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1.
Hemicellulose-type polysaccharides were isolated from the pericarp of seeds of Argania spinosa (L.) Skeels fruit by sequential alkaline extractions and fractionated by precipitation. Water soluble and water insoluble fractions were obtained, purified and characterized by sugar analysis and 1H and 13C NMR spectroscopy. The water soluble fractions were assumed to be (4-O-methyl-D-glucurono)-D-xylans, with 4-O-methyl-D-glucopyranosyluronic acid groups linked to C-2 of a (1-->4)-beta-D-xylan. The 1H NMR spectrum showed that the water soluble xylans have, on average, one non-reducing terminal residue of 4-O-methyl-D-glucuronic acid for every seven xylose units. The water insoluble fractions consisted of a neutral xylan with linear (1-->4)-beta-D-xylopyranosyl units.  相似文献   

2.
Xylans were isolated from the pericarp of prickly pear seeds of Opuntia ficus-indica (OFI) by alkaline extraction, fractionated by precipitation and purified. Six fractions were obtained and characterized by sugar analysis and NMR spectroscopy. They were assumed to be (4-O-methyl-d-glucurono)-d-xylans, with 4-O-α-d-glucopyranosyluronic acid groups linked at C-2 of a (1→4)-β-d-xylan. The sugar composition and the 1H and 13C NMR spectra showed that their chemical structures were very similar, but with different proportions of d-Xyl and 4-O-Me-d-GlcA. Our results showed that, on average, the water soluble xylans have one nonreducing terminal residue of 4-O-methyl-d-glucuronic acid for every 11 to 14 xylose units, whereas in the water non-soluble xylans, xylose units can varied from 18 to 65 residues for one nonreducing terminal residue of 4-O-methyl-d-glucuronic acid.  相似文献   

3.
A beta-1,4-xylan hydrolase (xylanase A) produced by Erwinia chrysanthemi D1 isolated from corn was analyzed with respect to its secondary structure and enzymatic function. The pH and temperature optima for the enzyme were found to be pH 6.0 and 35 degrees C, with a secondary structure under those conditions that consists of approximately 10 to 15% alpha-helices. The enzyme was still active at temperatures higher than 40 degrees C and at pHs of up to 9.0. The loss of enzymatic activity at temperatures above 45 degrees C was accompanied by significant loss of secondary structure. The enzyme was most active on xylan substrates with low ratios of xylose to 4-O-methyl-D-glucuronic acid and appears to require two 4-O-methyl-D-glucuronic acid residues for substrate recognition and/or cleavage of a beta-1,4-xylosidic bond. The enzyme hydrolyzed sweetgum xylan, generating products with a 4-O-methyl-glucuronic acid-substituted xylose residue one position from the nonreducing terminus of the oligoxyloside product. No internal cleavages of the xylan backbone between substituted xylose residues were observed, giving the enzyme a unique mode of action in the hydrolysis compared to all other xylanases that have been described. Given the size of the oligoxyloside products generated by the enzyme during depolymerization of xylan substrates, the function of the enzyme may be to render substrate available for other depolymerizing enzymes instead of producing oligoxylosides for cellular metabolism and may serve to produce elicitors during the initiation of the infectious process.  相似文献   

