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Ten Feizi - One of the best experts on this subject based on the ideXlab platform.
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Neoglycolipid (NGL)-Based Glycan Microarray System for Ligand Discovery
Glycoscience: Biology and Medicine, 2021Co-Authors: R. A. Childs, Ten FeiziAbstract:It is now appreciated that carbohydrate-protein interactions are integral to many physiological processes and are directly or indirectly involved in the majority of disease processes, infective or noninfective, including cancer. Carbohydrate microarrays have emerged as powerful tools for elucidating the ligands involved in these interactions. However, as oligosaccharides cannot be cloned and expressed as with DNA and proteins, few laboratories have libraries of sequence-defined oligosaccharide probes with sufficient breadth to tackle the unraveling of diverse carbohydrate-protein interactions. Microarray screening analyses are offered to the scientific community by the Wellcome Trust-supported carbohydrate microarray facility in the Glycosciences Laboratory at Imperial College London and by the NIH-supported Consortium for Functional Glycomics. This chapter gives a brief account of a technology, the Neoglycolipid (NGL) technology, first introduced in 1985 and converted into a glycan microarray system in 2002. Results are highlighted from analyses using this system, also including designer arrays, which entail microarrays of NGLs specifically derived from relevant ligand-bearing glycomes in order to reveal the oligosaccharide ligands they harbor and lead to their isolation and characterization. These include discoveries of new ligands in endogenous recognition and pathogen-host interactions and assignments of long-sought cancer-associated antigens.
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The Neoglycolipid (NGL) technology-based microarrays and future prospects.
FEBS letters, 2018Co-Authors: Ten FeiziAbstract:The Neoglycolipid (NGL) technology is the basis of a state-of-the-art oligosaccharide microarray system, which we offer for screening analyses to the broad scientific community. We review here the sequential development of the technology and its power in pinpointing and isolating naturally occurring ligands for glycan-binding proteins (GBPs) within glycan populations. We highlight our Designer Array approach and Beam Search Array approach for generating natural glycome arrays to identify novel ligands of biological relevance. These two microarray approaches have been applied for assignments of ligands or antigens on glucan polysaccharides for effector proteins of the immune system (Dectin-1, DC-SIGN and DC-SIGNR) and carbohydrate-binding modules (CBMs) on bacterial hydrolases. We also discuss here the more recent applications to elucidate the structure of a prostate cancer- associated antigen F77 and identify ligands for adhesins of two rotaviruses, P[10] and P[19], expressed on an epithelial mucin glycoprotein.
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Issue:Glycobiology of the Immune Response
2016Co-Authors: Ten FeiziAbstract:Carbohydrate recognition in the immune system: contributions of Neoglycolipid-based microarrays to carbohydrate ligand discover
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The Neoglycolipid (NGL)-based oligosaccharide microarray system poised to decipher the meta-glycome.
Current opinion in chemical biology, 2014Co-Authors: Angelina S. Palma, Ten Feizi, Robert A Childs, Wengang Chai, Yan LiuAbstract:The Neoglycolipid (NGL) technology is the basis of a state-of-the-art oligosaccharide microarray system. The NGL-based microarray system in the Glycosciences Laboratory Imperial College London ( http://www3.imperial.ac.uk/glycosciences ) is one of the two leading platforms for glycan microarrays, being offered for screening analyses to the broad biomedical community. Highlighted in this review are the sensitivity of the analysis system and, coupled with mass spectrometry, the provision for generating ‘designer’ microarrays from glycomes to identify novel ligands of biological relevance. Among recent applications are assignments of ligands for apicomplexan parasites, pandemic 2009 influenza virus, polyoma and reoviruses, an innate immune receptor against fungal pathogens, Dectin-1, and a novel protein of the endoplasmic reticulum, malectin; also the characterization of an elusive cancer-associated antigen. Some other contemporary advances in glycolipid-containing arrays and microarrays are also discussed.
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Neoglycolipid-based oligosaccharide microarray system: preparation of NGLs and their noncovalent immobilization on nitrocellulose-coated glass slides for microarray analyses.
