- In vitro synthesis of mucin-type O-glycans using saccharide primers comprising GalNAc-Ser and GalNAc-Thr residues
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Mucin-type O-glycosylation of serine or threonine residue in proteins is known to be one of the major post-translational modifications. In this study, two novel alkyl glycosides, Nα-lauryl-O-(2-acetamido-2-deoxy-α-D-galactopyranosyl)-L-serineamide (GalNAc-Ser-C12) and Nα-lauryl-O-(2-acetamido-2-deoxy-α-D-galactopyranosyl)-L-threonineamide (GalNAc-Thr-C12) were synthesized as saccharide primers to prime mucin-type O-glycan biosynthesis in cells. Upon incubating human gastric cancer MKN45 cells with the saccharide primers, 22 glycosylated products were obtained, and their structures were analyzed using liquid chromatography-mass spectrometry and enzyme digestion. The amounts of glycosylated products were dependent on the amino acid residues in the saccharide primers. For example, in vitro synthesis of T antigen (Galβ1-3GalNAc), fucosyl-T (Fucα1-2Galβ1-3GalNAc), and sialyl-T (NeuAcα2-3Galβ1-3GalNAc) preferred a serine residue, whereas sialyl-Tn (NeuAcα2-6GalNAc) preferred a threonine residue. Furthermore, the glycosylated products derived from GalNAc-Ser/Thr-C12 and Gal-GalNAc-Ser/Thr-C12 using cell-free synthesis showed the same amino acid selectivity as those in the cell experiments. These results indicate that glycosyltransferases involved in the biosynthesis of mucin-type O-glycans distinguish amino acid residues conjugated to GalNAc. The saccharide primers developed in this study might be useful for comparing mucin-type oligosaccharides in cells and constructing oligosaccharide libraries to study cell function.
- Sakura, Ryuma,Nagai, Kaori,Yagi, Yuka,Takahashi, Yoshihisa,Ide, Yoshimi,Yagi, Yuki,Yamamoto, Daiki,Mizuno, Mamoru,Sato, Toshinori
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- Efficient and reproducible synthesis of an Fmoc-protected Tn antigen
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This concise total synthesis of the Thomsen-Nouveau (Tn) glycoconjugate was accomplished using a palladium-catalyzed coupling between the glycosyl donor and Fmoc-protected serine acceptor. This is, to the best of our knowledge, the shortest synthesis reported from galactose for preparing this essential building block for large-scale solid phase peptide synthesis.
- Chapa-Villarreal, Fabiola A.,Chiaramonte, Jonathan,Piazza, Sabrina M.,Reynolds, Michael R.,Trant, John F.,Xu, Peihan
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supporting information
p. 19224 - 19227
(2021/11/09)
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- COMPOSITIONS AND METHODS OF TREATING CANCER AND INFECTIONS USING BACTERIOPHAGE AND ITS MUTANTS
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Provided herein are vaccine composition comprising an antigen conjugated to a capsid, wherein the capsid comprises wild type or native sequence. Provided herein are also vaccine composition comprising an antigen conjugated to a capsid, wherein said capsid comprises at least one mutation, such as a non-natural mutation. Such compositions are useful in the treatment and prevention of pathogenic infections, inflammatory diseases, and neurodegenerative disease, and cancer, among others.
- -
-
Page/Page column 8; 65
(2019/03/17)
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- Design and synthesis of trivalent Tn glycoconjugate polymers by nitroxide-mediated polymerization
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A new synthetic method for preparing Tn glycoconjugate polymers, containing tumor-associated carbohydrate antigens, by controlled living radical polymerization is reported. To mimic the authentic structures of Tn glycopeptide antigens and to explore the controlled living radical polymerization, three tumor-associated carbohydrate antigens (GalNAc, GalNAcα1-O-Ser, and GalNAcα1-O-Thr) were attached to a styrene-type monomer through a diethylene glycol spacer. Under nitroxide-mediated polymerization, controlled living radical polymerization proceeded to afford defined glycopeptide polymers with different Tn densities and compositions. The polydispersity index (PDI) and molecular weights were increased and conversions were decreased upon increasing the concentration of Tn glycoconjugate monomers. The resulting Tn glycoconjugate polymers were characterized by NMR and IR. The spectral data indicate that the Tn glycoconjugate moiety did attach to the polymer chain and Tn glycoconjugate density could be adjusted through the nitroxide-mediated polymerization conditions. The number of Tn units containing in the polymer chains could be estimated by NMR integration. This synthetic approach provides a new and efficient tool for constructing novel Tn glycoconjugate polymers.
