RESUMO
Transaldolase deficiency predisposes to chronic liver disease progressing from cirrhosis to hepatocellular carcinoma (HCC). Transition from cirrhosis to hepatocarcinogenesis depends on mitochondrial oxidative stress, as controlled by cytosolic aldose metabolism through the pentose phosphate pathway (PPP). Progression to HCC is critically dependent on NADPH depletion and polyol buildup by aldose reductase (AR), while this enzyme protects from carbon trapping in the PPP and growth restriction in TAL deficiency. Although AR inactivation blocked susceptibility to hepatocarcinogenesis, it enhanced growth restriction, carbon trapping in the non-oxidative branch of the PPP and failed to reverse the depletion of glucose 6-phosphate (G6P) and liver cirrhosis. Here, we show that inactivation of the TAL-AR axis results in metabolic stress characterized by reduced mitophagy, enhanced overall autophagy, activation of the mechanistic target of rapamycin (mTOR), diminished glycosylation and secretion of paraoxonase 1 (PON1), production of antiphospholipid autoantibodies (aPL), loss of CD161+ NK cells, and expansion of CD38+ Ito cells, which are responsive to treatment with rapamycin in vivo. The present study thus identifies glycosylation and secretion of PON1 and aPL production as mTOR-dependent regulatory checkpoints of autoimmunity underlying liver cirrhosis in TAL deficiency.
RESUMO
Exopolysaccharides (EPS) produced by some lactic acid bacteria are often used by the dairy industry to improve the rheological and physical properties of yogurt, but the relationship between their structure and functional effect is still unclear. The EPS from different species, or different strains from the same species, may differ in terms of molar mass, repeating unit structure, and EPS yield during fermentation of milk. This study aimed to characterize the detailed properties of EPS produced from 7 strains of Streptococcus thermophilus, which is one of the key cultures used for yogurt manufacture. Milk was fermented with strains DGCC 7698, DGCC 7710, DGCC 7785, ST-10255y, St-143, STCth-9204, and ST4239. These strains were selected because they have been used in previous studies on yogurt texture, but a complete description of their EPS structural properties has not yet been reported. All strains were fermented under a similar acidification rate by adjusting the level of supplementation with peptone or the inoculation level, which allowed for a comparison of EPS yields under similar growth conditions (reconstituted skim milk at 40°C). The EPS from each strain was isolated and the weight-average molar mass and z-average root mean square radius determined using size-exclusion chromatography multiangle laser light scattering. The monosaccharide composition of EPS was determined using gas chromatography-mass spectrometry, and repeating unit structure was determined using nuclear magnetic resonance spectroscopy. The weight-average molar mass values of EPS ranged from 0.14 to 1.61 × 106 g/mol. All 7 EPS samples were uncharged. The strains ST-10255y and ST4239 had EPS with the same repeating unit structure. The monosaccharide compositions of the various EPS were mainly composed of glucose and galactose, with low levels of rhamnose in the EPS isolated from DGCC 7698, and N-acetylgalactosamine in the EPS from DGCC 7785, ST-10255y, and ST4239. The yields of EPS (measured when fermented milks reached pH 4.6) ranged from 8.0 to 76.4 mg of glucose equivalents/kg. In addition to (free) EPS, some strains were also able to produce capsular polysaccharide (associated with the bacterial cells) when observed with negative staining technique. The results of our study will help the dairy industry to better understand the mechanism by which different strains of Streptococcus thermophilus affect yogurt texture.
Assuntos
Polissacarídeos Bacterianos/metabolismo , Streptococcus thermophilus/metabolismo , Animais , Fermentação , Cromatografia Gasosa-Espectrometria de Massas , Streptococcus/metabolismo , IogurteRESUMO
In this study, we show that Salmonella produces an O-antigen capsule coregulated with the fimbria- and cellulose-associated extracellular matrix. Structural analysis of purified Salmonella extracellular polysaccharides yielded predominantly a repeating oligosaccharide unit similar to that of Salmonella enterica serovar Enteritidis lipopolysaccharide O antigen with some modifications. Putative carbohydrate transport and regulatory operons important for capsule assembly and translocation, designated yihU-yshA and yihVW, were identified by screening a random transposon library with immune serum generated to the capsule. The absence of capsule was confirmed by generating various isogenic Deltayih mutants, where yihQ and yihO were shown to be important in capsule assembly and translocation. Luciferase-based expression studies showed that AgfD regulates the yih operons in coordination with extracellular matrix genes coding for thin aggregative fimbriae and cellulose. Although the capsule did not appear to be important for multicellular behavior, we demonstrate that it was important for survival during desiccation stress. Since the yih genes are conserved in salmonellae and the O-antigen capsule was important for environmental persistence, the formation of this surface structure may represent a conserved survival strategy.
