Stereoselective synthesis of 1,2-cis- and 2-deoxyglycofuranosyl azides from glycosyl halides

Protected 1,2-cis glycofuranosyl azides with alpha-D-ribo, beta-D-arabino and 2-deoxy-2-fluoro-beta-D-arabino configurations were efficiently prepared from the appropriate 1,2-trans glycosyl halides bearing non-participating 0-2 substituent by inversion with sodium azide under phase transfer catalytic conditions (80-85% yields, 90-96% de). The same method failed to result in sufficiently good beta-selectivity starting from 2-deoxy-3,5-di-O-(p-toluoyl)-alpha-D-ery-thro-pentofuranosyl chloride (5alpha) (40% de). The selectivity in favour of the protected 2-deoxy-beta-D-erythro-pentofura-nosyl azides was greatly improved (74-80% de) by treating 5alpha and its p-chlorobenzoyl analog 6alpha with cesium or potassium azide in dimethylsulfoxide at room temperature (83-85% yields).     

N-Glycosidation of D-arabino-hex-2-ulosonic acid

Two approaches to N-functionalized D-arabino-hex-2-ulosonic acid derivatives were established by nucleophilic substitution of methyl (3,4,5-tri-O-acetyl-beta-D-arabino-hex-2-ulopyranosyl)onate bromide (1). Reaction of 1 with amino compounds in the presence of mercury(II) cyanide led to the 2,3-cis configured beta-D-arabino N-glycosides. On the other hand, the reaction of bromide 1 with azide, followed by catalytic hydrogenation led to 2,3-trans alpha-D-arabino glycosyl amine methyl 3,4,5-tri-O-acetyl-2-amino-alpha-D-arabino-hex-2-ulopyranosonate, which was easily rearranged to the thermodynamically more stable beta-D-arabino N-acetyl derivative methyl 4,5-di-O-acetyl-2-acetylamino-3-hydroxy-beta-D-arabino-hex-2-ulopyranosonate. The assignment of configuration of the tertiary anomeric centre and conformation of all products was based on 1H NMR H,H coupling constants and NOE difference experiments.     

Synthesis and characterization of some anomeric pairs of per-O-acetylated aldohexopyranosyl cyanides (per-O-acetylated 2,6-anhydroheptononitriles). On the reaction of per-O-acetylaldohexopyranosyl bromides with mercuric cyanide in nitromethane

The synthesis and characterization of the anomeric pairs of the per-O-acetylaldohexopyranosyl cyanides of D-galactose, L-fucose, D-glucose, and D-mannose, as well as of 3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-beta-D-glucopyranosyl cyanide, are described. Cyanation of the readily available, per-O-acetylaldohexopyranosyl bromides with mercuric cyanide in nitromethane, and subsequent purification, gave the corresponding, crystalline glycosyl cyanides with a high degree of 1,2-trans stereoselectivity. Thus, per-O-acetylated aldohexopyranosyl cyanides of the 1,2-trans configuration were obtained in yields ranging from 20 to 79%, whereas the corresponding 1,2-cis anomers were obtained in yields of less than or equal to 8.4%, the ratios of the 1,2-trans:1,2-cis anomers so prepared being greater than or equal to 8.5:1. The principal by-products of these irreversible, cyanation reactions were the per-O-acetylated 1,2-O-[1-(exo- and endo-cyano)ethylidene]aldohexopyranoses, obtained in yields of up to 40%. The structural assignments of the per-O-acetylaldohexopyranosyl cyanides were unequivocally established by elemental analysis, chemical transformation, vibrational spectroscopy, and 13C- and 1H-nuclear magnetic resonance spectroscopy. Correlations between the physical properties and the anomeric configurations of these C-aldohexopyranosyl compounds are described.     

Syntheses of amphiphilic glycosylamides from glycosyl azides without transient reduction to glycosylamines

Protected glycosyl azides react with acyl chlorides in the presence of triphenylphosphine to afford glycosylamides in high yields, at room temperature. Starting from the beta-glycosyl azides, the reaction is highly stereoselective and occurs with retention of configuration, whereas the alpha-azido anomers display a lower stereoselectivity giving rise to alpha/beta mixtures of glycosylamides. The reaction was applied to several monosaccharidic azides and to lactosyl azide with various acyl chlorides; it was shown to be of general use for preparing 1,2-trans beta-glycosylamides.     