- Liu, Si-Xian,Tsai, Yun-Tzu,Lin, Yu-Tung,Li, Jia-Yue,Chang, Che-Chien
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- Utilization of bench-stable and readily available nickel(II) triflate for access to 1,2-cis-2-aminoglycosides
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The utilization of substoichiometric amounts of commercially available nickel(II) triflate as an activator in the reagent-controlled glycosylation reaction for the stereoselective construction of biologically relevant targets containing 1,2-cis-2-amino gl
- Sletten, Eric T.,Ramadugu, Sai Kumar,Nguyen, Hien M.
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p. 195 - 207
(2016/11/23)
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- Fluorenylmethoxycarbonyl-Protected O-Glycosyl-N-methyl Amino Acids: Building Blocks for the Synthesis of Conformationally Tuned Glycopeptide Antigens
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Peptide antibiotics often contain N-methylated amino acids. These N-methylamino components enhance the metabolic stability and strongly influence the conformational behavior of these peptide drugs. N-Methyl-O-glycosyl amino acids, in particular, threonine and serine derivatives, are unknown so far. Fmoc-protected N-methyl-O-glycosyl-threonine and -serine building blocks, including sialyl TN antigens, have been synthesized for the first time by converting the Fmoc-protected O-glycosyl amino acids or their tert-butyl esters into the corresponding oxazolidinones followed by reductive ring-opening. These new components are considered interesting for the construction of modified mucin glycopeptide anti-tumor vaccines with extended biological half-life. Fmoc-protected building blocks of N-methylated O-glycosyl amino acids, inaccessible so far, have been synthesized by the conversion of Fmoc-O-glycosyl amino acids into the corresponding oxazolidines followed by reductive ring-opening. These N-methyl-O-glycosyl amino acids could be used in solid-phase glycopeptide syntheses under particular conditions.
- Buba, Annette E.,L?we, Holger,Kunz, Horst
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supporting information
p. 5764 - 5774
(2015/09/15)
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- One-pot multi-enzyme (OPME) chemoenzymatic synthesis of sialyl-Tn-MUC1 and sialyl-T-MUC1 glycopeptides containing natural or non-natural sialic acid
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A series of STn-MUC1 and ST-MUC1 glycopeptides containing naturally occurring and non-natural sialic acids have been chemoenzymatically synthesized from Tn-MUC1 glycopeptide using one-pot multienzyme (OPME) approaches. In situ generation of the sialyltran
- Malekan, Hamed,Fung, Gabriel,Thon, Vireak,Khedri, Zahra,Yu, Hai,Qu, Jingyao,Li, Yanhong,Ding, Li,Lam, Kit S.,Chen, Xi
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p. 4778 - 4785
(2013/08/23)
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- Synthesis of Tn/T antigen MUC1 glycopeptide BSA conjugates and their evaluation as vaccines
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The tumor-associated mucin MUC1 over-expressed in most epithelial tumor tissues is considered a promising target for immunotherapy. The extracellular part of MUC1 contains a domain of numerous tandem repeats of the amino acid sequence HGVTSAPDTRPAPGSTAPPA, including five potential O-glycosylation sites. In this study, T9 and S15 have been chosen as the positions of glycosylation. The glycopeptides N-terminally equipped with a triethylene glycol spacer were synthesized by microwave-assisted Fmoc solid-phase peptide synthesis. After detachment from the resin and deprotection, the MUC1 glycopeptides were conjugated to bovine serum albumin (BSA). To evaluate the immunological properties, balb/c mice were immunized with these BSA vaccines. Copyright
- Cai, Hui,Huang, Zhi-Hua,Shi, Lei,Zou, Peng,Zhao, Yu-Fen,Kunz, Horst,Li, Yan-Mei
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p. 3685 - 3689
(2011/09/21)
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- Microwave-assisted synthesis of fluorescein-labelled GalNAcα1-O-Ser/ Thr (Tn) glycopeptides as immunological probes
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Fluorescently labelled glycopeptides containing GalNAcα1-O-Ser/Thr residues provide valuable immunological probes for the development of cancer vaccines. The microwave-assisted automated Fmoc solid-phase synthesis of a series of 5(6)-carboxyfluorescein-labelled GalNAcα1-O-Ser/Thr peptides is described. Lys(Dde)-Gly-Wang polystyrene resin was elongated using Fmoc SPPS with incorporation of several GalNAcα1-O-Ser/Thr residues. Deprotection of the Lys(Dde) then allowed attachment of the 5(6)-carboxyfluorescein label. The synthetic methodology described is flexible and suitably robust enabling the incorporation of three contiguous GalNAcα1-O-Ser residues into the peptide chain. Georg Thieme Verlag Stuttgart · New York.
