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1.
A Cahour P Debeire L Hartmann J Montreuil H van Halbeek J F Vliegenthart 《FEBS letters》1984,170(2):343-349
The carbohydrate chains of the pathological human immunoglobulins M from two patients with Waldenström's macroglobulinemia were released by hydrazinolysis. The N-acetyllactosamine-type glycans were obtained by affinity chromatography on concanavalin A and fractionated by high-voltage paper electrophoresis. The primary structure of the major compounds was elucidated on the basis of carbohydrate analysis, methylation analysis, including mass-spectrometry, and 500 MHz 1H-NMR spectroscopy. For both patients, this appeared to be a monosialyl monofucosyl biantennary structure; the compounds differed by the presence of an intersecting N-acetylglucosamine residue. 相似文献
2.
500 MHz 1H-NMR and NOE measurements of d(GGATCC) and d(GGm6ATCC) show that both oligo-nucleotides assume a B-DNA conformation at low temperature. Around the melting temperature, however, the single and double strands of the N-methylated form are in slow exchange on the NMR time scale. The preferred conformation of the adenine methyl group, cis to N1, retards base pairing and also destabilizes the double helix. 相似文献
3.
Iveta Uhliariková Mária Vršanská Barry V. McCleary Peter Biely 《Biochimica et Biophysica Acta (BBA)/General Subjects》2013
Background
Microbial degradation of acetylated plant hemicelluloses involves besides enzymes cleaving the glycosidic linkages also deacetylating enzymes. A detailed knowledge of the mode of action of these enzymes is important in view of the development of efficient bioconversion of plant materials that did not undergo alkaline pretreatment leading to hydrolysis of ester linkages.Methods
In this work deacetylation of hardwood acetylglucuronoxylan by acetylxylan esterases from Streptomyces lividans (carbohydrate esterase family 4) and Orpinomyces sp. (carbohydrate esterase family 6) was monitored by 1H-NMR spectroscopy.Results
The 1H-NMR resonances of all acetyl groups in the polysaccharide were fully assigned. The targets of both enzymes are 2- and 3-monoacetylated xylopyranosyl residues and, in the case of the Orpinomyces sp. enzyme, also the 2,3-di-O-acetylated xylopyranosyl residues. Both enzymes do not recognize as a substrate the 3-O-acetyl group on xylopyranosyl residues α-1,2-substituted with 4-O-methyl-d-glucuronic acid.Conclusions
The 1H-NMR spectroscopy approach to study positional and substrate specificity of AcXEs outlined in this work appears to be a simple way to characterize catalytic properties of enzymes belonging to various CE families.Significance
The results contribute to development of efficient and environmentally friendly procedures for enzymatic degradation of plant biomass. 相似文献4.
Bernard Overdijk Elly F J Hiensch-Goormachtig Edvard P Beem Gé J Van Steijn Leo A W Trippelvitz Jan J W Lisman Herman Van Halbeek Johanna H G M Mutsaers Johannes F G Vliegenthart 《Glycoconjugate journal》1986,3(4):339-350
Acid -galactosidase (EC3.2.1.23) was obtained from human liver in a pure monomeric state (Mr63 000). The carbohydrate content of the enzyme was established to be, 9% by weight; mannose,N-acetylglucosamine, galactose andN-acetylneuraminic acid were found to be the constituent monosaccharides. The carbohydrate structures of the enzyme were studied at the glycopeptide level by employing 500 MHz1H-NMR spectroscopy, carbohydrate composition analysis and methylation analysis involving GLCMS. Based upon the intensities of relevant signals in the1H-NMR spectrum, approximately 60% of the chains were found to be of theN-acetyllactosamine type, having the structure
The rest appeared to be of the oligomannoside type (Man5-6GlcNAc2Asn). The carbohydrate composition and methylation analysis results sustained these findings, although the calculation of the distribution based upon these techniques indicated a somewhat lower percentage ofN-acetyllactosamine type chains. There are approximately three oligosaccharide chains per molecule. These findings offer an explanation for the abnormal distribution of -galactosidase in tissues and cultured fibroblasts of patients with I-cell disease. 相似文献
5.