Assuntos
Cápsulas Bacterianas/genética , Proteínas de Bactérias/genética , Matriz Extracelular/genética , Regulação Bacteriana da Expressão Gênica , Salmonella/fisiologia , Fatores de Transcrição/genética , Cápsulas Bacterianas/metabolismo , Transporte Biológico , Metabolismo dos Carboidratos , Carboidratos/genética , Celulose/genética , Elementos de DNA Transponíveis , Dessecação , Microbiologia Ambiental , Matriz Extracelular/metabolismo , Fímbrias Bacterianas/genética , Genes Bacterianos/genética , Genes Bacterianos/fisiologia , Mutação , Antígenos O/química , Óperon/genética , Óperon/fisiologia , Salmonella/genética , Salmonella/metabolismoRESUMO
Polysaccharide from Tinospora cordifolia was isolated, purified, methylated, hydrolyzed, reduced and acetylated. The partially methylated alditol acetate (PMAA) derivative thus obtained was subjected to GC-MS studies. The following types of linkages were noticed: terminal-glucose, 4-xylose, 4-glucose, 4,6-glucose and 2,3,4,6-glucose.
Assuntos
Glicosídeos/química , Polissacarídeos/química , Tinospora/química , Acetilação , Sequência de Carboidratos , Cromatografia Gasosa-Espectrometria de Massas , Hidrólise , Indicadores e Reagentes , Metilação , Dados de Sequência Molecular , Monossacarídeos/química , OxirreduçãoRESUMO
The coccolithophores are valuable models for the design and synthesis of composite materials, because the cellular machinery controlling the nucleation, growth, and patterning of their calcitic scales (coccoliths) can be examined genetically. The coccoliths are formed within the Golgi complex and are the major CaCO(3) component in limestone sediments-particularly those of the Cretaceous period. In this study, we describe mutants lacking a sulfated galacturonomannan and show that this polysaccharide in conjunction with the Golgi-derived membrane is directly linked to the growth and shaping of coccolith calcite but not to the initial orientated nucleation of the mineral phase.
Assuntos
Calcificação Fisiológica/fisiologia , Carbonato de Cálcio/química , Membrana Celular/metabolismo , Complexo de Golgi/metabolismo , Mananas/química , Fitoplâncton/metabolismo , Divisão Celular , Cristalografia por Raios X , Microscopia Eletrônica , Mutação , FenótipoRESUMO
Bacillus thuringiensis Cry1Ac insecticidal toxin binds specifically to 120kDa aminopeptidase N (APN) (EC 3.4.11.2) in the epithelial brush border membrane of Manduca sexta midguts. The isolated 120-kDa APN is a member of a functional Cry1 toxin receptor complex (FEBS Lett. 412 (1997) 270). The 120-kDa form is glycosyl-phosphatidylinositol (GPI) anchored and converted to a 115-kDa form upon membrane solubilization. The 115-kDa APN also binds Cry1A toxins and Cry1Ac binding is inhibited by N-acetylgalactosamine (GalNAc). Here we determined the monosaccharide composition of APN. APN is 4.2mol% carbohydrate and contains GalNAc, a residue involved in Cry1Ac interaction. APN remained associated with non-covalently bound lipids through anion-exchange column purification. Most associated lipids were separated from APN by hydrophobic interaction chromatography yielding a lipid aggregate. Chemical analyses of the lipid aggregate separated from APN revealed neutral lipids consisting mostly of diacylglycerol and free fatty acids. The fatty acids were long, unsaturated chains ranging from C:14 to C:22. To test the effect of APN-associated lipids on Cry1Ac function, the lipid aggregate and 115-kDa APN were reconstituted into phosphatidylcholine (PC) vesicles. The lipid aggregate increased the amount of Cry1Ac binding, but binding due to the lipid aggregate was not saturable. In contrast the lipid aggregate promoted Cry1Ac-induced release of 86Rb(+) at the lowest Cry1Ac concentration (50nM) tested. The predominant neutral lipid component extracted from the lipid aggregate promoted Cry1Ac-induced 86Rb(+) release from membrane vesicles in the presence of APN.
