two uronic acids were conveniently determined by reducing the carboxyl functions with lithium borohydride and measuring the ratio of D-glucose to 4-O-methyl-D-glucose. Periodate oxidation of the carboxyl-reduced gum gave inter alia 2-O-methyl-D-erythritol and 4-O-methyl-D-glucose in amounts suggesting that 37% of the 4-O-methyl-D-glucuronic acid residues are unsubstituted in the polysaccharide. Acetolysis of the carboxyl-reduced gum gave O-α-D-glucopyranosyl-(1→2)-(4-O-β-D-xylopyranosyl)_0,1,2,-D-xylose, a hitherto undescribed series of oligosaccharides, together with 2-O-(4-O-methyl-α-D-glucopyranosyl)-D-xylose. Methylation confirmed that sapote gum has a highly branched structure, and commercial xylanases did not depolymerize the gum. An α-L-arabinofuranosidase liberated a substantial part of the arabinose residues. Sapote gum is a member of the uncommon class of plant gums having a D-xylose backbone and structurally resembles brea gum.