4.
Li B  Wei XJ  Sun JL  Xu SY 《Carbohydrate research》2006,341(9):1135-1146
A fucoidan, obtained from the hot-water extract of the brown seaweed, Hizikia fusiforme, was separated into five fractions by DEAE Sepharose CL-6B and Sepharose CL-6B column chromatography. All five fractions contained predominantly fucose, mannose and galactose and also contained sulfate groups and uronic acid. The fucoidans had MWs from 25 to 950 kDa. The structure of fraction F32 was investigated by desulfation, carboxyl-group reduction, partial hydrolysis, methylation analysis and NMR spectroscopy. The results showed that the sugar composition of F32 was mainly fucose, galactose, mannose, xylose and glucuronic acid; sulfate was 21.8%, and the MW was 92.7 kDa. The core of F32 was mainly composed of alternating units of -->2)-alpha-D-Man(1--> and -->4)-beta-D-GlcA(1-->, with a minor portion of -->4)-beta-D-Gal(1--> units. The branch points were at C-3 of -->2)-Man-(1-->, C-2 of -->4)-Gal-(1--> and C-2 of -->6)-Gal-(1-->. About two-thirds of the fucose units were at the nonreducing ends, and the remainder were (1-->4)-, (1-->3)- and (1-->2)-linked. About two-thirds of xylose units were at the nonreducing ends, and the remainder were (1-->4)-linked. Most of the mannose units were (1-->2)-linked, and two-thirds of them had a branch at C-3. Galactose was mainly (1-->6)-linked. The absolute configurations of the sugar residues were alpha-D-Manp, alpha-L-Fucp, alpha-D-Xylp, beta-D-Galp and beta-D-GlcpA. Sulfate groups in F32 were at C-6 of -->2,3)-Man-(1-->, C-4 and C-6 of -->2)-Man-(1-->, C-3 of -->6)-Gal-(1-->, C-2, C-3 or C-4 of fucose, while some fucose had two sulfate groups. There were no sulfate groups in either the GlcA or xylose residues.  相似文献   

5.
Cell wall and soluble polysaccharides that reacted with Trichosporon domesticum factor III serum were isolated from the type strain of T. domesticum. The fractions contained O-acetyl groups, which contributed to the serological reactivity. The antigenic structure was characterized by chromatographic and spectroscopic methods. The polysaccharide has an alpha-(1-->3)-D-mannan backbone with hetero-oligosaccharide side chains consisting of a 2-O-substituted beta-D-glucuronic acid residue bound to O-2 of the mannose residue, beta-D-xylopyranosyl residues located in the middle of the side chain, and a nonreducing terminal alpha-L-arabinopyranosyl residue bound to 0-4 of xylose. The mannan backbone is O-acetylated at O-6 of the mannose residues.  相似文献   

6.
从猴头菌子实体中分离得到一种新型的水溶性杂多糖HEPF2,分子量大小为1.66′104Da,该多糖由岩藻糖、半乳糖和葡萄糖以1.00:3.69:5.42比例构成,同时也含有微量的3-O-甲基鼠李糖。进一步利用傅立叶变换红外光谱法、糖组成分析、甲基化分析、部分酸水解法和核磁共振法等方法进行结构鉴定,检测结果表明,该杂多糖中包含1→4、1→6结合的葡萄糖和1→6结合的半乳糖残基,连接于主链的侧链残基,包括岩藻糖残基、少数的端基葡萄糖和半乳糖残基。核磁共振法检测结果还表明,1→4结合葡萄糖为β构型,(1→6)结合半乳糖、(1→2,6)结合半乳糖和端基葡萄糖均为α构型。  相似文献   

7.
The carbohydrate binding specificity of Psathyrella velutina lectin (PVL) was thoroughly investigated by analyzing the behavior of various complex-type oligosaccharides and human milk oligosaccharides on a PVL-Affi-Gel 10 column. Basically, the lectin interacts with the nonreducing terminal beta-N-acetylglucosamine residue, but does not show any affinity for the nonreducing terminal N-acetylgalactosamine or N-acetylneuraminic acid residue. Substitution of the terminal N-acetylglucosamine residues of oligosaccharides by galactose completely abolishes their affinity to the column. GlcNAc beta 1----3Gal beta 1----4sorbitol binds to the column, but GlcNAc beta 1----6Gal beta 1----4sorbitol is only retarded in the column. The behavior of degalactosylated N-linked oligosaccharides is quite interesting. Although all degalactosylated monoantennary sugar chain isomers are retarded in the column, those with the GlcNAc beta 1----2Man group interact more strongly with the column than those with the GlcNAc beta 1----4Man group or the GlcNAc beta 1----6Man group. The degalactosylated bi- and triantennary sugar chains bind to the column, but the tetraantennary ones are only retarded in the column. These results indicated that the binding affinity is not simply determined by the number of terminal N-acetylglucosamine residues. Addition of the bisecting N-acetylglucosamine residue reduces the affinity of oligosaccharides to the column, but addition of an alpha-fucosyl residue at the C-6 position of the proximal N-acetylglucosamine residue does not affect the behavior of oligosaccharides in the column. These results indicated that the binding specificity of PVL is quite different from those of other N-acetylglucosamine-binding lectins from higher plants, which interact preferentially with the GlcNAc beta 1----4 residue.  相似文献   