Assuntos
Bacillus thuringiensis/metabolismo , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/metabolismo , Antígenos CD13/metabolismo , Metabolismo dos Carboidratos , Endotoxinas/metabolismo , Metabolismo dos Lipídeos , Manduca/metabolismo , Animais , Toxinas de Bacillus thuringiensis , Antígenos CD13/isolamento & purificação , Cromatografia em Gel/métodos , Cromatografia em Camada Fina/métodos , Cromatografia Gasosa-Espectrometria de Massas/métodos , Proteínas Hemolisinas , Marcação por Isótopo , Lipídeos/isolamento & purificação , Lipossomos/metabolismo , Fosfatidilcolinas/metabolismo , RubídioRESUMO
Structures of five water-soluble xylans have been determined. Four purified xylanase enzymes have been studied for the hydrolysis of the xylans. Different xylanases have different activities against various xylan structures. The key factors that influence the rate of xylan hydrolysis are chain length and degree of substitution. Two family 11 xylanases, Orpinomyces pc2 xylanase and Trichoderma longibrachiatum xylanase, can rapidly hydrolyze xylans that have a chain length greater than 8 xylose residues, and their hydrolytic rates are not sensitive to substituents on the xylan backbone. A family 11 xylanase from Aureobasidium pullulans is most effective on xylans that have a long chain (greater than 19 xylose residues), and also is effective against substituent groups. Although Thermatoga maritima xylanase is also more active on a long xylan chain (greater than 19 xylose residues), its hydrolytic rate is greatly reduced by substituents on xylan backbones.
RESUMO
A specific and highly potent inhibitor of diguanylate cyclase, the key regulatory enzyme of the cellulose synthesizing apparatus in the bacterium Acetobacter xylinum, was isolated from extracts of etiolated pea shoots (Pisum sativum). The inhibitor has been purified by a multistep procedure, and sufficient amounts of highly purified compound (3-8 mg) for spectral analysis were obtained. The structure of this compound was established as 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1--> 2)-beta-D-glucuronopyranosyl soyasapogenol B 22-O-alpha-D-glucopyranoside. The structure was elucidated on the basis of susceptibility to various enzymes, chemical and spectral methods, such as GC-MS, FAB-MS, and the following types of 2D-NMR: COSY, ROESY, TOCSEY, HMQC, HMBC analyses. An identical or a very similar compound with identical biological activity was also isolated from A. xylinum, strongly suggesting that at least certain aspects of cellulose synthesis in the bacteria and in higher plants may be regulated in a similar manner. The content of this saponin in etiolated plants was about 0.04 mumol (g fresh tissue)-1.
Assuntos
Inibidores Enzimáticos/química , Gluconacetobacter xylinus/enzimologia , Fósforo-Oxigênio Liases/antagonistas & inibidores , Pisum sativum/química , Saponinas/química , Triterpenos/química , Configuração de Carboidratos , Sequência de Carboidratos , Escuridão , Inibidores Enzimáticos/isolamento & purificação , Inibidores Enzimáticos/farmacologia , Proteínas de Escherichia coli , Cromatografia Gasosa-Espectrometria de Massas , Dados de Sequência Molecular , Estrutura Molecular , Ressonância Magnética Nuclear Biomolecular , Saponinas/isolamento & purificação , Saponinas/farmacologia , Espectrometria de Massas de Bombardeamento Rápido de Átomos , Triterpenos/isolamento & purificação , Triterpenos/farmacologiaRESUMO
The anti-tuberculosis drug, ethambutol (Emb), was previously shown to inhibit the synthesis of arabinans of both the cell wall arabinogalactan (AG) and lipoarabinomannan (LAM) of Mycobacterium tuberculosis and other mycobacteria. However, an Emb-resistant mutant, isolated by consecutive passage of the Mycobacterium smegmatis parent strain in media containing increasing concentrations of Emb, while synthesizing a normal version of AG, produced truncated forms of LAM when maintained on 10 microg/ml Emb (Mikusová, K., Slayden, R. A., Besra, G. S., and Brennan, P. J. (1995) Antimicrob. Agents Chemother. 39, 2482-2489). We have now isolated and characterized the truncated LAMs made by both the resistant mutant and a recombinant strain transfected with a plasmid containing the emb region from Mycobacterium avium which encodes for Emb resistance. By chemical analysis, endoarabinanase digestion, high pH anion exchange chromatography, and mass spectrometry analyses, truncation was demonstrated as primarily a consequence of selective and partial inhibition of the synthesis of the linear arabinan terminal motif, which constitutes a substantial portion of the arabinan termini in LAM but not of AG. However, at higher concentrations, Emb also affected the general biosynthesis of arabinan destined for both AG and LAM, resulting in severely truncated LAM as well as AG with a reduced Ara:Gal ratio. The results suggested that Emb exerts its antimycobacterial effect by inhibiting an array of arabinosyltransferases involved in the biosynthesis of arabinans unique to the mycobacterial cell wall. It was further concluded that the uniquely branched terminal Ara6 motif common to both AG and LAM is an essential structural entity for a functional cell wall and, consequently, that the biosynthetic machinery responsible for its synthesis is the effective target of Emb in its role as a potent anti-tuberculosis drug.