C H Hokke A A Bergwerff G W van Dedem J van Oostrum J P Kamerling J F Vliegenthart 《FEBS letters》1990,275(1-2):9-14
HPLC analysis of sialic acids released from recombinant variants of human tissue plasminogen activator, human chimeric plasminogen activator, human erythropoietin, and human follitropin, expressed in Chinese hamster ovary cells, demonstrates for each glycoprotein the presence of N-acetylneuraminic acid and N-glycolylneuraminic acid in a ratio of 97:3. Structural analysis by 500 MHz1H-NMR spectroscopy, of the enzymatically released N-linked carbohydrate chains of chimeric plasminogen activator and of erythropoietin, showed that alpha 2-3 linked N-glycolylneuraminic acid can occur in different N-acetyllactosamine type antennary structures. 相似文献
6.
Lorella Navazio Barbara Baldan Paola Mariani Gerrit J. Gerwig Johannes F. G. Vliegenthart 《Glycoconjugate journal》1996,13(6):977-983
Calreticulin is a multifunctional Ca2+-binding protein of the endoplasmic reticulum of most eukaryotic cells. The 56 kDa Calreticulin glycoprotein isolated from spinach (Spinacia oleracea L.) leaves was N-deglycosylated by PNGase-F digestion. The carbohydrate moiety was isolated by gel permeation chromatography and purified by high-pH anion-exchange chromatography. The fractions were investigated by 500 MHz1H-NMR spectroscopy, in combination with monosaccharide analysis and fast-atom bombardment-mass spectrometry. The following carbohydrate structure could be established as the major component (Man8GlcNAc2): Heterogeneity was demonstrated by the presence of two minor components being Man7GlcNAc2 lacking a terminal residue (D1 or D3), compared to the major component. A cross-reactivity with an antibody against the endoplasmic reticulum retention signal HDEL was also found. 相似文献
7.
Glycosylation sites and site-specific glycosylation in human Tamm- Horsfall glycoprotein 总被引:4,自引:1,他引:3
The N-glycosylation sites of human Tamm-Horsfall glycoprotein from one
healthy male donor have been characterized, based on an approach using
endoproteinase Glu-C (V-8 protease, Staphylococcus aureus ) digestion and a
combination of chromatographic techniques, automated Edman sequencing, and
fast atom bombardment mass spectrometry. Seven out of the eight potential
N-glycosylation sites, namely, Asn52, Asn56, Asn208, Asn251, Asn298,
Asn372, and Asn489, turned out to be glycosylated, and the potential
glycosylation site at Asn14, being close to the N-terminus, is not used.
The carbohydrate microheterogeneity on three of the glycosylation sites was
studied in more detail by high-pH anion-exchange chromatographic profiling
and 500 MHz1H-NMR spectroscopy. Glycosylation site Asn489 contains mainly
di- and tri-charged oligosaccharides which comprise, among others, the
GalNAc4 S (beta1-4)GlcNAc terminal sequence. Only glycosylation site Asn251
bears oligomannose-type carbohydrate chains ranging from Man5GlcNAc2to
Man8GlcNAc2, in addition to a small amount of complex- type structures.
Profiling of the carbohydrate moieties of Asn208 indicates a large
heterogeneity, similar to that established for native human Tamm-Horsfall
glycoprotein, namely, multiply charged complex-type carbohydrate
structures, terminated by sulfate groups, sialic acid residues, and/or the
Sda-determinant.
相似文献
8.
The roots of Cichorium pumilum afforded two new guaianolides, 10β-hydroxyguaia-4,13-dien-6,12-olide and the corresponding 11β,13-dihydro derivative which could be separated only after transforming the methylene lactone into the corresponding pyrazoline. The structures were elucidated by 400 MHz 1H NMR spectroscopy. The chemotaxonomic situation is discussed briefly. 相似文献
9.