DFT studies of the role of C-2-O-2 bond rotation in neighboring-group glycosylation reactions

Although the synthetic utility of the 1,2-trans relationship of the products of neighboring group participation is well established, it is still common to find glycosylation reactions where the stereochemical purity of the products is not 100%. As part of an ongoing series of density functional theory (DFT) studies of the factors that affect glycosylation reactions which are aimed at allowing synthetic chemists to achieve such selectivities, the structures of four oxacarbenium ions and eight methanol complexes of these ions were optimized for the prototypical ions 2-O-acetyl-3,4,6-tri-O-methyl-D-gluco- (1) and mannopyranos-1-yl (2). These studies corroborate the two-conformer hypothesis and further demonstrate that glycopyranosyl oxacarbenium ions exhibit facial selectivity that depends on, besides the inherent steric and Van der Waals effects, the conformational effect associated with the change from sp(2) to sp(3) hybridization at C-1 during nucleophilic attack and H-bonding between the incoming nucleophile and the electronegative atoms of the electrophile. Further studies based on systematic C-2-O-2 bond rotations found TSs that connect the monocyclic ions with the bicyclic ions associated with neighboring-group participation. It was also possible to find two TSs that connect nucleophilic attack at C-1 with C-2-O-2 bond rotation ultimately leading to 1,2-trans O-glycosides, that is, the probable TS that determines the stereochemistry of neighboring-group participation. Both of these TSs exhibit intramolecular H-bonding, which is considered the first step in proton transfer. It is further hypothesized that this coupling of proton transfer and nucleophilic attack is integral to glycosylation. It is further hypothesized that in many cases analogous intermolecular H-bonding is also favorable with the most likely acceptor the anion that is ion-paired to the oxacarbenium ion. These general features are found for both 1 and 2, but characteristic features of each isomer are found that provide further insights into the origins of stereoselectivity.     

Transglucosidation of methyl and ethyl D-glucofuranosides by alcoholysis

The acid catalyzed ethanolysis of methyl 5-O-methyl-alpha- and -beta-D-glucofuranoside and the analogous methanolysis of ethyl 5-O-methyl-alpha- and -beta-D-glucofuranoside have been investigated. For all four reactions, the primarily formed transglycosylation product was a single glycoside that had the opposite anomeric configuration to the starting material. This strongly indicates that a D-glucose methyl ethyl acetal is first formed and is then ring closed by a nucleophilic attack by HO-4, giving either the starting material or a transglycosylation product with the opposite anomeric configuration. Low percentages of the methyl ethyl acetals and of dimethyl acetals were also observed in the reaction product during the methanolysis reactions.     

Lifetimes and reactivities of some 1,2-didehydroazepines commonly used in photoaffinity labeling experiments in aqueous solutions

The reactive 1,2-didehydroazepine (cyclic ketenimine) intermediates produced upon photolysis of phenyl azide, 3-hydroxyphenyl azide, 3-methoxyphenyl azide, and 3-nitrophenyl azide in water and in HEPES buffer were studied by laser flash photolysis techniques with UV-vis detection of the transient intermediates. The lifetimes of the 1,2-didehydroazepines were obtained along with the absolute rate constants of their reactions with typical amino acids, nucleosides, and other simple reagents present in a biochemical milieu. The nitro substituent greatly accelerates the bimolecular reactions of the cyclic ketenimines, and the 3-methoxy group greatly decelerates the absolute reactivity of 1,2-didehydroazepines. The intermediate produced by photolysis of 3-hydroxyphenyl azide is much more reactive than the intermediate produced by photolysis of 3-methoxyphenyl azide. We propose that the hydroxyl-substituted 1,2-didehydoazepines rapidly (<10 micros) tautomerize in water to form azepinones and much more rapidly than the corresponding 3-methoxy-substituted cyclic ketenimines undergo hydrolysis. Azepinones react more rapidly with nucleophiles than do methoxy-substituted 1,2-didehydroazepines and are the active species present upon the photolysis of 3-hydroxyphenyl azide in aqueous solution.     