- Lee, Dong Jun,Harris, Paul W.R.,Kowalczyk, Renata,Dunbar, P. Rod,Brimble, Margaret A.
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experimental part
p. 763 - 769
(2010/10/01)
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- Chemoenzymatic synthesis of glycosylphosphatidylinositol-anchored glycopeptides
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MUC1 glycopeptide was efficiently coupled to glycosylphosphatidylinositol (GPI) derivatives by sortase A (SrtA), verifying that SrtA can accept sterically hindered glycopeptide as substrate for ligation with GPIs. This work has established a practical method for the chemoenzymatic synthesis of GPI-linked glycopeptides.
- Wu, Zhimeng,Guo, Xueqing,Guo, Zhongwu
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supporting information; experimental part
p. 5773 - 5774
(2010/10/19)
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- Cowpea mosaic virus capsid: A promising carrier for the development of carbohydrate based antitumor vaccines
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Immunotherapy targeting tumor cell surface carbohydrates is a promising approach for cancer treatment. However, the low immunogenecity of carbohydrates presents a formidable challenge. We describe here the enhancement of carbohydrate immunogenicity by an ordered display on the surface of the cowpea mosaic virus (CPMV) capsid. The Tn glycan, which is overexpressed on numerous cancer cell surfaces, was selected as the model antigen for our study. Previously it has been shown that it is difficult to induce a strong T cell-dependent immune response against the monomeric form of Tn presented in several ways on differeent carriers. In this study, we first synthesized Tn antigens derivatized with either a maleimide or a bromoacetamide moiety that was conjugated selectively to a cysteine mutant of CPMV. The glycoconjugate was then injected into mice and pre- and post-immune antibody levels in the mice sera were measured by enzyme-linked immunosorbant assays. High total antibody titers and, more importantly, high IgG titers specific for Tn were obtained in the post-immune day 35 serum, suggesting the induction of T cell-dependent antibody isotype switching by the glycoconjugate. The antibodies generated were able to recognize Tn antigens presented in their native conformations on the surfaces of both MCF-7 breast cancer cells and the multidrug resistant breast cancer cell line NCI-ADR RES. These results suggest that the CPMV capsid can greatly enhance the immunogenicity of weak antigens such as Tn and this can provide a promising tool for the development of carbohydrate based anti-cancer vaccines.
- Miermont, Adeline,Barnhill, Hannah,Strable, Erica,Lu, Xiaowei,Wall, Katherine A.,Wang, Qian,Finn,Huang, Xuefei
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experimental part
p. 4939 - 4947
(2009/04/21)
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- Methyl esters: an alternative protecting group for the synthesis of O-glycosyl amino acid building blocks
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The glycosyl amino acids α-GalNAc-Ser and α-GalNAc-Thr are fundamental building blocks for glycopeptide synthesis, Schmidt's synthesis method often being chosen for this purpose. Methyl esters used as orthogonal carboxylic acid protecting group in this pr
- Mayato, Carlos,Dorta, Rosa L.,Vázquez, Jesús T.
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p. 1396 - 1398
(2008/09/18)
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- Saccharide-induced peptide conformation in glycopeptides of the recognition region of LI-cadherin
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(Chemical Equation Presented) The type and the size of the saccharide side chain can affect the conformation of the peptide portion of glycopeptides. This was shown by conformational analysis in solution for a series of glycoundecapeptides with systematic
- Kuhn, Axel,Kunz, Horst
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p. 454 - 458
(2008/02/04)
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- Syntheses of α-d-galactosamine neoglycolipids
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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.