Methods in molecular biology (Clifton N.J.), 2011Co-Authors: Yan Liu, Robert A Childs, Wengang Chai, Maria A. Campanero-rhodes, Mark S. Stoll, Angelina S. Palma, Ten FeiziAbstract:Carbohydrate microarrays, since their advent in 2002, are revolutionizing studies of the molecular basis of protein-carbohydrate interactions both in endogenous recognition systems and pathogen-host interactions. We have developed a unique carbohydrate microarray system based on the Neoglycolipid (NGL) technology, a well-validated microscale approach for generating lipid-tagged oligosaccharide probes for use in carbohydrate recognition studies. This chapter provides an overview of the principles and key features of the NGL-based oligosaccharide microarrays, and describes in detail the basic techniques - from the preparation of NGL probes to the generation of microarrays using robotic arraying hardware, as well as a general protocol for probing the microarrays with carbohydrate-binding proteins.
Shigeyuki Fukui - One of the best experts on this subject based on the ideXlab platform.
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Detection of oligosaccharide ligands for Hepatocyte growth factor/Scatter factor (HGF/SF), Keratinocyte growth factor (KGF/FGF-7), RANTES and Heparin cofactor II by Neoglycolipid microarrays of glycosaminoglycan-derived oligosaccharide fragments
Glycoconjugate Journal, 2006Co-Authors: Keiko Yamaguchi, Hirotoshi Tamaki, Shigeyuki FukuiAbstract:Neoglycolipid technology is eminently adaptable for microarray design for high-throughput detection and specificity assignments of carbohydrate-protein interactions. Dermatan sulfate (DS) is known to play an important role because of its ability to bind growth factors as well as chemokines and to modulate their biological activities during inflammation and response to injury. We prepared various iduronic acid-rich fragments from DS by complete digestion with chondroitinase ACI, and investigated whether the DS-binding proteins, such as HGF/SF, RANTES, KGF/FGF-7 and HCII, can detect their oligosaccharide ligands in a Neoglycolipid microarray. First, a comparison of the intensity of binding signals obtained from chondroitin oligosaccharides with those of heparin oligosaccharides showed that our microarray system is feasible not only to single-out the oligosaccharide ligands, but also to detect the difference between an intrinsic interaction unrelated only to electrostatic interaction and non-specific electrostatic interaction. Second, HGF/SF, KGF/FGF-7 and HCII showed preferential binding to iduronic acid-rich fragments of DS oligosaccharides that are greater than 8-mers in lengths. In contrast, RANTES binding seemed to depend only on the negative charges; their binding intensity towards the DS oligosaccharides was somewhat stronger than the binding of HGF/SF, KGF/FGF-7 and HCII. Third, the use of polyvinylpyrrolidone-40 (PVP-40), ovalbumin (OV) and Tween 20 in place of BSA as a blotting agent was useful in these glycosaminoglycan dependent reactions to minimize background due to non-specific interactions.
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detection of oligosaccharide ligands for hepatocyte growth factor scatter factor hgf sf keratinocyte growth factor kgf fgf 7 rantes and heparin cofactor ii by Neoglycolipid microarrays of glycosaminoglycan derived oligosaccharide fragments
Glycoconjugate Journal, 2006Co-Authors: Keiko Yamaguchi, Hirotoshi Tamaki, Shigeyuki FukuiAbstract:Neoglycolipid technology is eminently adaptable for microarray design for high-throughput detection and specificity assignments of carbohydrate-protein interactions. Dermatan sulfate (DS) is known to play an important role because of its ability to bind growth factors as well as chemokines and to modulate their biological activities during inflammation and response to injury. We prepared various iduronic acid-rich fragments from DS by complete digestion with chondroitinase ACI, and investigated whether the DS-binding proteins, such as HGF/SF, RANTES, KGF/FGF-7 and HCII, can detect their oligosaccharide ligands in a Neoglycolipid microarray. First, a comparison of the intensity of binding signals obtained from chondroitin oligosaccharides with those of heparin oligosaccharides showed that our microarray system is feasible not only to single-out the oligosaccharide ligands, but also to detect the difference between an intrinsic interaction unrelated only to electrostatic interaction and non-specific electrostatic interaction. Second, HGF/SF, KGF/FGF-7 and HCII showed preferential binding to iduronic acid-rich fragments of DS oligosaccharides that are greater than 8-mers in lengths. In contrast, RANTES binding seemed to depend only on the negative charges; their binding intensity towards the DS oligosaccharides was somewhat stronger than the binding of HGF/SF, KGF/FGF-7 and HCII. Third, the use of polyvinylpyrrolidone-40 (PVP-40), ovalbumin (OV) and Tween 20 in place of BSA as a blotting agent was useful in these glycosaminoglycan dependent reactions to minimize background due to non-specific interactions.