The hydrolysis of glycosyl fluorides by glycosidases. Determination of the anomeric configuration of the products of glycosidase action

The enzymic hydrolysis of glycosyl fluorides is conveniently followed by using a pH-stat. Reactions involving glucosyl or galactosyl fluorides can also be followed by using glucose oxidase or galactose oxidase respectively. The pH-stat allows the rapid assay of intestinal alpha-glucosidase in crude homogenates. Use of glycosyl fluorides as substrates for glycosidases facilitates the polarimetric or g.l.c. determination of the anomeric nature of the initial product of hydrolysis. Hydrolysis by fungal amyloglucosidase proceeds with inversion of configuration whereas that by yeast and rat intestinal alpha-glucosidase, coffee-bean alpha-galactosidase and almond emulsin beta-glucosidase proceeds with retention of configuration. beta-d-Glucopyranosyl azide was not a detectable substrate for almond emulsin beta-d-glucosidase.     

Recent studies on reaction pathways and applications of sugar orthoesters in synthesis of oligosaccharides

Formation of sugar-sugar orthoesters consisting of a fully acylated mono- or disaccharide donor and a partially protected mono- or disaccharide acceptor is regioselective, and rearrangement of the orthoesters via RO-(orthoester)C bond cleavage gives a dioxolenium ion intermediate leading to 1,2-trans glycosidic linkage. The activity order of hydroxyl groups in the partially protected mannose and glucose acceptors is 6-OH>3-OH>2- or 4-OH. The coupling reactions with acylated glycosyl trichloroacetimidates as the donors usually give orthoesters as the intermediates specially when the coupling is carried out at slowed rates, and this is successfully used in regio- and stereoselective syntheses of oligosaccharides. Mannose and rhamnose orthoesters readily undergo O-2-(orthoester)C bond breaking, and this is used for synthesis of alpha-(1-->2)-linked oligosaccharides. (1-->3)-Glucosylation is special since the rearrangement of its sugar orthoester intermediates can occur with either RO-(orthoester)C bond cleavage with formation of the dioxolenium ion leading to 1,2-trans linkage, or C-1-O-1 bond cleavage leading to 1,2-cis linkage, and this is dependent upon the structures of donor and acceptor that compose the orthoester.     

Synthesis and antioxidant activity of a novel class of 4,6-O-protected O-glycosides and their utility in disaccharide synthesis

BF₃·Et₂O-catalysed O-glycosylation of 1,2,3-tri-O-acetyl-4,6-O-butylidene- and ethylidene-β-d-glucopyranose with different aliphatic and aromatic alcohols proceeds for the most part with complete retention of anomeric configuration. Antioxidant activity of O-glycosides shows significant inhibition (IC₅₀ ∼77%). 1,3-Dipolar cycloaddition of terminal alkyne derivatives of O-glycosides with glycosyl azide results in disaccharides.     

Glycosyl fluorides in glycosidations

This short review deals with the recent progress in chemical O-glycosidation and C-glycosylation methods using glycosyl fluorides as glycosyl donors. Pyranosyl and furanosyl fluorides were effectively activated by fluorophilic reagents such as SnCl2-AgClO4, SnCl2-TrClO4, SnCl2-AgOTf, TMSOTf, SiF4, BF3 x Et2O, TiF4, SnF4, Cp2MCl2-AgClO4 (M = Zr or Hf), Cp2ZrCl2-AgBF4, Cp2HfCl2-AgOTf, Bu2Sn(ClO4)2, Me2GaCl, Tf2O, LiClO4, Yb(OTf)3, La(ClO4)3 x nH2O, La(ClO4)3 x nH2O-Sn(OTf)2, Yb-Amberlyst 15, SO4/ZrO2, Nafion-H, montmorillonite K-10, and TrB(C6F5)4 to react with alcohols to give the corresponding O-glycosides in high yields. Furthermore, several types of C-glycosyl compounds, such as aryl, allyl and alkyl C-glycosyl derivatives, were also obtained by the glycosylation using glycosyl fluorides and the corresponding nucleophile with or without a Lewis acid.     

Identification of the catalytic acid base residue of arthrobacter endo-beta-N-acetylglucosaminidase by chemical rescue of an inactive mutant

Arthrobacter endo-beta-N-acetylglucosaminidase (Endo-A), a member of glycoside hydrolase (GH) family 85, catalyses the hydrolysis and transglycosylation of asparagine-linked oligosaccharides of glycoproteins with retention of anomeric configuration. Glu-173 of Endo-A is a catalytically essential amino acid residue, and the corresponding residue is conserved in all GH family 85 members. The catalytic activity of Endo-A E173A mutant was rescued by the addition of sodium azide or sodium formate. Furthermore, the produced beta-glycosyl azide (Man(5)GlcNAc-beta-N(3)) retained the anomeric configuration, indicating that Glu-173 is the catalytic acid-base residue of Endo-A. This is the first identification of the catalytic residue for GH family 85 endo-beta-N-acetylglucosaminidases.     