- Laurent, Nicolas,Lafont, Dominique,Boullanger, Paul
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p. 823 - 835
(2007/10/03)
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- Synthesis of Peptide and Glycopeptide Partial Structures of the Homophilic Recognition Domain of Epithelial Cadherin
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Peptide and glycopeptide sequences of the homophilic recognition site of epithelial cadherin (E-cadherin) were synthesized by solid-phase technique based on acid-sensitive Wang anchor according to Fmoc strategy. Fmoc serine building blocks with TN/s
- Reipen, Tanja,Kunz, Horst
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p. 2487 - 2502
(2007/10/03)
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- Solid-phase synthesis of O-linked glycopeptide analogues of enkephalin
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The synthesis of 18 N-α-FMOC-amino acid glycosides for solid-phase glycopeptide assembly is reported. The glycosides were synthesized either from the corresponding O'Donnell Schiff bases or from N-α-FMOC-amino protected serine or threonine and the appropriate glycosyl bromide using Hanessian's modification of the Koenigs-Knorr reaction. Reaction rates of D-glycosyl bromides (e.g., acetobromoglucose) with the L- and D-forms of serine and threonine are distinctly different and can be rationalized in terms of the steric interactions within the two types of diastereomeric transition states for the D/L and D/D reactant pairs. The N-α-FMOC-protected glycosides [monosaccharides Xyl, Glc, Gal, Man, GlcNAc, and GalNAc; disaccharides Gal-β(1-4)-Glc (lactose), Glc-β(1-4)-Glc (cellobiose), and Gal-α(1-6)-Glc (melibiose)] were incorporated into 22 enkephalin glycopeptide analogues. These peptide opiates bearing the pharmacophore H-Tyr-c[DCys-Gly-Phe-DCys]- were designed to probe the significance of the glycoside moiety and the carbohydrate-peptide linkage region in blood-brain barrier (BBB) transport, opiate receptor binding, and analgesia.
- Mitchell,Pratt,Hruby,Polt
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p. 2327 - 2342
(2007/10/03)
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- Nitroglycal concatenation: A broadly applicable and efficient approach to the synthesis of complex O-glycans
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Base-catalyzed glycosylations provide the basis for a new and general entry to the synthesis of mucin-type O-glycans. The desired α-linked 2-acetamidoglycosyl amino acids B are accessible selectively starting from glycals of type A. Fmoc = 9-fluorenylmeth
- Winterfeld, Gottfried A.,Schmidt, Richard R.
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p. 2654 - 2657
(2007/10/03)
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- A novel and efficient route towards α-GalNAc-Ser and α-GalNAc-Thr building blocks for glycopeptide synthesis
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Michael addition of serine and threonine derivatives 4a-4c to 3,4,6-tri- O-benzyl-2-nitro-D-galactal (1) afforded the corresponding 2-deoxy-2-nitro- α-D-galactopyranosides 5a-5c in good yield and stereoselectivity. 2-deoxy-2- nitroglycosides 5a and 5b were reduced to the 2-acetamido compounds by platinized Raney nickel T4. Manipulation of the protecting groups afforded known N-Fmoc-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-α-D- galactopyranosyl)-L-serine (8a) and -threonine (8b), valuable building blocks for O-glycopeptide synthesis.
- Winterfeld, Gottfried A.,Ito, Yukishige,Ogawa, Tomoya,Schmidt, Richard R.
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p. 1167 - 1171
(2007/10/03)
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- Synthetic and immunological studies on clustered modes of mucin-related Tn and TF O-linked antigens: The preparation of a glycopeptide-based vaccine for clinical trials against prostate cancer
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The syntheses of two tumor-associated carbohydrate antigens, Tn and TF, have been achieved using glycal assembly and cassette methodologies. These synthetic antigens were subsequently clustered (c) and immunoconjugated to a carrier protein (KLH or BSA) or
- Kuduk, Scott D.,Schwarz, Jacob B.,Chen, Xiao-Tao,Glunz, Peter W.,Sames, Dalibor,Ragupathi, Govindaswami,Livingston, Philip O.,Danishefsky, Samuel J.