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detection of oligosaccharide ligands for hepatocyte growth factor scatter factor hgf sf keratinocyte growth factor kgf fgf 7 rantes and heparin cofactor ii by Neoglycolipid microarrays of glycosaminoglycan derived oligosaccharide fragments
Glycoconjugate Journal, 2006Co-Authors: Keiko Yamaguchi, Hirotoshi Tamaki, Shigeyuki FukuiAbstract:Neoglycolipid technology is eminently adaptable for microarray design for high-throughput detection and specificity assignments of carbohydrate-protein interactions. Dermatan sulfate (DS) is known to play an important role because of its ability to bind growth factors as well as chemokines and to modulate their biological activities during inflammation and response to injury. We prepared various iduronic acid-rich fragments from DS by complete digestion with chondroitinase ACI, and investigated whether the DS-binding proteins, such as HGF/SF, RANTES, KGF/FGF-7 and HCII, can detect their oligosaccharide ligands in a Neoglycolipid microarray. First, a comparison of the intensity of binding signals obtained from chondroitin oligosaccharides with those of heparin oligosaccharides showed that our microarray system is feasible not only to single-out the oligosaccharide ligands, but also to detect the difference between an intrinsic interaction unrelated only to electrostatic interaction and non-specific electrostatic interaction. Second, HGF/SF, KGF/FGF-7 and HCII showed preferential binding to iduronic acid-rich fragments of DS oligosaccharides that are greater than 8-mers in lengths. In contrast, RANTES binding seemed to depend only on the negative charges; their binding intensity towards the DS oligosaccharides was somewhat stronger than the binding of HGF/SF, KGF/FGF-7 and HCII. Third, the use of polyvinylpyrrolidone-40 (PVP-40), ovalbumin (OV) and Tween 20 in place of BSA as a blotting agent was useful in these glycosaminoglycan dependent reactions to minimize background due to non-specific interactions.
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Carbohydrate Microarray for Deciphering the Information Embedded in Oligosaccharide Structures
Experimental Glycoscience, 1Co-Authors: Shigeyuki FukuiAbstract:The Neoglycolipid (NGL) technology for generating lipid-linked oligosaccharide probes has been shown to be an extremely useful tool for detection and characterization of bioactive oligosaccharides that are extremely minor components (Feizi and Child 1994). As the affinities of most carbohydrate-protein interactions are thought to be very low, and the oligosaccharide portions that play a role in biological function are generally believed to be of a small region ranging from monosaccharide to decasaccharide, the Neoglycolipid technology is eminently adaptable for microarray design for high-throughput detection and specificity assignments of carbohydrate-protein interactions.
Paul Boullanger - One of the best experts on this subject based on the ideXlab platform.
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Syntheses of deuterium‐labelled cholesteryl Neoglycolipids
Journal of Labelled Compounds and Radiopharmaceuticals, 2012Co-Authors: Dominique Lafont, Paul Boullanger, Antoine GambettaAbstract:Four deuterium-labelled Neoglycolipids derived from cholesterol were synthesized for embedment into liposomes. Deuterium atoms were either incorporated by CH2 replacement with a CD2 group in the triethylene glycol spacer arm between the cholesteryl residue and the sugar moiety (products 2–4) or incorporated directly on the acetamido function in the sugar head (compound 5).