Assuntos
Antígenos de Bactérias/química , Etambutol/farmacologia , Lipopolissacarídeos/química , Mycobacterium/efeitos dos fármacos , Polissacarídeos/biossíntese , Antígenos de Bactérias/biossíntese , Resistência Microbiana a Medicamentos , Eletroforese em Gel de Poliacrilamida , Lipopolissacarídeos/biossíntese , Espectroscopia de Ressonância Magnética , Modelos Químicos , Espectrometria de Massas de Bombardeamento Rápido de Átomos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
Rhamnogalacturonan I (RG-I), a major pectic component of the primary walls of plant cells, is believed to play an important role in determining both the structure and functions of the walls. A more detailed structural description of RG-I is likely to lead to a greater understanding of the biological roles of this polysaccharide. Two enzymes secreted by Aspergillus aculeatus that have been cloned and expressed in a fungal system (Kofod et al., J. Biol. Chem., 269, 29182-29189, 1994) cleave the RG-I backbone in an endo fashion and should assist in the further structural characterization of this polysaccharide. We found that both of the available preparations of the cloned enzymes were contaminated with exoglycanases, reducing their utility in structurally characterizing RG-I. We purified the enzymes to apparent homogeneity by ion-exchange chromatography and then used the purified enzymes to generate backbone oligosaccharide fragments from partially debranched sycamore RG-I. The backbone oligosaccharides, which were separated from larger pieces of partially debranched RG-I by gel-permeation chromatography, have been structurally characterized by 1H-NMR spectroscopy, electrospray MS, GC-MS, high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and UV spectroscopy. The results of these analyses establish that rhamnogalacturonase A (RGase A) is an endohydrolase that cleaves the -4)-alpha-D-GalpA-(1-2)-alpha-L-Rhap glycosidic linkage. However, the purported rhamnogalacturonase B (RGase B) is, in fact, an endolyase that cleaves the -2)-alpha-L-Rhap-(1-4)-alpha-D-GalpA glycosidic linkage, thereby generating oligosaccharides terminating at the non-reducing end with a hex-4-enopyranosyluronic acid residue.
Assuntos
Parede Celular/química , Pectinas/química , Árvores/química , Sequência de Carboidratos , Células Cultivadas , Cromatografia Líquida de Alta Pressão , Glicosídeo Hidrolases/metabolismo , Espectroscopia de Ressonância Magnética , Dados de Sequência Molecular , Pectinas/metabolismo , Polissacarídeo-Liases/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
The N-terminal glycopeptide of pro-opiomelanocortin (POMC), designated as the 16K fragment, is highly conserved throughout vertebrates from amphibians to mammals and is likely therefore to have an important functional role. In this paper, we report the first structural characterization of N-glycans attached to asparagine-65 of a 16K glycopeptide. The 16K fragment was isolated from bovine pituitaries and the N-glycans were analysed using fast atom bombardment mass spectrometry together with sugar and linkage analysis. Sulphated-N-acetylgalactosamine-capped antennae, typical of the pituitary glycohormones, were present in the major acidic components. The POMC oligosaccharides are distinct from those of the pituitary glycohormones because the sulphate is exclusively located on the 3-arm of biantennary structures and, in addition, a significant proportion of the molecules carry the Lewis x epitope. It is probable that these differences reflect the absence of a tripeptide motif in POMC which fully conforms to the criteria previously defined for the recognition sequence for the N-acetylgalactosamine transferase that is specific for the pituitary glycohormones [Smith and Baenziger (1992) Proc. Natl. Acad. Sci. USA, 89, 329-333]. It remains to be seen whether the Lewis x epitope is involved in selectin-mediated events, but previous studies suggest that the sulphated moieties are unlikely to play a major role in clearance. The Lewis x epitope is also present in the neutral N-linked oligosaccharides, together with a variety of other antennae including a rarely found fucosylated GalNAc-GlcNAc structure.