8.
The structures of water-soluble birch and beech xylans, extracted from holocellulose using dimethyl sulfoxide, were determined employing 1H and 13C NMR spectroscopy together with chemical analysis. These polysaccharides were found to be O-acetyl-(4-O-methylglucurono)xylans containing one 4-O-methylglucuronic acid substituent for approximately every 15 D-xylose residues. The average degree of acetylation of the xylose residues in these polymers was 0.4. The presence of the structural element -->4)[4-O-Me-alpha-D-GlcpA-(1-->2)][3-O-Ac]-beta-D-Xylp-(1--> was demonstrated. Additional acetyl groups were present as substituents at C-2 and/or C-3 of the xylopyranosyl residues. Utilizing size-exclusion chromatography in combination with mass spectroscopy, the weight-average molar masses (and polydispersities) were shown to be 8000 (1.09) and 11,100 (1.08) for birch and beech xylan, respectively.  相似文献   

9.
Beta-D-xylosidases are hemilcellulases that hydrolyze short xylooligosaccharides into xylose units. Here, we describe the characterization and kinetic analysis of a family 43 beta-xylosidase from Geobacillus stearothermophilus T-6 (XynB3). Enzymes in this family use an inverting single-displacement mechanism with two conserved carboxylic acids, a general acid, and a general base. XynB3 was most active at 65 degrees C and pH 6.5, with clear preference to xylose-based substrates. Products analysis indicated that XynB3 is an exoglycosidase that cleaves single xylose units from the nonreducing end of xylooligomers. On the basis of sequence homology, amino acids Asp15 and Glu187 were suggested to act as the general-base and general-acid catalytic residues, respectively. Kinetic analysis with substrates bearing different leaving groups showed that, for the wild-type enzyme, the k(cat) and k(cat)/K(m) values were only marginally affected by the leaving-group reactivity, whereas for the E187G mutant, both values exhibited significantly greater dependency on the pK(a) of the leaving group. The pH-dependence activity profile of the putative general-acid mutant (E187G) revealed that the protonated catalytic residue was removed. Addition of the exogenous nucleophile azide did not affect the activities of the wild type or the E187G mutant but rescued the activity of the D15G mutant. On the basis of thin-layer chromatography and (1)H NMR analyses, xylose and not xylose azide was the only product of the accelerated reaction, suggesting that the azide ion does not attack the anomeric carbon directly but presumably activates a water molecule. Together, these results confirm the suggested catalytic role of Glu187 and Asp15 in XynB3 and provide the first unequivocal evidence regarding the exact roles of the catalytic residues in an inverting GH43 glycosidase.  相似文献   

10.
The structures of the polysaccharide chains of the LPS from Bordetella bronchiseptica strains 110H and Bp512 were analysed by NMR spectroscopy and mass spectrometry. The polysaccharides consist of alpha-(1-4)-linked 2,3-diacetamido-2,3-dideoxy-L-galacturonic acid repeating units. Polysaccharides from both strains have 2,3, 4-triamino-2,3,4-trideoxy-alpha-galacturonamide derivatives at their nonreducing ends, a monosaccharide identified for the first time in nature. The polymers from the two strains differ in the nature of the acylation of the amino groups of this monosaccharide. In the strain 110H, the residue is formylated at positions 3 and 4, and has N-formyl-L-alanyl or L-alanyl substituents at N-2. In the strain Bp512, the amino group at position 2 is acetylated, at position 3 it is formylated, and the amino group at position 4 bears a 2-methoxypropionyl substituent. The distribution of the acyl groups was determined from long range 1H-13C correlation (HMBC) NMR spectra. Measurement of the spectra under different pH conditions showed that carboxyl groups of the inner uronic acid residues of the polymeric chain are free, and that carboxyl groups of the terminal residues are amidated. These conclusions were confirmed by the results of mass spectrometric analysis.  相似文献   

