Comparison of glycosyl donors: a supramer approach.

The development of new methods for chemical glycosylation commonly includes comparison of various glycosyl donors. An attempted comparison of chemical properties of two sialic acid-based thioglycoside glycosyl donors, differing only in the substituent at O-9 (trifluoroacetyl vs chloroacetyl), at different concentrations (0.05 and 0.15 mol·L-1) led to mutually excluding conclusions concerning their relative reactivity and selectivity, which prevented us from revealing a possible influence of remote protective groups at O-9 on glycosylation outcome. According to the results of the supramer analysis of the reaction solutions, this issue might be related to the formation of supramers of glycosyl donors differing in structure hence chemical properties. These results seem to imply that comparison of chemical properties of different glycosyl donors may not be as simple and straightforward as it is usually considered.

Synthesis of photoactivable oligosaccharide derivatives from 1,2-cyclic carbamate building blocks and study of their interaction with carbohydrate-binding proteins.

Despite the broad occurrence of carbohydrate-protein interactions in biology, the low binding affinities of such interactions hamper the characterization of carbohydrate binding sites in the absence of three-dimensional structural models. To allow the identification of proteins interacting with specific carbohydrate epitopes, we have developed new photoactivable oligosaccharide probes. Oligosaccharides containing the 1,2-cyclic carbamate group were attached to building blocks with a primary amino group to yield the corresponding urea derivatives. Cyclic carbamates of lactose, and 3- and 2'-fucosyl lactose, were used for the conjugation with building blocks containing photoactivable diazirine, benzophenone or aryl azido groups. The resulting oligosaccharide derivatives were tested for binding to Erythrina cristagalli lectin (ECL), Aleuria aurantia lectin (AAL) and Ulex europaeus agglutinin-I (UEA I). We found that ligands containing an aryl azido photoactivable group were successfully attached to lectins. The photoactivation reaction preserved lectin integrity, as no sign of protein degradation was visible. Mass spectrometric analysis confirmed the covalent binding of between one to three oligosaccharide probes, which matched with the expected carbohydrate-binding properties of the lectins tested. The conjugation of cyclic carbamate-derivatized oligosaccharides with photoactivable aryl azido groups thus represents a convenient approach to study protein-carbohydrate interactions.

Preparation and in vivo evaluation of glyco-gold nanoparticles carrying synthetic mycobacterial hexaarabinofuranoside.

A number of bacterial glycans are specific markers for the detection and the serological identification of microorganisms and are also widely used as antigenic components of vaccines. The use of gold nanoparticles as carriers for glyco-epitopes is becoming an important alternative to the traditional conjugation with proteins and synthetic polymers. In this study, we aimed to prepare and evaluate in vivo glyco-gold nanoparticles (glyco-GNPs) bearing the terminal-branched hexaarabinofuranoside fragment (Ara6) of arabinan domains of lipoarabinomannan and arabinogalactan, which are principal polysaccharides of the cell wall of Mycobacterium tuberculosis, the causative agent of tuberculosis. In particular, we were interested whether the antibodies generated against Ara6-GNPs would recognize the natural saccharides on the cell surface of different mycobacterial strains. Two synthetic Ara6 glycosides with amino-functionalized spacer aglycons differing in length and hydrophilicity were directly conjugated with spherical gold nanoparticles (d = 15 nm) to give two sets of glyco-GNPs, which were used for the immunization of rabbits. Dot assays revealed cross-reactions between the two obtained antisera with the hexaarabinofuranoside and the 2-aminoethyl aglycon used for the preparation of glyco-GNPs. Both antisera contained high titers of antibodies specific for Mycobacteria as shown by enzyme-linked immunosorbent assay using M. bovis and M. smegmatis cells as antigens while there was only a weak response to M. phlei cells and no interaction with E. coli cells. The results obtained suggest that glyco-GNPs are promising agents for the generation of anti-mycobacterial antibodies.

Arabinofuranose 1,2,5-orthobenzoate as a single precursor of linear α(1 → 5)-linked oligoarabinofuranosides.

Selectively protected mono-, di- and trisaccharide thioglycoside building blocks with unprotected primary hydroxy group at the non-reducing end, available in only one step from 3-O-benzoyl ß-d-arabinofuranose 1,2,5-orthobenzoate, were used in the synthesis of linear α(1 â†’ 5)-linked oligoarabinofuranosides up to octasaccharide. The obtained oligosaccharides contain 4-(2-chloroethoxy)phenyl (CEP) or 4-(2-azidoethoxy)phenyl (AEP) pre-spacer aglycons that allow preparation of neoglycoconjugates.

Stereoselective sialylation with O-trifluoroacetylated thiosialosides: hydrogen bonding involved?

A series of novel sialyl donors containing O-trifluoroacetyl (TFA) groups at various positions was synthesized. The choice of protecting groups in sialyl donors was based on hypothesis that variations in ability of different acyl groups to act as hydrogen bond acceptors would influence the supramolecular structure of reaction mixture (solution structure), hence the outcome of sialylation. These glycosyl donors were examined in the model glycosylation of the primary hydroxyl group of 1,2:3,4-di-O-isopropylidene-α-D-galactopyranose in comparison with sialyl donors without O-TFA groups. The presence of O-TFA groups in a sialyl donor strongly affected the outcome of sialylation. Several sialyl donors studied showed promising results: yields of disaccharides can be as high as 86% as can be the stereoselectivities (α/ß up to 15:1). The results obtained suggest that varying acyl O-protecting groups in sialyl donor may result in dramatic changes in the outcome of sialylation although further studies are required to dissect the influence of intermolecular hydrogen bonding and intramolecular substituent effects related to variations of electron-withdrawing properties of different acyl groups.

