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Carbohydrate Chains (carbohydrate + chain)
Selected AbstractsFull Structure of the Carbohydrate Chain of the Lipopolysaccharide of Providencia rustigianii,O34CHEMISTRY - A EUROPEAN JOURNAL, Issue 20 2008Abstract A lipopolysaccharide isolated from an opportunistic pathogen of the Enterobacteriaceae family Providencia rustigianii,O34 was found to be a mixture of R-, SR-, and S - forms consisting of a lipid moiety (lipid,A) that bears a core oligosaccharide, a core with one O-polysaccharide repeating unit attached, and a long-chain O-polysaccharide, respectively. The corresponding carbohydrate moieties were released from the lipopolysaccharide by mild acid hydrolysis and studied by sugar and methylation analyses along with one- and two-dimensional NMR spectroscopy and high-resolution electrospray ionization mass spectrometry. As a result, the structures of the core and the O-polysaccharide were established, including the structure of the biological repeating unit (an oligosaccharide that is preassembled and polymerized in biosynthesis of the O-polysaccharide), as well as the mode of the linkage between the O-polysaccharide and the core. Combining the structure of the carbohydrate moiety thus determined and the known structure of lipid,A enabled determination of the full lipopolysaccharide structure of P. rustigianii,O34. [source] Advances on the compositional analysis of glycosphingolipids combining thin-layer chromatography with mass spectrometryMASS SPECTROMETRY REVIEWS, Issue 3 2010Johannes Müthing Abstract Glycosphingolipids (GSLs), composed of a hydrophilic carbohydrate chain and a lipophilic ceramide anchor, play pivotal roles in countless biological processes, including infectious diseases and the development of cancer. Knowledge of the number and sequence of monosaccharides and their anomeric configuration and linkage type, which make up the principal items of the glyco code of biologically active carbohydrate chains, is essential for exploring the function of GSLs. As part of the investigation of the vertebrate glycome, GSL analysis is undergoing rapid expansion owing to the application of novel biochemical and biophysical technologies. Mass spectrometry (MS) takes part in the network of collaborations to further unravel structural and functional aspects within the fascinating world of GSLs with the ultimate aim to better define their role in human health and disease. However, a single-method analytical MS technique without supporting tools is limited yielding only partial structural information. Because of its superior resolving power, robustness, and easy handling, high-performance thin-layer chromatography (TLC) is widely used as an invaluable tool in GSL analysis. The intention of this review is to give an insight into current advances obtained by coupling supplementary techniques such as TLC and mass spectrometry. A retrospective view of the development of this concept and the recent improvements by merging (1) TLC separation of GSLs, (2) their detection with oligosaccharide-specific proteins, and (3) in situ MS analysis of protein-detected GSLs directly on the TLC plate, are provided. The procedure works on a nanogram scale and was successfully applied to the identification of cancer-associated GSLs in several types of human tumors. The combination of these two supplementary techniques opens new doors by delivering specific structural information of trace quantities of GSLs with only limited investment in sample preparation. © 2009 Wiley Periodicals, Inc. Mass Spec Rev 29:425-479, 2010 [source] Heparan sulfate proteoglycans in experimental models of diabetes: a role for perlecan in diabetes complicationsDIABETES/METABOLISM: RESEARCH AND REVIEWS, Issue 6 2001Karin Conde-Knape Abstract Proteoglycans are ubiquitous extracellular proteins that serve a variety of functions throughout the organism. Unlike other glycoproteins, proteoglycans are classified based on the structure of the glycosaminoglycan carbohydrate chains, not the core proteins. Perlecan, a member of the heparan sulfate proteoglycan (HSPG) family, has been implicated in many complications of diabetes. Decreased levels of perlecan have been observed in the kidney and in other organs, both in patients with diabetes and in animal models. Perlecan has an important role in the maintenance of the glomerular filtration barrier. Decreased perlecan in the glomerular basement membrane has a central role in the development of diabetic albuminuria. The involvement of this proteoglycan in diabetic complications and the possible mechanisms underlying such a role have been addressed using a variety of models. Due to the importance of nephropathy among diabetic patients most of the studies conducted so far relate to diabetes effects on perlecan in different types of kidney cells. The various diabetic models used have provided information on some of the mechanisms underlying perlecan's role in diabetes as well as on possible factors affecting its regulation. However, many other aspects of perlecan metabolism still await full elucidation. The present review provides a description of the models that have been used to study HSPG and in particular perlecan metabolism in diabetes and some of the factors that have been found to be important in the regulation of perlecan. Copyright © 2001 John