11.
Hemicellulose polymers were isolated from Argania spinosa leaf cell walls by sequential extractions with alkali. The structure of the two main polymers, xylan and xyloglucan, was investigated by enzyme degradation with specific endoglycosidases followed by analysis of the resulting fragments by high performance anion exchange chromatography (HPAEC) and matrix-assisted laser desorption ionisation-time of flight mass spectrometry (MALDI-TOF MS). The results show that A. spinosa xylan is composed of a beta-(1-->4)-linked-D-xylopyranose backbone substituted with 4-O-methyl-D-glucuronic acid residues. Xyloglucan oligosaccharide subunits were generated by treatment with an endo-(1-->4)-beta-D-glucanase of the xyloglucan-rich hemicellulosic fractions. MALDI-TOF mass spectra and HPAE-PAD chromatography of the pool of endoglucanase-generated xyloglucan oligomers indicated that A. spinosa cell wall contains a XXXG-type xyloglucan. In addition to XXXG, XXFG, XLXG/XXLG, XLFG fragments previously characterised in various plants, a second group of XXXG-type fragments was detected. The primary structure of the major subunit was determined by a combination of sugar analysis, methylation analysis, post-source decay (PSD) fragment analysis of MALDI-TOF MS and 1H NMR spectroscopy. This fragment, termed XUFG, contains a novel beta-D-Xylp-(1-->2)-alpha-D-Xylp side chain linked to C-6 of the second glucose unit from the nonreducing end of the cellotetraose sequence.  相似文献   

12.
Oligomers of hyaluronic acid were prepared by digestion of hyaluronic acid from rooster combs with testicular hyaluronidase (hyaluronate 4-glycanohydrolase, EC 3.2.1.35), leech head hyaluronidase (hyaluronate 3-glycanohydrolase, EC 3.2.1.36), and with fungal hyaluronidase (hyaluronate lyase from Streptomyces hyalurolyticus). The oligomers were fractionated by gel permeation, using Sephadex G-50. Oligomers isolated after incubation of the hyaluronic acid with the testicular hyaluronidase were further modified. To prepare oligomers with N-acetylglucosamine at both ends, terminal nonreducing glucuronic acid residues were removed with beta-glucuronidase. Reducing terminal N-acetylglucosamine residues were removed by reaction under mildly alkaline conditions. The reducing terminal N-acetylglucosamine residues were also reduced with sodium borohydride to form N-acetylglucosaminitol. The potentials of the various oligosaccharides to bind to the proteoglycan from bovine nasal septum cartilage were estimated by determining their effectiveness as inhibitors of the proteoglycan-hyaluronate interaction. The present study shows that, to bind maximally to the proteoglycan, the hyaluronate oligosaccharide must be at least 10 sugar residues in length and be terminated at the nonreducing and reducing ends with a glucuronate residue and an N-acetylglucosamine residue, respectively. Sugar residues extended beyond this basic decasaccharide, do not interact with the hyaluronate binding site on the proteoglycan.  相似文献   

13.
The major water-soluble arabinoxylan fraction from rye grain, containing 4-linked β- -xylopyranosyl residues of which about 43% were substituted solely at O-3 and 7% at both O-2 and O-3 with terminal - -arabinofuranosyl units, was hydrolysed to different extents using semi-purified xylanase from Trichoderma reesei. Products were fractionated on Biogel P-2 and structurally elucidated by sugar, methylation and high-field 1H-NMR analysis. Moderate hydrolysis released arabinose, xylose, xylobiose, xylotriose and xylotetraose together with xylo-oligosaccharides (DP ≥ 4) in which one or more of the residues were substituted at O-3 with a terminal arabinose unit. The xylose residues substituted with arabinose units at both O-2 and O-3 became enriched in the remaining polymeric fraction. Extensive hydrolysis with the enzyme released arabinose, xylose and xylobiose as major products together with small amounts of two oligosaccharides and a polymeric fraction. One of the oligosaccharides was identified as xylotriose in which the non-reducing end was substituted at O-2 and O-3 with terminal arabinose units and the other as xylotetraose in which one of the interjacent residues was substituted with arabinose units in the same way. The polymeric fraction contained a main chain of 4-linked xylose residues in which 60–70% of the residues were substituted at both O-2 and O-3 with arabinose units.