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p. 12474 - 12485
(2007/10/03)
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- Chemoselective removal of protecting groups from O-glycosyl amino acid and peptide (methoxyethoxy)ethyl esters using lipases and papain
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The selective C-terminal deprotection of O-glycopeptide (methoxyethoxy)ethyl esters is achieved under mild conditions (pH 6.6, 37°C) by enzymatic hydrolysis using papain or lipase M from Mucor javanicus to give building blocks useful for chain-extending glycopeptide synthesis. On the other hand, the selective removal of acetyl protecting groups from the saccharide portion of glycopeptides is accomplished by alternative enzymatic hydrolysis with lipase WG from wheat germ to furnish model substrates for enzymatic glycosyl transfer reactions in order to extend the carbohydrate side chain of these conjugates.
- Eberling, Joerg,Braun, Peter,Kowalczyk, Danuta,Schultz, Michael,Kunz, Horst
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p. 2638 - 2646
(2007/10/03)
-
- Piperidine is preferred to morpholine for Fmoc cleavage in solid phase glycopeptide synthesis as exemplified by preparation of glycopeptides related to HIV gp120 and mucins
-
Protected derivatives of the Tn antigens [Fmoc-Ser/Thr(Ac3GalNAcα)-OH, compounds 5 and 8] have been prepared by glycosylation of Fmoc-Ser/Thr- OAllyl with 3,4,6-tri-O-acetyl-2-azido-2-deoxy-D-galactopyranosyl chloride (2), followed by conversion of the azido group to an acetamide and deallylation. The derivatives 5 and 8 were used for solid phase synthesis of glycopeptides related to HIV gp120 and mucins. In these syntheses piperidine was found to give efficient Fmoc removal whereas deprotection with morpholine was slow and incomplete for some steps. In contrast to previous concerns β- elimination and epimerization of glycopeptide stereocenters was not encountered when piperidine was used for Fmoc deprotection. However, it was found that for glycopeptides which contained cysteine residues, de-O- acetylation with methanolic ammonia had to be performed before side-chain deprotection and cleavage from the solid phase.
- Vuljanic, Tatjana,Bergquist, Karl-Erik,Clausen, Henrik,Roy, Sarbani,Kihlberg, Jan
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p. 7983 - 8000
(2007/10/03)
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- Stereoselective synthesis of O-serinyl/threoninyl-2-acetamido-2-deoxy-α- or β-glycosides
-
General glycosidation methodology has been developed which can selectively provide 2-acetamido-2-deoxy-α- or β-glycosides of β-hydroxy-α-amino acid derivatives [glucopyranoside-(8, 43), galactopyranoside- (9, 13), mannopyranoside- (10), lactoside analogs (11, 38) and 3-O-β-galactopyranosyl-mannopyranoside (12)] stereoselectively in excellent yield from the highly nucleophilic α-imino esters (Schiff bases) of L-serine and L-threonine. Various glycosides were converted via their amino and acetamido derivatives to Fmoc-protected serinyl- or threoninyl-glycosides (24-28, 37, 41, 46) which are all suitable building blocks for the solid-phase synthesis of O-glycopeptides. Complete 1H- and 13C-NMR data are provided for all compounds.
- Szabo,Ramza,Langdon,Polt
-
-
- Synthesis of O-Glycopeptides of the N-Terminus of Interleukin-2
-
The synthesis of a number of O-glycopeptides with different lengths of chain of between 3 and 10 amino acids that are part of the N-terminus of interleukin-2 is described.The key intermediate for the syntheses is O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-α-D-galactopyranosyl)-N-(9-fluorenylmethoxycarbonyl)-L-threonine tert-butyl ester (9b), to which suitably protected amino acids and peptides can be added C- or N-terminally. 9b is an element, which allows to insert glycosylated hydroxyamino acids with the Fmoc/tBu combination of protective groups, which are approved in peptide synthesis.During the syntheses peptide blocks are used.With the deprotected products it is possible to test the structural specificity of glycosyl transferases.
- Paulsen, Hans,Adermann, Knut
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p. 751 - 770
(2007/10/02)
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- Synthesis of Varied O-Glycopeptides of Interleukin-2
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A number of O-glycopeptides, which are varieties of the N-terminus of interleukin-2, is synthesized.In these compounds a modified amino acid sequence, an altered glycosylation site, or a higher level of glycosylation is achieved.The syntheses are carried
- Paulsen, Hans,Adermann, Knut
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p. 771 - 780
(2007/10/02)
-