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Establishment of mass spectrometric fingerprints of novel synthetic cholesteryl Neoglycolipids: The presence of a unique C-glycoside species during electrospray ionization and during collision-induced dissociation tandem mass spectrometry
Journal of the American Society for Mass Spectrometry, 2007Co-Authors: Anas El-aneed, Paul Boullanger, Joseph Banoub, Mariano Koen-alonso, Dominique LafontAbstract:In this study we evaluated the fragmentation pattern of 16 novel amphiphilic Neoglycolipid cholesteryl derivatives that can be efficiently used to increase cationic liposomal stability and to enhance gene transfer ability. These Neoglycolipids bear different sugar moieties, such as D-glucosamine, N -acetyl-D-glucosamine_, N -trideuterioacetyl-D-glucosamine, N -acetyllactosamine, L-fucose, N -allyloxycarbonyl-D-glucosamine, and some of their per- O -acetylated derivatives. Regardless of the structure of the tested Neoglycolipid, QqToF-MS analysis using electrospray ionization (ESI) source showed abundant protonated [M+H]^+ species. We also identified by both QqToF-MS and low-energy collision tandem mass spectrometry (CID-MS/MS) of the [M+H]^+ ion, the presence of specific common fingerprint fragment ions: [Cholestene]^+, sugar [oxonium]^+, [(Sugar-spacer-OH)+H]^+, [oxonium-H_2O]^+, and [(Cholesterol-spacer-OH)+H]^+. In addition, we observed a unique ion that could not be rationally explained by the expected fragmentation of these amphiphilic molecules. The structure of this ion was tentatively proposed with that of a C -glycoside species formed by a chemical reaction between the sugar portion and the cholesterol. MS/MS analysis of this unique [ C -glycoside]^+ confirmed the validity of the proposed structure of this ion. The presence of an amino group at position C-2 and free hydroxyl groups of the sugar motif is crucial for the formation of a “reactive” sugar oxonium ion that can form the [ C -glycoside]^+ species. In summary, we precisely established the fragmentation patterns of the tested series of Neoglycolipid cholesteryl derivatives and authenticated their structure as well; moreover, we speculated on the formation of a C -glycoside with the ESI source under atmospheric pressure and in the collision cell during MS/MS analysis.
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Syntheses of α-d-galactosamine Neoglycolipids
Carbohydrate research, 2006Co-Authors: Nicolas Laurent, Dominique Lafont, Paul BoullangerAbstract:Abstract Several N-acetyl-α- d -galactosamine Neoglycolipids, as well as hydrophobized T and TN antigen analogues, were prepared for embedment onto liposomes. Three different lipidic structures were used for the anchoring, that is cholesterol, 1,3-bis(undecyloxy)propan-2-ol and 1,3-bis(3,7,11,15-tetramethylhexadecyloxy)propan-2-ol. Oligoethyleneglycol spacers were used to link the carbohydrate and the hydrophobic moieties; their lengths were varied in order to obtain model compounds for the selective recognition by sialyl transferases involved in cancer processes. Glycosylation reactions were optimized to sluggish amphiphilic acceptor alcohols, in order to reach good 1,2-cis-stereoselectivities and acceptable yields. This aim was achieved by using 3,4,6-tri-O-acetyl-2-azido-2-deoxy- d -galactopyranosyl trichloroacetimidate as the donor, trimethylsilyl trifluoromethanesulfonate as the promoter and diethyl ether or mixtures of diethyl ether and dichloromethane as solvents.
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Specific interaction of lectins with liposomes and monolayers bearing Neoglycolipids
Chemistry and physics of lipids, 2003Co-Authors: Vincent Faivre, Paul Boullanger, Maria De Lourdes Costa, Adam Baszkin, Véronique RosilioAbstract:Abstract The interaction of three lectins (wheat germ, Ulex europaeus I, and Lotus tetragonolobus agglutinins: WGA, UEA-I and LTA) with either N-acetyl- d -glucosamine or l -fucose Neoglycolipids incorporated into phospholipid monolayers and liposome bilayers was studied at the air/water interface and in bulk solution. The results show that for both systems studied, synthesized Neoglycolipids were capable of binding their specific lectin and that, in general, the binding of lectins increased with the increase in the molar fraction of the saccharide derivative incorporated in either the monolayers or bilayers. However, whereas for UEA-I, molecular recognition was enhanced by a strong hydrophobic interaction, for WGA and LTA successful recognition was predominantly related to the distance between neighboring sugar groups. The observed lengthy adsorption times of these lectins onto their specific ligands were attributed to interfacial conformational changes occurring in the proteins upon their adsorption at the interfaces.
