Anatoly Ya. Chernyak

Synthesis of glycuronamides of amino acids, constituents of microbial polysaccharides and their conversion into neoglycoconjugates of copolymer type

Glycopyranosiduronic acids, amidically linked to amino acids (alanine, serine, threonine, and lysine) were prepared.O-tert-Butyl andNε-tert-butyloxycarbonyl protected amino acidtert-butyl esters were used in ethyl 2-ethoxy-1,2-dihydroquinoline-1-carboxylate promoted condensation with 2-azidoethyl glycosides of glucuronic and galacturonic acid. Reduction of the azido-function followed byN-acryloylation and removal of blocking groups with trifluoroacetic acid gave the target monomers. These were converted into neoglycoconjugates of copolymer type, potentially useful for immunochemical studies.

Synthesis of carbohydrate-amino acid conjugates related to the capsular antigen K54 from Escherichia coli O6:K54:H10 and artificial antigens therefrom.

The disaccharides alpha-L-Rhap-(1----3)-beta-D-GlcpA and beta-D-GlcpA-(1----3)-alpha-L-Rhap bearing amide-linked L-serine or L-threonine, which represent the repeating unit(s) of the capsular polysaccharide from E. coli O6:K54:H10, have been synthesised. O-tert-Butyl-protected amino acid tert-butyl esters were condensed with the corresponding biouronic acid as the 2-acrylamidoethyl or 2-azidoethyl glycosides. The azido function was replaced by the acrylamido group by catalytic hydrogenation followed by N-acryloylation. The tert-butyl groups were removed by treatment with trifluoroacetic acid to give the target monomers which were copolymerised with acrylamide to give neoglycoconjugates that are potentially useful for immunochemical studies.

Synthesis of lysine-containing fragments of the Proteus mirabilis O27 O-specific polysaccharide and neoglyco- conjugates therefrom

Amide-linked lysine mono- and di-uronic acid fragments of the O-specific polysaccharide from P. mirabilis O27 have been synthesised. N epsilon-Boc-L-lysine tert-butyl ester was condensed with 2-azidoethyl glycosides of glucuronic acid and beta-D-GlcpNAc-(1----3)-beta-D-GlcpA. Transformation of the products into 2-acrylamidoethyl glycosides, followed by deprotection using trifluoroacetic acid, gave the target monomers that were converted into high-molecular-weight copolymer-type neoglycoconjugates.

The β-configuration of the rhamnosidic linkage in salmonella serogroups C2 and C3, lipopolysaccharide is important for the immunochemistry of the o-antigen 8

Pairs of synthetic di- and trisaccharide-polyacrylamide (PAA) conjugates, isomers in configuration of the rhamnose residue and related to the sequence abequosyl-(alpha 1-3)-rhamnosyl-(beta 1-2)-mannose (ARM), found in Salmonella serogroup C2 (O-antigens 6,8) and C3 (O-antigens 8,20) lipopolysaccharides, have been used as coating antigens and inhibitors in enzyme immunoassay to evaluate the immunochemical importance of the beta-rhamnosidic linkage in the O-antigen 8. In each pair, the reaction with the factor O:6,8 serum was more pronounced for the synthetic antigen with the beta-rhamnosidic linkage. The ARM-PAA conjugate with the beta-rhamnosidic linkage was found to be 2000 fold more efficient as inhibitor of binding of the factor O:6,8 serum to the AR beta M-PAA conjugate as compared to the alpha-linked analogue. The discrepancy in immunochemical behaviour of the alpha and beta-rhamnose containing ARM oligosaccharides can be explained by conformational differences of the oligosaccharides. A slight cross-reactivity observed in the interaction of antiserum against abequosyl-(alpha 1-3)-mannose, representative of Salmonella O-antigen 4, coupled to BSA, with Salmonella O-factor 8 specific abequosyl-(alpha 1-3)-rhamnose containing conjugates is due to the common terminal immunodominant sugar, abequose.

Glycopolymers from Synthetic Fragments (Amides of α-d-Galacturonic Acid with Amino Acids) of Proteus O-Antigens

Abstract Galacturonamides of amino acids (alanine, lysine, serine, and threonine), constituents of Proteus O-specific polysaccharides, have been synthesised. O-tert-Butyl and N∊-tert-butyloxycarbonyl protected amino acid tert-butyl esters were condensed with the 2-azidoethyl α-glycoside of d-galacturonic acid, prepared by Fischer glycosidation. Reduction of the azido group followed by N-acryloylation and deprotection gave the target monomers. By copolymerisation with acryl-amide, these were converted into glycopolymers potentially useful for defining epitopes in Proteus O-antigens.

Acid-catalyzed isopropylidenation of some heptoses

Abstract Acid-catalyzed acetonation of d - glycero - d - galacto -heptose yields solely the 1,2:3,4:6,7-tri- O -isopropylidene pyranoid derivative, whereas d - glycero - l - gluco - and d - glycero - l - manno -heptose react in the furanose form to give 1,2:5,6-(major) and 1,2:6,7-di- O -isopropylidene- d - glycero - l - gluco -heptose (minor), and 2,3:5,6-(major) and 2,3:6,7-di- O -isopropylidene- d - glycero - l - manno -heptose (minor), respectively.

Glycopeptidolipids--a new class of artificial antigens with carbohydrate determinants. Synthesis of artificial antigen with type-specific oligosaccharide hapten from Neisseria meningitidis group B.

Synthetic lipopeptideN-palmitoyltyrosyl-seryl-seryl-asparaginyl-alanine, an analogue of B-mitogenic tripalmitoyl-pentapeptide fromEscherichia coli lipoprotein, was coupled with an oligosaccharide hapten fromNeisseria meningitidis lipooligosaccharide to give a glycopeptidolipid conjugate — the artificial antigen of a new type possessing the type-specific microbial determinant.

S19.10 Chemical structure and serological specificity of proteus mirabilis 028 O-specific polysaccharide and lipopolysaccharide from a transient form of proteus mirabilis R14/1959

The aminoacyl analysis of endotoxic lipopolysaccharides (LPS) isolated from several bacteria revealed the essential amounts of glycine, among the inherent LPS components. This prompts for the detailed investigation in view of the search for common epitopes suitable for construction of antimicrobial vaccine with broad specificity. The glycine, possibly unifying the LPS structures, might create the common epitope. Significant amounts of glycine was detected in lipopolysaccharides isolated from over 30 strains of Shigella, Escherichia coli, Salmonella, Hafnia and Citrobacter. When the O-specific polysaccharide, lipid A and core fractions of Shigella flexneri and sonnei LPSs were subjected to aminoacyl analysis, glycine as a single aminoacid was found only in core oligosaccharide. The quantity of the aminoacid was not stoichiometric in core oligosaccharides. Molar ratio of glycine ranged from 0.2 to 0.6 per 3 heptoses. Using the HPLC the oligosaccharide enriched in glycine was isolated. Mild acid treatment (0.05 M HC1, 15 min, 100°C) released free glycine. The aminoacid could be also partially cleaved (in 50°70) by pronase digestion. Growing E. coli C600 or Sh. sonnei phase II in the presence of radioactive ~4C-Gly, the labelling of their lipopolysaccharide cores was achieved. The experiments indicate that glycine is covalently bound component of core oligosaccharide in bacterial lipopolysaccharides.