J. William Lown Christopher C. Hanstock Corrine G. Lobe Christopher Bleackley 《Journal of biomolecular structure & dynamics》2013,31(6):1107-1124
Abstract The non self complementary heptadeoxyribonucleotides d(GTCGTCA) and d(TGACGAC) were synthesized by the phosphotriester method. While complete 1H-NMR assignments of the former were obtained by a combination of one and two-dimensional techniques at room temperature, extensive stacking of the latter under these conditions dictated analysis at 50°C when the lines were sharply resolved. The duplex form of the annealed strands under the conditions of the 1H-NMR experiment was established independently of the NMR evidence by 32P end labeling with T4 polynucleotide kinase followed by butt end joining using the absolute specificity of T4 ligase for double strand DNA. Analysis of the resulting ladder of polymers was performed using gel electrophoresis and autoradiography. Complete 1H-NMR assignments of the non-exchangeable protons in the self complementary heptamer was achieved. The assignments were confirmed using NOE differences, and two-dimensional COSY, and HH-INADEQUATE experiments at 400 and 500 MHz. The assignments are in accord with a conformation for the heptamer belonging to the B family of structures. 相似文献
10.
Cuvillier Olivier; Alonso Catherine; Wieruszeski Jean-Michel; Brassart Colette; Strecker Gerard; Bouquelet Stephane; Michalski Jean-Claude 《Glycobiology》1995,5(3):281-289
During a systematic study of carbohydrate material present inhuman meconium, in addition to the previously described mucins,glycolipids and free oligosaccharides, we have now characterizeda significant quantity of free glycoasparagines. These glycoasparagineshave been isolated from human meconium by a combination of ion-exchange,concanavalin A (ConA)-affinity and high-performance liquid (HPLC)chromatographies. Their structures have been established by400 MHz 1H-NMR spectroscopy. These compounds are related toN-acetyllactosaminic type structures and are based on the commoncore These glycoasparagines are probably derived from both proteaseand partial exoglycosidase hydrolysis of fetal gastrointestinalN-glycosyl proteins. Their structures are discussed in the contextof the known catabolic pathways of N-glycans glycoasparagine N-glycosyl protein catabolism meconium NMR 相似文献
11.
Thomas E. Cedel Patricia F. Cottam Michael D. Meadows Chien Ho 《Biophysical chemistry》1984,19(4):279-287
High-resolution 1 H-NMR spectroscopy at 600 MHz has been used to investigate the conformational transitions of the histidine-binding protein J of Salmonella Typhinmrium in solution as a function of pH and of l-histidine concentration. The dissociation constant for the binding of l-histidine to histidine-binding protein J increases from 6.0 × 10?8 to 5.1 × 10?7 M in going from pH 5.57 to 8.00. The conformation of this protein as observed by 1H-NMR also changes over this range of pH. However, when l-histidine is bound, the changes in conformation with pH are much smaller. Also, the pk for the single histidyl residue in histidine-binding protein J changes from 6.75 in the absence of l-histidine to 6.52 when l-histidine is bound. Earlier work in this laboratory resulted in the identification of several proton resonances believed to be at or near the l-histidine-binding site. Two of these resonances have been assigned to a tyrosine and the single histidyl residue in the histidine-binding protein J molecule. 相似文献
12.
C. A.G. Haasnoot U. K. Pandit C. Kruk C. W. Hilbers 《Journal of biomolecular structure & dynamics》2013,31(2):449-467
Abstract 1H-NMR spectra of bleomycin A2 recorded at 500 MHz in D2O and H2O at 24°C and 3°C were investigated. Resonances of the individual spin systems were identified by using two-dimensional correlated spectroscopy (COSY), two-dimensional spin echo correlated spectroscopy (SECSY) and by the application of two-dimensional Nuclear Overhauser Effect spectroscopy (NOESY). Employment of these techniques allowed the assignment of 13 exchangeable and 59 non-exchangeable protons in the 1H NMR spectrum of bleomycin A2. By means of 2D NOE spectroscopy also interresidual connectivities could be observed. Comparison of the NOESY spectra at 3°C and 24°C suggest that at low temperatures the central part of the bleomycin A2 molecule tends to adopt an extended conformation. 相似文献
13.