Assuntos
Glicopeptídeos/química , Antígenos do Grupo Sanguíneo de Lewis/química , Pró-Opiomelanocortina/química , Acetilgalactosamina/análise , Sequência de Aminoácidos , Animais , Configuração de Carboidratos , Sequência de Carboidratos , Bovinos , Sequência Conservada , Epitopos/análise , Epitopos/química , Fucose/análise , Indicadores e Reagentes , Dados de Sequência Molecular , Oligopeptídeos/química , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/isolamento & purificação , Pró-Opiomelanocortina/genética , Homologia de Sequência de Aminoácidos , Serina Endopeptidases , Espectrometria de Massas de Bombardeamento Rápido de Átomos , Ácidos Sulfúricos/análise , TripsinaRESUMO
Rat C-reactive protein (CRP) is a pentameric glycoprotein composed of five apparently identical monomers, two of which form a disulfide-linked dimer (Rasosouli, M., Sambasivam, H., Azadi, P., Dell, A., Morris, H. R., Nagpurkar, A., Mookerjea, S., and Murray, R. K. (1992) J. Biol. Chem. 267, 2947-2954). In this study, the nature of the oligosaccharide chain of rat CRP was investigated by fast atom bombardment-mass spectrometry (FAB-MS), and general features of its biosynthetic pathway were also analyzed. FAB-MS, electrospray-mass spectrometry, and linkage analysis demonstrated that each monomer of rat CRP contained one oligosaccharide chain, predominantly a disialylated biantennary structure, attached to Asn-128. The biosynthesis of rat CRP was studied by immunoprecipitation of CRP synthesized in vitro and by cultured hepatocytes. The results revealed that each monomer of rat CRP was synthesized individually as a single-chain precursor with a cleavable signal sequence. The translocated species was sensitive to digestion by endoglycosidase H (endo H), indicating that it possessed a high mannose oligosaccharide. Rat CRP acquired the ability to bind to phosphorylcholine-Sepharose and to form the dimeric and oligomeric species prior to acquiring resistance to endo H. Studies using tunicamycin revealed that the N-linked oligosaccharide present in rat CRP was not required for formation of its dimeric component, oligomerization, ability to bind to phosphorylcholine, or secretion. The non-glycosylated rat CRP, however, was still able to bind to phosphorylcholine-Sepharose and to be secreted by hepatocytes.
Assuntos
Proteína C-Reativa/biossíntese , Proteína C-Reativa/química , Glicoproteínas/biossíntese , Glicoproteínas/química , Animais , Asparagina , Proteína C-Reativa/genética , Configuração de Carboidratos , Sequência de Carboidratos , Sistema Livre de Células , Eletroforese em Gel de Poliacrilamida , Substâncias Macromoleculares , Dados de Sequência Molecular , Peso Molecular , Oligossacarídeos/síntese química , Oligossacarídeos/isolamento & purificação , Biossíntese de Proteínas , Processamento de Proteína Pós-Traducional , Coelhos , Ratos , Reticulócitos/metabolismo , Ácidos Siálicos/análise , Espectrometria de Massas de Bombardeamento Rápido de ÁtomosRESUMO
Rat C-reactive protein (CRP) is unique among mammalian CRPs in being a glycoprotein and in containing a covalently linked dimer in its pentameric structure. To investigate these features, cDNA clones encoding rat CRP were isolated from an expression library, and the primary structure of the protein was derived. Taken along with the results of Northern blotting, we conclude that a single mRNA of approximately 2,500 nucleotides codes for a precursor of rat CRP with a signal sequence of 19 amino acids and a polypeptide of 211 amino acids, the latter sharing extensive homology with human, rabbit, and mouse CRPs. The deduced sequence agreed with results obtained from partial microsequencing and mapping by fast atom bombardment-mass spectrometry. Two potential sites for N-glycosylation (Asn-128 and Asn-147) and a C-terminal heptapeptide (Leu-205 to Ser-211, containing two cysteines at positions 208 and 209) were unique to rat CRP. The protein was also shown to be composed of five apparently identical monomers, two of which form a dimer linked by two interchain disulfide bonds involving Cys-208 and Cys-209. These same cysteines form an intrachain disulfide bond in the other three monomers. The primary structure of rat CRP and the basis of dimer formation have, therefore, been elucidated.