The semi-purified enzyme contained xylanase and arabinosidase activities which rapidly degraded un- and mono-substituted xylose residues while the degradation of double-substituted xylose residues was much slower. The results show that the mono- and double-substituted xylose residues were present in different polymers or different regions of the same polymer.  相似文献   


14.
The synthesis of conjugates consisting of two or three mannose units interconnected by a 1,2,3-triazole linker installed by the "click" reaction is reported. These conjugates were evaluated in mycobacterial mannosyltransferase (ManT) assay. Detailed analysis of the reaction products showed that these compounds with triazole linker between sugar moieties were tolerated by the enzyme, which elongated them by one or two sugar units with α-(1→6) linkage. The effectiveness of this transfer was reduced in comparison to that observed for the acceptor analogues containing a glycosidic linkage, but still, this is the first report on such unnatural compounds serving as substrates for mycobacterial ManT. The ability of the studied compounds to function as acceptors for the ManT suggests that the relative distance and spatial orientation of acceptor octyl hydrophobic aglycone (optimal length for the ManT) and free primary C-6 hydroxy group of the nonreducing terminal mannose unit (to which glycosyl residue is transferred by the mycobacterial ManT) are important for ManT activity, but at the same time, their variations are tolerated by the enzyme in a relatively wide range.  相似文献   

15.
Cell wall and soluble polysaccharides that reacted with Trichosporon domesticum factor III serum were isolated from the type strain of T. domesticum. The fractions contained O-acetyl groups, which contributed to the serological reactivity. The antigenic structure was characterized by chromatographic and spectroscopic methods. The polysaccharide has an α-(1→3)- -mannan backbone with hetero-oligosaccharide side chains consisting of a 2-O-substituted β- -glucuronic acid residue bound to O-2 of the mannose residue, β- -xylopyranosyl residues located in the middle of the side chain, and a nonreducing terminal α- -arabinopyranosyl residue bound to O-4 of xylose. The mannan backbone is O-acetylated at O-6 of the mannose residues.  相似文献   

16.
We have previously cloned chondroitin-4-sulfotransferase (C4ST) cDNA from mouse brain. In this paper, we report cloning and characterization of GalNAc 4-sulfotransferase (GalNAc4ST), which transfers sulfate to position 4 of the nonreducing terminal GalNAc residue. The obtained cDNA contains a single open reading frame that predicts a type II transmembrane protein composed of 424 amino acid residues. Identity of the amino acid sequence between GalNAc4ST and human C4ST was 30%. When the cDNA was transfected in COS-7 cells, sulfotransferase activity toward carbonic anhydrase VI was overexpressed but no sulfotransferase activity toward chondroitin or desulfated dermatan sulfate was increased over the control. Sulfation of carbonic anhydrase VI by the recombinant GalNAc4ST occurred at position 4 of the GalNAc residue of N-linked oligosaccharides. The recombinant GalNAc4ST transferred sulfate to position 4 of GalNAc residue of p-nitrophenyl GalNAc, indicating that this sulfotransferase transfers sulfate to position 4 at the nonreducing terminal GalNAc residue. Dot blot analysis showed that the message of GalNAc4ST was expressed strongly in the human pituitary, suggesting that the cloned GalNAc4ST may be involved in the synthesis of the nonreducing terminal GalNAc 4-sulfate residues found in the N-linked oligosaccharides of pituitary glycoprotein hormones.  相似文献   

