Chemico-enzymatic synthesis of Salmonella newington O-specific polysaccharide and its modified derivatives

Treatment of the DMan (beta 1-4) LRha (alpha 1-3) D [3H]Gal-derivative of moraprenyl pyrophosphate with the cell envelope preparation from S. newington results in the formation of polysaccharide with beta 1-6 linkage between the trisaccharide units (polymerization degree approximately 8). The synthetic derivatives of moraprenyl pyrophosphate which contain D-talose, 4-deoxy-D-xylo-hexose, LRha (alpha 1-3) DGlc or LRha (alpha 1-3) DGlc (alpha 1-6) DGal were found to serve as substrates for the biosynthesis of the corresponding modified polysaccharides.     

Incorporation of abequose residues into O-specific polysaccharides of Salmonella serotypes B, C2 and C3

The possibility of chemical-enzymatic synthesis of O-specific polysaccharide and its modified derivatives was demonstrated with a cell envelope preparation from Salmonella typhimurium using synthetic polyprenyl pyrophosphate oligosaccharides and CDP-[14C]Abe. It was shown that during biosynthesis of O-specific polysaccharides from S. newport and S. kentucky abequosylation reaction occurs prior to polymerization of oligosaccharide repeating units.     

Introduction of modified hexose residues into O-specific polysaccharides of Salmonella of serotype E, B, C2 and C3 using synthetic nucleotide sugars

A series of synthetic nucleotide sugars have been studied as monosaccharide donors in the biosynthetic assembly of the O-antigen repeating units of Salmonella sero-groups E, B, C2 and C3. UDP-Tal did not serve as a substrate of galactosyl phosphate transferase, and GDP-Tal did not substitute GDP-Man in mannosyltransferase reactions. dTDP-L-Man proved to be a substrate of rhamnosyl transferases; GDP-2dMan, GDP-3dMan, GDP-D-Rha and GDP-Glc served as substrates for mannosyltransferases. The modified tri- and tetrasaccharide derivatives can be converted into the modified O-specific polysaccharides after incubation with Salmonella membranes.     

Chemico-enzymatic synthesis of O-specific polysaccharide of Salmonella anatum and its analogs using partially purified polymerase preparation

A partial purified polymerase from S. anatum was used for the synthesis of polysaccharide [-6) [14C]Man(beta 1-4)Rha(alpha 1-3)Gal(alpha 1-]n and its analogues containing D-glucose residue instead of D-galactose or D-mannose. Structures of these polymers were confirmed by methylation analysis and radioimmunochemical tests.     

Incorporation of paratose residue into Salmonella O-specific polysaccharides using enzymatic reactions]

The block mechanism of O-specific polysaccharides biosynthesis was demonstrated for Salmonella nitra (serogroup A) and S. haifa (serogroup B). Due to the moderate specificity of glycosyl transferases from S. nitra, S. typhimurium, S. haifa and S. kentucky (serogroup C3) towards the 3,6-dideoxyhexose structure a paratose residue can be incorporated into the polysaccharide chain instead of an abequose residue, and vice versa.     

Synthesis of trisaccharide fragment of O-specific polysaccharides of Salmonella of serologic groups C2 and C3 and its conversion into synthetic antigen of a copolymer type

Abequosyl-(alpha 1----3)-L-rhamnopyranosyl-(1----2)-D-mannopyranosides anomeric in the rhamnose residue have been synthesised. 2-Benzyloxycarbonylaminoethyl group used as the aglycon can be transformed into 2-acrylamidoethyl aglycon in the final stages of the syntheses. These isomeric alpha-glycosides were converted into copolymer artificial antigens, which are of interest for studying immunochemistry of factor O:8 of Salmonella O-antigens (serological groups C2 and C3).     

Enzymatic synthesis of polyprenyl phosphosugars in Salmonella serotype C1

The cell envelopes of serogroup C1 Salmonella, viz. S. thompson and S. montevideo, catalyze the transfer of radiolabeled sugars from UDP-[14C]Glc and UDP-[14C]GlcNAc into the lipid-linked sugars. Using TLC and DEAE-cellulose chromatography, the radiolabeled products were identified as polyprenyl pyrophosphate N-acetylglucosamine (I), polyprenyl monophosphate N-acetylglucosamine and polyprenyl monophosphate glucose. The derivative (I) served as an acceptor for mannose transfer from GDP-Man with formation of Man1-2GlcNAc1PPPre. A similar reaction was observed after addition of synthetic GlcNAc1PPPre to the cell envelopes.     

Synthesis of moraprenylpyrophosphate disaccharides--precursors of the biosynthesis of Salmonella O-antigenic polysaccharides of C2 and C3 serotypes

Disaccharides Man(alpha 1-3)Gal and Man(beta 1-3)Gal and their acetates have been synthesized, from which alpha-glycosyl phosphates were obtained by the MacDonald reaction. These compounds were converted into corresponding moraprenylpyrophasphate disaccharides by treating with moraprenylphosphoimidazolidate.     

Chemico-enzymatic method of synthesis of mixed oligonucleotides

Oligo (ribodeoxyribo)nucleotide d(TGTGTAT)r(GCCAU) was synthesized from oligonucleotide blocks by chemical ligation, the oligoribonucleotide and oligodeoxynucleotide blocks being prepared by enzymatic and chemical synthesis, resp. A general procedure of the hybride duplex strand separation is described.     