Wiley & Sons, Ltd. [source] Lactosamine modulates the rate of migration of GnRH neurons during mouse developmentEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 3 2006Elizabeth Bless Abstract Gonadotropin-releasing hormone (GnRH) neurons are derived from progenitor cells in the olfactory placodes and migrate from the vomeronasal organ (VNO) across the cribriform plate into the forebrain. At embryonic day (E)12 in the mouse most of these neurons are still in the nasal compartment but by E15 most GnRH neurons have migrated into the forebrain. Glycoconjugates with carbohydrate chains containing terminal lactosamine are expressed by neurons in the main olfactory epithelium and in the VNO. One of the key enzymes required to regulate the synthesis and expression of lactosamine, ,1,3-N-acetylglucosaminyltransferase-1 (,3GnT1), is strongly expressed by neurons in the olfactory epithelium and VNO, and on neurons migrating out of the VNO along the GnRH migratory pathway. Immunocytochemical analysis of lactosamine and GnRH in embryonic mice reveals that the percentage of lactosamine+,GnRH+ double-labeled neurons decreases from >,80% at E13, when migration is near its peak, to ,,30% at E18.5, when most neurons have stopped migrating. In ,3GnT1,/, mice, there is a partial loss of lactosamine expression on GnRH neurons. Additionally, a greater number of GnRH neurons were retained in the nasal compartment of null mice at E15 while fewer GnRH neurons were detected later in embryonic development in the ventral forebrain. These results suggest that the loss of lactosamine on a subset of GnRH neurons impeded the rate of migration from the nose to the brain. [source] Human lactoferrin activates NF-,B through the Toll-like receptor 4 pathway while it interferes with the lipopolysaccharide-stimulated TLR4 signalingFEBS JOURNAL, Issue 9 2010Ken Ando Lactoferrin (LF) has been implicated in innate immunity. Here we reveal the signal transduction pathway responsible for human LF (hLF)-triggered nuclear factor-,B (NF-,B) activation. Endotoxin-depleted hLF induces NF-,B activation at physiologically relevant concentrations in the human monocytic leukemia cell line, THP-1, and in mouse embryonic fibroblasts (MEFs). In MEFs, in which both tumor necrosis factor receptor-associated factor 2 (TRAF2) and TRAF5 are deficient, hLF causes NF-,B activation at a level comparable to that seen in wild-type MEFs, whereas TRAF6-deficient MEFs show significantly impaired NF-,B activation in response to hLF. TRAF6 is known to be indispensable in leading to NF-,B activation in myeloid differentiating factor 88 (MyD88)-dependent signaling pathways, while the role of TRAF6 in the MyD88-independent signaling pathway has not been clarified extensively. When we examined the hLF-dependent NF-,B activation in MyD88-deficient MEFs, delayed, but remarkable, NF-,B activation occurred as a result of the treatment of cells with hLF, indicating that both MyD88-dependent and MyD88-independent pathways are involved. Indeed, hLF fails to activate NF-,B in MEFs lacking Toll-like receptor 4 (TLR4), a unique TLR group member that triggers both MyD88-depependent and MyD88-independent signalings. Importantly, the carbohydrate chains from hLF are shown to be responsible for TLR4 activation. Furthermore, we show that lipopolysaccharide-induced cytokine and chemokine production is attenuated by intact hLF but not by the carbohydrate chains from hLF. Thus, we present a novel model concerning the biological function of hLF: hLF induces moderate activation of TLR4-mediated innate immunity through its carbohydrate chains; however, hLF suppresses endotoxemia by interfering with lipopolysaccharide-dependent TLR4 activation, probably through its polypeptide moiety. [source] Regulated expression and intracellular localization of cystatin F in human U937 cellsFEBS JOURNAL, Issue 22 2002Carl-Michael Nathanson Cystatin F is a cysteine peptidase inhibitor recently discovered in haematopoietic cells by cDNA cloning. To further investigate the expression, distribution and properties of the native human inhibitor the promyeloid cell line U937 has been studied. The cells expressed relatively large quantities of cystatin F, which was found both secreted and intracellularly. The intracellular levels were unusually high for a secreted cystatin (, 25% of the cystatin F in 2- or 4-day culture medium). By contrast, U937 cells contained only 3,4% of the related inhibitor, cystatin C. Cystatin F purified from lysates of U937 cells showed three major forms carrying two, one or no carbohydrate chains. Immunocytochemistry demonstrated a marked cytoplasmic cystatin F staining in a granular pattern. Double staining with a marker for endoplasmic reticulum revealed no colocalization for cystatin F. Analysis of the promoter region of the cystatin F gene (CST7) showed that it, like that of the cystatin C gene (CST3), is devoid of typical TATA- and CAAT-box elements. In contrast to the cystatin C promoter, it does not contain multiple Sp1 binding sites, but has a unique site for C/EBP,, possibly explaining the restricted expression of the cystatin F gene. Cells stimulated with all- trans retinoic acid to differentiate them towards a granulocytic pathway, showed a strong (, 18-fold) down-regulation of intracellular cystatin F and almost abolished secreted levels of the inhibitor. Stimulation