C. A.G. Haasnoot H. P. Westerink G. A. van der Marel J. H. van Boom 《Journal of biomolecular structure & dynamics》2013,31(1):131-149
Abstract The double helical structure of the self-complementary DNA-RNA-DNA hybrid d(CG)r(CG) d(CG) was studied in solution by 500 MHz 1H-NMR spectroscopy. The non-exchangeable base protons and the (deoxy)ribose H1′, H2′ and H2″ protons were unambiguously assigned using 2D-J-correlated (COSY) and 2D-NOE (NOESY) spectroscopy techniques. A general strategy for the sequential assignment of 1H-NMR spectra of (double) helical DNA and RNA fragments by means of 2D-NMR methods is presented. Conformational analysis of the sugar rings of d(CG)r(CG)d(CG) at 300 K shows that the central ribonucleotide part of the helix adopts an A-type double helical conformation. The 5′- and 3′-terminal deoxyribose base pairs, however, take up the normal DNA-type conformation. The A-to-B transition in this molecule involves only one (deoxyribose) base pair. It is shown that this A-to-B conformational transition can only be accomodated by two specific sugar pucker combinations for the junction base pair, i.e. N·S (C3′-endo-C2′-endo, 60%, where the pucker given first is that assigned to the junction nucleotide residue of the strand running 5′ → 3′ from A-RNA to B-DNA) and S·S (C2′-endo-C2′-endo, 40%). 相似文献
14.
S Hase S Koyama H Daiyasu H Takemoto S Hara Y Kobayashi Y Kyogoku T Ikenaka 《Journal of biochemistry》1986,100(1):1-10
An asparagine-linked sugar chain of a protease inhibitor from barbados pride (Caesalpinia pulcherrima Sw.) was liberated by hydrazinolysis. After N-acetylation, the reducing end residue of this carbohydrate unit was coupled with 2-aminopyridine and the pyridylamino (PA-) derivative was purified by gel-filtration and reversed-phase HPLC. The structure of the resulting PA-sugar chain was determined mainly by stepwise exoglycosidase digestions and 500 MHz 1H-NMR spectroscopy and proved to be as follows: (formula; see text). 相似文献
15.
The three-dimensional structure of native SHL-I, a lectin from the venom of the Chinese bird spider Selenocosmia huwena, has been determined from two-dimensional 1H NMR spectroscopy recorded at 500 and 600 MHz. The best 10 structures have NOE violation <0.3 Å, dihedral violation <2 deg, and average root-mean-square differences of 0.85 + 0.06 Å over backbone atoms. The structure consists of a three-stranded antiparallel β-sheet and three turns. The three disulfide bridges and three-stranded antiparallel β-sheet form a inhibitor cystine knot motif which is adopted by several other small proteins, such as huwentoxin-I, ω-conotoxin, and gurmarin. The C-terminal fragment from Leu28 to Trp32 adopts two sets of conformations corresponding to the cis and trans conformations of Pro31. The structure of SHL-I also has high similarity with that of the N-terminus of hevein, a lectin from rubber-tree latex. 相似文献
16.
Determination of the structure of the carbohydrate chains of prostaglandin endoperoxide synthase from sheep 总被引:2,自引:0,他引:2
J H Mutsaers H van Halbeek J P Kamerling J F Vliegenthart 《European journal of biochemistry》1985,147(3):569-574
Prostaglandin endoperoxide synthase was isolated from sheep seminal vesicles. Sugar analysis of the glycoprotein revealed the presence of mannose and N-acetylglucosamine only. The carbohydrate moiety was released from the polypeptide backbone by hydrazinolysis. After re-N-acetylation and reduction, the resulting mixture of oligosaccharide-alditols was fractionated on Bio-Gel P-4 and their structures were investigated by 500-MHz1H-NMR spectroscopy. The carbohydrate chains turned out to be of the oligomannoside type containing six to nine mannose residues. The largest and most abundant compound was established to be: (formula; see text) For the smaller structures heterogeneity occurs with respect to the outer alpha(1----2)-linked mannose residues. Furthermore, a small amount of Man6GlcNAc-ol (artefact of the hydrazinolysis procedure) was detected by 1H-NMR spectroscopy and fast atom bombardment mass spectrometry. 相似文献
17.