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Syntheses and interfacial behaviour of Neoglycolipid analogues of glycosyl ceramides
Carbohydrate research, 2001Co-Authors: Dominique Lafont, Marie-noëlle Bouchu, Agnès Girard-egrot, Paul BoullangerAbstract:Abstract Four glycosyl ceramides analogues having d -galactose or 2-acetamido-2-deoxy- d -glucose moieties linked to enantiomeric lipids have been synthesised to study their interfacial behaviour at the air ∣ water interface. The lipid chains were prepared in two steps by opening 1,2-epoxyhexadecane using Jacobsen kinetic hydrolytic resolution (KHR) followed by an azidosilylation reaction of the diol so obtained. Glycosylation reactions were realised either with 2,3,4,6-tetra- O -benzoyl-α- d -galactopyranosyl trichloroacetimidate or 1,3,4,6-tetra- O -acetyl-2-allyloxycarbonylamino-2-deoxy-β- d -glucopyranose as donors and (2 R )- or (2 S )-2-azidohexadecanol derivatives as acceptors. Transformation of the azido glycosides into N -acylated products was done by a modified Staudinger reaction in the presence of fatty acyl chlorides. The four Neoglycolipids are able to form a condensed monolayer at the air ∣ water interface; their π-A isotherm diagrams are similar to that described for the natural glycosyl ceramides. The detailed analysis of the isotherms, taking into account the chirality of the lipid chains, allowed to determine the contribution of the different parts of the molecule under the monolayer packing.
Wengang Chai - One of the best experts on this subject based on the ideXlab platform.
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The Neoglycolipid (NGL)-based oligosaccharide microarray system poised to decipher the meta-glycome.
Current opinion in chemical biology, 2014Co-Authors: Angelina S. Palma, Ten Feizi, Robert A Childs, Wengang Chai, Yan LiuAbstract:The Neoglycolipid (NGL) technology is the basis of a state-of-the-art oligosaccharide microarray system. The NGL-based microarray system in the Glycosciences Laboratory Imperial College London ( http://www3.imperial.ac.uk/glycosciences ) is one of the two leading platforms for glycan microarrays, being offered for screening analyses to the broad biomedical community. Highlighted in this review are the sensitivity of the analysis system and, coupled with mass spectrometry, the provision for generating ‘designer’ microarrays from glycomes to identify novel ligands of biological relevance. Among recent applications are assignments of ligands for apicomplexan parasites, pandemic 2009 influenza virus, polyoma and reoviruses, an innate immune receptor against fungal pathogens, Dectin-1, and a novel protein of the endoplasmic reticulum, malectin; also the characterization of an elusive cancer-associated antigen. Some other contemporary advances in glycolipid-containing arrays and microarrays are also discussed.
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Neoglycolipid-based oligosaccharide microarray system: preparation of NGLs and their noncovalent immobilization on nitrocellulose-coated glass slides for microarray analyses.
Methods in molecular biology (Clifton N.J.), 2011Co-Authors: Yan Liu, Robert A Childs, Wengang Chai, Maria A. Campanero-rhodes, Mark S. Stoll, Angelina S. Palma, Ten FeiziAbstract:Carbohydrate microarrays, since their advent in 2002, are revolutionizing studies of the molecular basis of protein-carbohydrate interactions both in endogenous recognition systems and pathogen-host interactions. We have developed a unique carbohydrate microarray system based on the Neoglycolipid (NGL) technology, a well-validated microscale approach for generating lipid-tagged oligosaccharide probes for use in carbohydrate recognition studies. This chapter provides an overview of the principles and key features of the NGL-based oligosaccharide microarrays, and describes in detail the basic techniques - from the preparation of NGL probes to the generation of microarrays using robotic arraying hardware, as well as a general protocol for probing the microarrays with carbohydrate-binding proteins.