Convenient preparation of 3,5-anhydro- and 2,5-anhydropentofuranosides, and 5,6-anhydro-D-glucofuranose by use of the Mitsunobu reaction

Methyl 3,5-anhydro-alpha-D-xylofuranosides are obtained by use of the Mitsunobu reaction from 2-O-protected methyl alpha-D-xylofuranosides, which are easily prepared from D-xylose. The Mitsunobu reaction of methyl 3-N-benzylamino-3-deoxy- and 3-azido-3-deoxyarabinofuranosides, which are prepared from the conveniently available methyl 2,3-anhydro-alpha-D- and 2,3-anhydro-alpha-l-lyxofuranosides by nucleophilic ring opening, yields the corresponding methyl 2,5-anhydro-alpha-D- and 2,5-anhydro-alpha-l-arabinofuranosides. Ring opening of 3,5-anhydro-1,2-O-isopropylidene-alpha-D-xylofuranose with azide yields the corresponding 5-azido derivative. The structure and configuration of the products is confirmed by NMR spectroscopy. 5,6-Anhydro-1,2-O-isopropylidene-alpha-D-glucofuranose is formed by the Mitsunobu reaction of 1,2-O-isopropylidene-alpha-D-glucofuranose. Its structure is verified by single-crystal X-ray diffraction analysis.     

The impact of oxacarbenium ion conformers on the stereochemical outcome of glycosylations

The search for stereoselective glycosylation reactions has occupied synthetic carbohydrate chemists for decades. Traditionally, most attention has been focused on controlling the S_N2-like substitution of anomeric leaving groups as highlighted by Lemieux’s in situ anomerization protocol and by the discovery of anomeric triflates as reactive intermediates in the stereoselective formation of β-mannosides. Recently, it has become clear that also S_N1-like reaction pathways can lead to highly selective glycosylation reactions. This review describes some recent examples of stereoselective glycosylations in which oxacarbenium ions are believed to be at the basis of the selectivity. Special attention is paid to the stereodirecting effect of substituents on a pyranosyl ring with an emphasis on the role of the C-5 carboxylate ester in the condensations of mannuronate ester donors.     

Preparation of 2-amino-2-C-glycosyl-acetonitriles from C-glycosyl aldehydes by Strecker reaction

Synthesis of new 2-amino-2-C-d-glycosyl-acetonitriles in a Strecker reaction from various C-glycosyl aldehydes, chiral amines, and HCN was carried out. While aminonitriles from glycal and 2-deoxy-β-d-glycosyl aldehydes were prepared in satisfactory yields, lower yields were obtained with C-glycosyl aldehydes. Strecker reaction with the benzyl-protected 1-C-formyl-d-galactal and S- or R-1-phenylethylamine (S-PEA or R-PEA) yielded predominantly the R-configured C-glycosyl aminoacetonitrile. The direction of the nucleophilic addition appears to be governed by the configuration of the anomeric carbon with β-linked sugars. Since the stereochemistry of the transition state is unknown according to the configuration of the major product a Felkin–Ahn selectivity can be mainly presumed.     

Identification of the catalytic nucleophile of the family 29 alpha-L-fucosidase from Sulfolobus solfataricus via chemical rescue of an inactive mutant