Assuntos
Proteína C-Reativa/genética , DNA/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteína C-Reativa/isolamento & purificação , Proteína C-Reativa/metabolismo , Cromatografia Líquida de Alta Pressão , Clonagem Molecular , Eletroforese em Gel de Poliacrilamida , Biblioteca Gênica , Humanos , Cinética , Substâncias Macromoleculares , Masculino , Espectrometria de Massas , Dados de Sequência Molecular , Peso Molecular , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Homologia de Sequência do Ácido NucleicoRESUMO
A variety of modifications of sialic acids have been described in nature. There are currently many difficulties in the detection and quantitation of these modified sialic acids from biological sources. We report here that fast-atom bombardment-mass-spectrometry (FAB-MS) of native sialic acids provides specific detection and quantitation of many previously known compounds. Derivatization of the sialic acids by reduction and peracylation under acidic conditions prior to FAB-MS provides further confirmation of their identity and improves the sensitivity of detection. Samples containing as little as 100 ng of a derivatized sialic acid loaded onto the FAB target allowed accurate identification. Mixtures of sialic acids could be analyzed, and minor components were seen, at levels undetectable by other currently known techniques. Analysis of known mixtures of different sialic acids gave reproducible relative signal intensities, indicating that quantitative data can be derived from the FAB-MS spectra. After reduction and peracylation, each sialic acid gave two major molecular ions, corresponding to the fully derivatized linear species and a lactone form, and a minor ion, corresponding to an anhydro form. Lactone formation was minimal in the case of four substituted sialic acids, indicating that the hydroxyl group at the 4-position is involved in lactonization. Differentiation between different positional isomers of the modified sialic acids could be achieved using controlled degradation with periodate, tagging of the fragments with p-aminobenzoic acid ethyl ester under acid reducing conditions, peracylation, and FAB-MS of the derivatized products. We used this FAB-MS strategy to identify a novel sialic acid, 8-O-methyl-7,9-di-O-acetyl-N-glycolyl-neuraminic acid from the starfish Pisaster brevispinus, and to demonstrate the presence of a previously undetected sialic acid, 4,8-anhydro-N-acetyl-neuraminic acid in acid hydrolysates of horse serum. We also use FAB-MS to show that the alkaline conditions traditionally used for analytical de-O-acetylation of sialic acids causes substantial conversion of 4-O-acetylated sialic acids into the same anhydro compound.
Assuntos
Espectrometria de Massas , Ácido N-Acetilneuramínico/análogos & derivados , Ácido Periódico , Ácidos Siálicos/análise , Acetilação , Acilação , Animais , Bovinos , Cromatografia Líquida de Alta Pressão , Cavalos/sangue , Hidrólise , Isomerismo , Lactonas , Conformação Molecular , Estrutura Molecular , Mucinas/análise , Oxirredução , Estrelas-do-Mar , Glândula Submandibular/análiseRESUMO
A mass-spectrometric approach is presented for the analysis of the structures of lipopolysaccharide-derived oligosaccharides, which are frequently difficult to define by classical methods since they contain chemically labile components. The method involves f.a.b.-m.s. of the oligosaccharides, their peracetylated and permethylated derivatives, their deuterioacetylated and methylated analogues, and the fragments obtained during graded methanolysis of the methylated analogues. Data obtained from two representative meningococcal LPS oligosaccharides define the sequence, patterns of branching, and the extent and location of the phosphorylethanolamine and O-acetyl substituents.