Bimodal concentration-dependent reactivity pattern of a glycosyl donor: Is the solution structure involved?

Changes in concentration (0.001-0.1 M) of an arabinofuranosyl donor (1) have been shown to modulate the temperature T at which activation of 1 occurs (from -23 °C to +7 °C), the reaction time (from 1.5 h to 3 days) and the yield of the disaccharide formed (from 14% to 82%). At concentrations exceeding 0.01 M, these parameters, as well as the specific optical rotation of the solution of 1, virtually do not depend on concentration suggesting formation of reacting species (supramers) of glycosyl donor with similar structures, hence reactivities, but considerably different from those formed in more dilute solutions. The found critical concentration (0.01 M) separates two concentration ranges of reaction solutions corresponding to two types of solution structure that are featured by the presence of fundamentally different supramers of glycosyl donor, which have distinct chemical properties. These results allow a fresh look at the problems of reactivity of chemical compounds and selectivity of the reactions in which they participate.

Rapid synthesis of linear homologous oligoarabinofuranosides related to mycobacterial lipoarabinomannan and a neoglycoconjugate thereof.

Rapid and simple synthesis of oligosaccharides related to one of the terminal motifs of mycobacterial lipoarabinomannan is described. An array of homologous linear α(1 â†’ 5)-linked oligoarabinofuranosides with 4-(2-chloroethoxy)phenyl aglycon and selectively unprotected 5-OH group at the non-reducing end was obtained by oligomerization of 3-O-benzoyl ß-D-arabinofuranose 1,2,5-orthobenzoate. Subsequent introduction of ß(1 â†’ 2)-linked arabinofuranose disaccharide moiety by step-wise glycosylation furnished the target oligosaccharides which were conjugated with bovine serum albumin.

A Concise Synthesis of the Repeating Unit of Capsular Polysaccharide Staphylococcus aureus Type 8.

The first synthesis of the repeating unit of S. aureus capsular polysaccharide type 8 is described. The repeating unit is an unusual trisaccharide sequence of three uncommon sugars, all connected via 1,2-cis linkages. The synthetic trisaccharide was equipped with capping methyl groups at the points of propagation of the polysaccharide sequence.

The use of O-trifluoroacetyl protection and profound influence of the nature of glycosyl acceptor in benzyl-free arabinofuranosylation.

The influence of O-trifluoroacetyl (TFA) groups at different positions of thioglycoside glycosyl donors on stereoselectivity of α-arabinofuranosylation leading to corresponding disaccharides was studied. It was shown that TFA group in thioglycoside glycosyl donors, when combined with 2-O-(triisopropylsilyl) (TIPS) non-participating group, may be regarded as an electron-withdrawing protecting group that may enhance 1,2-cis-selectivity in arabinofuranosylation, the results strongly depending on the nature of glycosyl acceptor. The reactivities of the glycosyl donors were compared with those of a similar thioglycoside with O-pentafluoropropionyl groups and the known phenyl 3,5-O-(di-tert-butylsilylene)-1-thio-α-d-arabinofuranosides with 2-O-TIPS and 2-O-benzyl groups. The 'matching' in the donor-acceptor combination was found to be critical for achieving both high reactivity of glycosyl donor and ß-stereoselectivity of arabinofuranosylation. The use of glycosyl donors with TFA and silyl protection may be useful in the realization of the benzyl-free approach to oligoarabinofuranosides with azido group in aglycon-convenient building blocks for the preparation of neoglycoconjugates.

Synthesis of branched arabinofuranose pentasaccharide fragment of mycobacterial arabinans as 2-azidoethyl glycoside.

Branched arabinofuranose pentasaccharide with 2-azidoethyl aglycon was prepared for the first time by [3+1+1] bis-(1,2-cis)-glycosylation of trisaccharide diol with silyl-protected thioglycoside glycosyl donor. The presence of 2-azidoethyl aglycon would enable the preparation of neoglycoconjugates using the click chemistry approaches.

Ring opening of acylated ß-d-arabinofuranose 1,2,5-orthobenzoates with nucleophiles allows access to novel selectively-protected arabinofuranose building blocks.

ß-d-Arabinofuranose 1,2,5-orthobenzoates with 3-O-acetyl, 3-O-benzoyl, and 3-O-chloroacetyl groups were prepared in an efficient manner starting from readily available crystalline methyl 2,3,5-tri-O-benzoyl-α-d-arabinofuranoside, and ring-opening reactions of these compounds with O- and S-nucleophiles were studied. Optimized conditions leading to the formation of the respective monosaccharide adducts (up to 96% isolated yields) and to α-(1→5)-linked disaccharide thioglycosides with 5'-OH unprotected (up to 30% isolated yields) were found. Basing on these results, a novel approach for effective differentiation of 3,5-diol system and 2-hydroxy group in arabinofuranose thioglycosides was proposed. The selectively protected derivatives prepared are valuable building blocks for the assembly of linear and branched oligoarabinofuranosides.