with tetradecanoyl phorbol acetate, causing monocytic differentiation, also resulted in down-regulation (two fold to threefold) of cystatin F expression, whereas the cystatin C expression was essentially unaltered in both experiments. The results suggest that cystatin F as an intracellular cysteine peptidase inhibitor with readily regulated expression, may be a candidate to control the cysteine peptidase activity known to be essential for antigen presentation in different blood cell lineages. [source] Analysis of an antibody pharmaceutical, tocilizumab, by capillary electrophoresis using a carboxylated capillaryJOURNAL OF SEPARATION SCIENCE, JSS, Issue 5 2008Atsushi Taga Abstract Antibody pharmaceuticals are becoming more and more prevalent due to their excellent effectiveness in clinical medications, and are expected to allow tailor-made medical treatment for rheumatic diseases, immunosuppression in cardiac transplantation, and cancer. Antibody-type pharmaceuticals of immunoglobulin G (IgG) commonly have N -glycosylated carbohydrate chains attached to heavy chains. The carbohydrate chains play important roles in the effectiveness of antibodies. Therefore evaluation of a glycosylated species is important in the first step of quality control of antibody pharmaceuticals. In the present work, we examined capillary electrophoresis with a newly developed, chemically modified capillary, the inner surface of which is modified with carboxyl groups, for evaluation of IgG molecular species which have carbohydrate chains; tocilizumab was used as a model. The analytical system developed in the present study is useful for determining the content of non-glycosylated peptides. In the analysis of tocilizumab, the ratio of non-glycosylated peptide was estimated to be 1.23% with a relative standard deviation of 3.05%. The method affords high reproducibility with simple operation, and analysis can be completed within 6 min. [source] Antigenic properties of the GroEL-like protein of Campylobacter rectusMOLECULAR ORAL MICROBIOLOGY, Issue 1 2002D. Hinode The purpose of this study was to clarify the antigenic properties of the GroEL-like protein of Campylobacter rectus using a specific polyclonal antibody directed to the purified 64-kDa GroEL-like protein (pAb- CrGroEL), a polyclonal antibody directed to the Actinobacillus actinomycetemcomitans GroEL-like protein (pAb- AaGroEL) and a monoclonal antibody against the recombinant human HSP60 (mAb-HuHSP60). In SDS-PAGE/Western immunoblotting analysis, mAb-HuHSP60, pAb- CrGroEL and pAb- AaGroEL were found to react with the GroEL-like protein (64-kDa) present in all C. rectus strains. A 150-kDa protein in C. rectus ATCC 33238 also reacted strongly with pAb- CrGroEL. This 150-kDa protein was found to be present on the surface-associated material of bacterial cells, as determined by transmission electron microscopy and immunogold labelling of cells with pAb- CrGroEL. Analysis of the first 20 N -terminal amino acids of the sequence of the 150-kDa protein revealed a strong homology (80%) with the C. rectus surface layer (S-layer) protein. Investigation of the biochemical nature of antigenic determinants using periodic acid and proteolytic enzymes showed that the C. rectus GroEL-like protein possessed immunodominant epitopes in both peptide and carbohydrate chains, and that the immunoreactive determinants of the 150-kDa protein belonged to carbohydrate. These results suggest that the GroEL-like protein and the S-layer protein of C. rectus may share the same carbohydrate epitopes. [source] Tissue distribution of histo-blood group antigens.APMIS, Issue 1 2000Vibeke Ravn The introduction of immunohistochemical techniques and monoclonal antibodies to specific carbohydrate epitopes has made it possible to study in detail the tissue distribution of histo-blood group antigens and related carbohydrate structures. The present paper summarizes the available data concerning the histological distribution of histo-blood group antigens and their precursor structures in normal human tissues. Studies performed have concentrated on carbohydrate antigens related to the ABO, Lewis, and TTn blood group systems, i.e. histo-blood group antigens carried by type 1, 2, and 3 chain carrier carbohydrate chains. Histo-blood group antigens are found in most epithelial tissues. Meanwhile, several factors influence the type, the amount, and the histological distribution of histo-blood group antigens, i.e. the ABO, Lewis, and saliva-secretor type of the individual, and the cell-and tissue type. Oligosaccharides with blood-group specificity are synthesized by the stepwise action of specific gene-encoded glycosyltransferases. In general, this stepwise synthesis of histo-blood group antigens correlates with cellular differentiation. The H and the Se genes both encode an ,1,2fucosyltransferase, which is responsible for the synthesis of blood group antigen H from precursor disaccharides. A new model for the participation of the Se/H-gene-encoded glycosyl transferases in synthesis of terminal histo-blood group antigens in human tissues is proposed; the type and degree of differentiation rather than the embryologic origin determines whether it is the H or the Se gene-encoded transferases that influence expression of terminal histo-blood group antigens in tissues. 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