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n glycolyl gm1 ganglioside as a receptor for simian virus 40
Journal of Virology, 2007Co-Authors: Maria A Campanerorhodes, Alicia E Smith, Robert A Childs, Yibing Zhang, Helge Ewers, Laura Mauri, Sandro Sonnino, Wengang Chai, Ari Helenius, Anne ImbertyAbstract:Carbohydrate microarrays have emerged as powerful tools in analyses of microbe-host interactions. Using a microarray with 190 sequence-defined oligosaccharides in the form of natural glycolipids and Neoglycolipids representative of diverse mammalian glycans, we examined interactions of simian virus 40 (SV40) with potential carbohydrate receptors. While the results confirmed the high specificity of SV40 for the ganglioside GM1, they also revealed that N-glycolyl GM1 ganglioside [GM1(Gc)], which is characteristic of simian species and many other nonhuman mammals, is a better ligand than the N-acetyl analog [GM1(Ac)] found in mammals, including humans. After supplementing glycolipid-deficient GM95 cells with GM1(Ac) and GM1(Gc) gangliosides and the corresponding Neoglycolipids with phosphatidylethanolamine lipid groups, it was found that GM1(Gc) analogs conferred better virus binding and infectivity. Moreover, we visualized the interaction of NeuGc with VP1 protein of SV40 by molecular modeling and identified a conformation for GM1(Gc) ganglioside in complex with the virus VP1 pentamer that is compatible with its presentation as a membrane receptor. Our results open the way not only to detailed studies of SV40 infection in relation to receptor expression in host cells but also to the monitoring of changes that may occur with time in receptor usage by the virus.
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Oligosaccharide microarrays to decipher the glyco code
Nature Reviews Molecular Cell Biology, 2004Co-Authors: Ten Feizi, Wengang ChaiAbstract:The oligosaccharide moieties of glycoproteins, glycolipids, proteoglycans and polysaccharides are highly diverse, the reason for this diversity is not yet understood. Neoglycolipid technology allows the generation of oligosaccharide probes with lipid tags from desired sources and is showing promise as a basis for oligosaccharide microarrays. Such microarrays would allow surveys of glycomes and proteomes to be carried out, which would enable the molecular definition of carbohydrate-recognition systems in whole organisms.
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Neoglycolipid technology: deciphering information content of glycome.
Methods in enzymology, 2003Co-Authors: Wengang Chai, Mark S. Stoll, Alexander M. Lawson, Christine Galustian, Ten FeiziAbstract:Publisher Summary Neoglycolipid technology was introduced in 1985 to enable direct binding studies to be conveniently performed with oligosaccharides of glycoproteins. At that time, there was a lack of microscale methods for examining the carbohydrate–protein interactions, after release of the oligosaccharides from the carrier proteins. Most carbohydrate–protein interactions are of such low affinities that di- or multivalence of both oligosaccharide and recognition protein is required for detection by precipitation, radioimmunoassay, or enzyme-linked immunosorbent assay experiments. It was clear that a method was required to examine the released oligosaccharides in a multivalent state. The existing methods for examining the recognition of specific oligosaccharides by antibodies and other carbohydrate-recognizing proteins involved purification of the oligosaccharides, often from highly heterogeneous mixtures, and testing them individually as inhibitors of the binding of the recognition proteins to macromolecules or cells. The amounts of oligosaccharides required for inhibition of binding were frequently prohibitive. The approach chosen to address the problem was chemical conjugation of oligosaccharides to lipid. The artificial glycolipids, Neoglycolipids, formed would enable the presentation of the oligosaccharides in the clustered state. The conjugation principle selected was reductive amination to a phosphatidylethanolaminetype aminolipid. For O-glycans released by reductive alkali treatment, a mild periodate oxidation procedure was included to generate reactive aldehydes for the conjugation. A major advantage of conjugating each oligosaccharide in a mixture to a lipid molecule rather than to a protein, such as bovine serum albumin, is that each component remains discrete and can be isolated.
Heinz Egge - One of the best experts on this subject based on the ideXlab platform.
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Cholesterol-containing lactose derived Neoglycolipids serve as acceptors for sialyltransferases from rat liver Golgi vesicles.