17.
Thrombomodulin (TM) is an integral membrane glycoprotein, which occurs as both a chondroitin sulfate (CS) proteoglycan (PG) form (β-TM) and a non-PG form without a CS chain (α-TM) and hence is a part-time PG. An α-TM preparation isolated from human urine contained the glycosaminoglycan linkage region tetrasaccharide GlcUAβ1-3Galβ1-3Galβ1-4xylose, and the nonreducing terminal GlcUA residue is 3-O-sulfated. Because the human natural killer-1 sulfotransferase (HNK-1ST) transfers a sulfate group from 3'-phosphoadenosine 5'-phosphosulfate to the C-3 position of the nonreducing terminal GlcUA residue in the HNK-1 antigen precursor trisaccharide, GlcUAβ1-3Galβ1-4GlcNAc, the sulfotransferase activity toward the linkage region was investigated. In fact, the activity of HNK-1ST toward the linkage region was much higher than that toward the glucuronylneolactotetraosylceramide, the precursor of the HNK-1 epitope. HNK-1ST may be responsible for regulating the sorting of α- and β-TM. Furthermore, HNK-1ST also transferred a sulfate group from 3'-phosphoadenosine 5'-phosphosulfate to the C-3 position of the nonreducing terminal GlcUA residue of a chondroitin chain. Intriguingly, the HNK-1 antibody recognized CS chains and the linkage region if they contained GlcUA(3-O-sulfate), suggesting that HNK-1ST not only synthesizes the HNK-1 epitope but may also be involved in the generation of part-time PGs.  相似文献   

18.
A substantial level of xylose was detected in the pectic polysaccharides that had been extracted from carrot (Daucus carota L.) calli and purified by gel-permeation and ion-exchange chromatography. The results of the removal of neutral sugar chains and -elimination indicated that the xylose was not included in the neutral sugar chains but was directly bound to a polygalac-turonic-acid backbone. Methylation analysis confirmed that the xylose was directly linked to galacturonic acid at position 2 or 3, as a terminal residue. The amount of xylose was positively correlated with the size of cell clusters in several lines of cultured carrot cells.Abbreviations EC embryogenic callus - 4-GalA 4-linked galacturonic acid - NC non-embryogenic callus - T-Xyl terminal xylose - 3,4-GalA 3,4-linked galacturonic acid - 2,4-GalA 2,4-linked galacturonic acid Part of this work was supported by a research grant from the Science and Technology Agency of Japan and a Grand-in-Aid from the Ministry of Education, Science and Culture, Japan. The authors are grateful to Dr. Koichi Kakegawa of the Forestry and Forest Products Research Institute for his encouragement throughout this research.  相似文献   

19.
Mature fruit of Chorisia speciosa yield an exudate (E-I) following mechanical injury. The polysaccharide contains rhamnose, arabinose, xylose, mannose, glucose, galactose and glucuronic acid in molar ratios of 20:11:1:3:2:40:23. The main chain of the structure is composed by beta-galactopyranosyl units linked (1 --> 3) and (1 --> 6) as indicated by NMR spectra and methylation data. Arabinosef and rhamnose are terminal residues. In order to compare E-I with the polysaccharides from the fruit mesocarp, the latter was submitted to different extractions. The water fraction contains rhamnose, arabinose, xylose, mannose, glucose, galactose and uronic acid in molar ratios of 18:4:1:2:3:44:28. It was treated with CTAB yielding a precipitate which was decomplexed with NaCl, giving four fractions. The fraction obtained using 0.15 M NaCl had a quantitative composition similar that of E-I.  相似文献   

20.
The behaviour of different hardwood glucuronoxylans during the kraft pulping process was investigated. Woods and pulps xylans were isolated and characterized by size exclusion chromatography, methylation (linkage) analysis and 1H NMR. Eucalyptus globulus and Eucalyptus urograndis showed xylan retention significantly higher than that of Betula pendula. The higher retention of Eucalyptus xylans was assigned to (i) their higher average molecular weight (31 against 24 KDa in B. pendula) and to (ii) the presence of O-2 substituted 4-O-methyl--d-glucuronic acid groups ([→2)-GlcpA-(1→]) with galactopyranosyl/glucopyranosyl residues belonging to fragments of galactan/glucan chains that were absent in B. pendula xylans. A significant part of uronic acids, particularly [→2)-GlcpA-(1→] units, remain in fibres until the end of pulping. The acetylation degree and distribution of acetyl groups between Xylp units, in general terms, was similar in the three types of xylans. Unexpectedly, about 20% of the acetyl groups persisted in pulps xylans till the end of pulping.  相似文献   

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