《Bioscience, biotechnology, and biochemistry》2013,77(11):1866-1871
The structures of N-linked sugar chains (N-glycans) of storage glycoproteins in soybean seeds have been identified. Eight pyridylaminated (PA-) N-linked sugar chains were derived and purified from hydrazinolysates of the storage glycoproteins by reverse-phase HPLC and size-fractionation HPLC. The structures of the PA-sugar chains purified were first identified by two-dimensional PA-sugar chain mapping and ion-spray mass analysis, considering the results of sugar composition analysis or sequential exoglycosidase digestion. The deduced structures were further analyzed by ion-spray tandem mass spectrometry and 500 MHz 1H-NMR spectrometry. The eight structures fell into two categories; the major class (96.6%) was a typical high mannose-type, the minor class was a xylose containing-type (Man3Xyl1GlcNAc2, Man3Fuc1Xyl1GlcNAc2; 3.4%). 相似文献
18.
Heteroassociation of antibacterial antibiotic norfloxacin with aromatic vitamins nicotinamide and flavin mononucleotide in aqueous solution was studied by 1H NMR spectroscopy (500 MHz). Equilibrium constants, induced proton chemical shifts, and thermodynamic parameters (ΔH, ΔS) for the reactions of heteroassociation of the molecules were determined on the basis of the concentration and temperature dependences of proton chemical shifts for interacting aromatic molecules. The analysis of the results obtained indicates the formation of heterocomplexes between vitamin molecules and norfloxacin owing to stacking interactions between aromatic chromophores and additional intermolecular hydrogen bonding in norfloxacin-nicotinamide. The most probable spatial structures of 1:1 norfloxacin-flavin mononucleotide and norfloxacin-nicotinamide heterocomplexes were determined by molecular modeling methods using X-PLOR software on the basis of analysis of induced proton chemical shifts. 相似文献
19.
《Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology》1987,911(2):244-251
Acid α-glucosidase (α-d-glucoside glucohydrolase, EC 3.2.1.20) from human placenta (70 and 76 kDa) was found to contain 4 N-glycosidic carbohydrate chains per molecule. Sugar analysis of purified enzyme revealed the presence of mannose, N-acetylglucosamine and fucose at a molar ratio of 5.0:2.0:0.6. In addition, trace amounts of galactose and N-acetylneuraminic acid were detected. The sugar chains were liberated from the polypeptides by the hydrazinolysis procedure and subsequently fractionated by gel filtration and HPLC. Purified compounds were investigated by 500-MHz 1H-NMR spectroscopy. Oligomannoside-type chains of intermediate size, e.g., Man5GlcNAcGlcNAc-ol and Man7GlcNAcGlcNAc-ol, and N-type chains of smaller size e.g., Man2–3GlcNAc[Fuc]0–1GlcNAc-ol, were demonstrated to be present at a ratio of 2:3. In addition, a small amount of sialylated N-acetyllactosamine-type chains has been found. The possible biosynthetic route of the fucose-containing small-size chains is discussed. 相似文献
20.
Structure determination of the major asparagine-linked sugar chain of human factor VIII--von Willebrand factor 总被引:4,自引:0,他引:4
P Debeire J Montreuil B Samor C Mazurier M Goudemand H van Halbeek J F Vliegenthart 《FEBS letters》1983,151(1):22-26
N-glycosidically-linked glycans released by hydrazinolysis of human factor VIII/von Willebrand factor (FVIII/vWf) were separated by high-voltage electrophoresis. Five fractions were obtained, one of them representing 60% of the total amount of the N-glycosidically-linked glycans of FVIII/vWf. On the basis of the carbohydrate composition, methylation analysis and 500 MHz 1H-NMR spectroscopy, we describe the primary structure of this major glycan which is of the monosialylated and monofucosylated biantennary N-acetyllactosaminic type. 相似文献