Glycoconjugate journal, 1996Co-Authors: Gottfried Pohlentz, Anja Mokros, Heinz EggeAbstract:The cholesterol-containing lactose derived Neoglycolipids β-Lactosylcholesterol, Cholesteryl-β-lactosylpropane-1,3-diol, 3-Cholesteryl-1-β-lactosylglycerol, 2-Cholesteryl-1-β-lactosylglycerol, 2,3-Dicholesteryl-1-β-lactosylglycerol, 1-Deoxy-1-cholesterylethanolaminolactitol, 1-Deoxy-1-cholesteryl (N-acetyl)-ethanolaminolactitol, 1-Deoxy-1-cholesterylphosphoethanolaminolactitol, and 1-Deoxy-1-cholesterylphospho (N-acetyl)-ethanolaminolactitol were synthesized and used as acceptors for sialytransferases from rat liver Golgi vesicles. Relative activities with the Neoglycolipids as acceptors varied from 28 to 163% compared to those obtained with the authentic acceptor lactosylceramide. Product identification by thin layer chromatography and fast atom bombardment mass spectrometry showed that the Neoglycolipids yielded mono- and disialylated products. The results of competition experiments suggested that lactosylceramide and the Neoglycolipids were sialylated by the same enzymes.
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Neoglycolipids of 1-deoxy-1-phosphatidylethanolaminolactitol type: synthesis, structure analysis, and use as probes for characterization of glycosyltransferases.
Methods in enzymology, 1994Co-Authors: Gottfried Pohlentz, Heinz EggeAbstract:Publisher Summary Neoglycolipids (NeoGL) of the 1-deoxy- 1-phosphatidylethanolamino-lactitol type (Lac-PtdEtn), synthesized by coupling an oligosaccharide to phosphatidylethanolamine (PtdEtn) by reductive amination, have been used for studies on antigenicity, receptor function, and lectin and toxin binding by carbohydrates. Glycosyl-PtdEtns and especially the N-acetylated derivatives can serve as acceptors for glycosphingolipid (GSL) glycosyltransferases. The NeoGLs are as good acceptors as the authentic GSL analogs, and they are glycosylated by the same enzymes. This chapter describes an improved synthesis of Lac-PtdEtn-type Neoglycolipids, their N-acetylation, and their use as acceptors for glycosyl-transferases and for structure elucidation of oligosaccharides by fast atom bombardment-mass spectrometry (FAB-MS). Radiolabeled NeoGL can be useful tools for investigations on glycolipid metabolism in cell culture. For the glycosyl-PtdEtn compounds to serve as substitutes for natural GSL in glycosyltransferase assays, it must be ascertained that Neoglycolipids are converted to the analogous products by the same enzymes that act on natural GSL and that they exhibit similar apparent kinetic constants. This chapter illustrates the use of glycosyl-PtdEtns and the N-acetylated derivatives as acceptors for sialyltransferases from rat liver Golgi and discusses the transferase products by FAB-MS. By competition experiments it is demonstrated that the NeoGL and the authentic GSL analog are sialylated by the same respective enzymes.
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1 deoxy 1 phosphatidylethanolamino lactitol type Neoglycolipids serve as acceptors for sialyltransferases from rat liver golgi vesicles
FEBS Journal, 1992Co-Authors: Gottfried Pohlentz, Susanne Schlemm, Heinz EggeAbstract:1-Deoxy-1-phosphatidylethanolamino-lactitols (LacPtdEtns), 1-deoxy-1-phosphatidylethanol-amino-sialyllactitols (NeuAcLacPtdEtns) and their corresponding N-acetylated derivatives were synthesized and characterized by fast-atom-bombardment mass spectrometry (FAB MS). The Neoglycolipids were used as acceptors for sialyltransferases from rat liver Golgi vesicles. Sialylation rates were as good as or even better than those obtained with the corresponding authentic acceptors lactosylceramide (LacCer) and ganglioside GM3. The sialylation of LacPtdEtns and NeuAcLacPtdEtns yielded sialyl and disialyl compounds, respectively, as shown by FAB MS analysis of the reaction products. The results of competition experiments indicate that the Neoglycolipids and the authentic acceptors are sialylated by the same sialyltransferases.