We have recently reported that a functional alpha-L-fucosidase could be expressed by a single insertional mutation in the region of overlap between the ORFs SSO11867 and SSO3060 of the hyperthermophilic Archaeon Sulfolobus solfataricus [Cobucci-Ponzano et al. J. Biol. Chem. (2003) 278, 14622-14631]. This enzyme, belonging to glycoside hydrolase family 29 (GH29), showed micromolar specificity for p-nitrophenyl-alpha-L-fucoside (pNp-Fuc) and promoted transfucosylation reactions by following a reaction mechanism in which the products retained the anomeric configuration of the substrate. The active site residues in GH29 enzymes are still unknown. We describe here the identification of the catalytic nucleophile of the reaction in the alpha-L-fucosidase from S. solfataricus by reactivation with sodium azide of the mutant Asp242Gly that shows a 10(3)-fold activity reduction on pNp-Fuc. The detailed stereochemical analysis of the fucosyl-azide produced by the mutant reactivated on pNp-Fuc revealed its inverted (beta-fucosyl azide) configuration compared with the substrate. This allows for the first time the unambiguous assignment of Asp242, and its homologous residues, as the nucleophilic catalytic residues of GH29 alpha-L-fucosidases. This is the first time that this approach is used for alpha-L-glycosidases, widening the applicability of this method.     

Synthesis of N-glycan oxazolines: donors for endohexosaminidase catalysed glycosylation

Oxazoline mono-, di-, tri- and hexasaccharides, corresponding to the core components of N-linked glycoprotein high mannose glycans, are synthesised as potential glycosyl donors for endohexosaminidase catalysed glycosylation of glycopeptides and glycoprotein remodelling. The crucial beta-D-Manp-(1-->4)-D-GlcpNAc linkage is synthesised via epimerisation of gluco disaccharide substrates by sequential triflation and nucleophilic substitution. Oxazolines are formed directly from the anomeric OPMP protected N-acetyl glucosamine derivatives. Efficient endohexosaminidase catalysed glycosylation of a synthetic beta-D-GlcpNAcAsn glycosyl amino acid is demonstrated with the trisaccharide oxazoline donor.     

Antigenic bacterial polysaccharides. 30. The structure of the polysaccharide chain of lipopolysaccharide of Pseudomonas syringae pv. syringae 281 (serogroup I)

Anomeric methyl 3-O-(D-mannopyranosyl- and L-rhamnopyranosyl)-beta-D-talopyranosides were synthesised by the stereoselective 1,2-cis- and 1,2-trans manno- and rhamnosylation of methyl 2,4,6-tri-O-acetyl-beta-D-talopyranoside, which has been prepared from methyl beta-D-galactopyranoside by a synthetic scheme including conversion of the C2 configuration. From 13C-NMR spectra of the disaccharides obtained the spectral alpha- and beta-effects of O3-glycosylation of talopyranose were determined.     

Specific features of phase transfer catalytic glycosylation of aromatic hydroxy acids

Reactions of peracetylated α-D-glucosaminyl chloride with isomeric hydroxybenzoic and 1-hydroxy-2-naphthoic acids in a solid phase transfer system of potassium carbonate-acetonitrile were studied. It was found that the nature of carboxylic acids, lipophilicity of the phase transfer catalyst, and reaction temperatures affected the reaction composition and product yields. The O-β-glycosyl esters of ortho-hydroxyaromatic acids were first found to form anomeric 1,2-cis derivatives in the presence of potassium carbonate. The structures of the synthesized compounds were confirmed by ¹H NMR spectroscopy. As was shown in vivo experiments, the analgesic activities of glycosyl esters of salicylic acid and peracetylated 2-carboxyphenylglucosaminide were comparable with that of aspirin.     

The two-conformer hypothesis: 2,3,4,6-tetra-O-methyl-mannopyranosyl and -glucopyranosyl oxacarbenium ions

Computational chemistry can give information about the probable conformations of reactive intermediates that are difficult to determine experimentally. Based on density functional theory (DFT) calculations of tetra-O-methyl-D-mannopyranosyl and -glucopyranosyl oxacarbenium ions, two families of conformations, which we call B0 and B1, were found. For the manno configuration, a 4H3 and 3E almost isoenergetic pair were found, whereas for the gluco-configuration a 4H3 and 5S1 pair favouring 4H3 were calculated. These results corroborate earlier results and suggest that this two or more conformer hypothesis is general. Nucleophilic attack on these pairs of cations was modelled with methanol and led to four cases to consider namely alpha- or beta-attack on B0 or B1. The resulting complexes (G0, G1 and F0, F1) demonstrate facial selectivity. The relative energies of these complexes are dominated by intramolecular hydrogen bonding and the conformational consequences to the pyranose ring of changes in the C-5-O-5-C-1-C-2 torsion angle. Constrained variation of the nucleophilic oxygen (methanol) to C-1 distance shows that these ion dipole complexes are the only minima with this constraint.