Synthesis of cytidine diphosphate-3,6-dideoxyhexoses--donors of glycosyl residues in the biosynthesis of O-specific polysaccharides of Salmonella of serogroups A, B and C

Interaction of lithium alcoholates of 2,4-di-O-benzoates of paratose and abequose with tetrabenzyl pyrophosphate gave alpha-phosphates of the 3,6-dideoxyhexoses, further converted into the corresponding cytidine-5'-diphosphate derivatives. These synthetic nucleotides were shown to participate in the biosynthesis of the O-specific polysaccharides for Salmonella typhimurium and S. nitra.     

O-Acetylation in the O-specific polysaccharide isolated from Shigella flexneri serotype 2a

Shigella flexneri causes diarrheal diseases especially in infants and children in developing countries. Modifications of the lipopolysaccharide (LPS) molecule, like bacteriophage-mediated glucosylation and acetylation of the O-specific chain (O-SP), are important for the LPS antigenicity and consequently for the immunogenicity of the polysaccharide-based vaccines against shigellosis. Here, we report the degree of O-acetylation and the localisation of O-acetyl groups and side-chain glucose substitution in the O-SP (scheme) in different preparations of S. flexneri type 2a LPS. [structure: see text]     

Antigenic polysaccharides of bacteria. 33. The structure of O-specific polysaccharide chain of lipopolysaccharide from Pseudomonas cepacia serotype 6

On mild acid degradation of the Pseudomonas cepacia serotype 6 lipopolysaccharide, the O-specific polysaccharide was obtained, which contains D-mannose and D-galactose residues in the ratio approximately 1:1, as well as O-acetyl groups. On the basis of 1H and 13C NMR analysis, calculation of specific optical rotation, and methylation, it was concluded that the polysaccharide possesses the following structure: (formula; see text) Regularities in glycosidation effects in 13C NMR spectra of 1,3-linked disaccharides containing furanoside residues are discussed.     

Antigenic bacterial polysaccharides. 23. The structure of the O-specific polysaccharide chain of Salmonella arizonae 059 lipopolysaccharide

The O-specific polysaccharide of Salmonella arizonae O59 (Arizona 19) is composed of D-galactose, N-acetyl-D-glucosamine, and N-acetyl-L-fucosamine (FucNAc, 2-acetamido-2,6-dideoxy-L-galactose) in the ratio 1:1:1. The computerized calculation of the 13C NMR spectrum of the polysaccharide, based on the monosaccharide composition, spectra of the free monosaccharides and glycosydation effects, together with the chemical analysis (methylation and Smith degradation) showed that the polysaccharide is built up of trisaccharide repeating units of the following structure: ----3)-alpha-L-FucNAcp(1----3)-beta-D-GlcNAcp-(1----2)-beta- D-Galp-1(----. The molecular basis of serological interrelations between S. arizonae O59 and Pseudomonas aeruginosa O7 (Lányi) is discussed.     

Structures of two putative O-specific polysaccharides from the Rahnella aquatilis 3-95 lipopolysaccharide

Two polysaccharide preparations (OPSI and OPSII) were obtained by mild acid degradation of the lipopolysaccharide of Rahnella aquatilis 3-95. Studies by chemical methods and 1H and 13C NMR spectroscopy showed that OPSI is a linear alpha-D-mannan having a trisaccharide repeat and OPSII is a approximately 2:1 mixture of the same mannan and an alpha-d-glucan:     

Antigenic polysaccharides of bacteria. 32. The structure of O-specific polysaccharide chains of Pseudomonas cepacia serotype B and E lipopolysaccharides containing D-fucose

On the basis of acid hydrolysis, methylation, 1H and 13C NMR analysis, and calculation of specific optical rotation, the following structures were established for O-specific polysaccharides of Pseudomonas cepacia serotypes B and E: ----3)-beta-D-Galf-(1----3)-alpha-D-Fucp-(1----serotype B ----3)-beta-D-GlcpNAc-(1----3)-alpha-D-Fucp-(1----serotype E A characteristic feature of the polysaccharides is the presence of D-fucose, rather rare for bacterial antigens.     

Polyprenyl phosphates: synthesis and structure-activity relationship for a biosynthetic system of Salmonella anatum O-specific polysaccharide

A series of polyprenyl phosphates with modified structure of polyprenyl residue was prepared through phosphorylation of polyprenyl trichloroacetimidates with phosphoric acid. Interaction of polyprenols with tetra-n-butylammonium dihydrogen phosphate and trichloroacetonitrile was found to represent a very efficient, simple and general method for the synthesis of polyprenyl phosphates. A procedure was developed for smooth conversion of polyprenyl pyrophosphates into the monophosphates through hydrolysis in the presence of 4-dimethylaminopyridine. The polyprenyl phosphates prepared were studied as substrates for the enzymes of Salmonella anatum O-specific polysaccharide biosynthesis. Correct stereochemistry of alpha- and beta-isoprenic units was found to be essential for substrate efficiency. At the more remote positions of the hydrocarbon chain just the presence of isoprenic units of any configuration seems necessary. Some changes in position of the phosphate group may be permissible without significant loss of substrate properties.     

Immunochemical analysis of oligosaccharide fragments of O-specific Shigella flexneri polysaccharides

Comparison of inhibitory properties of several synthetic oligosaccharides related to Sh. flexneri O-specific polysaccharides, namely Glc alpha 1-3Rha alpha 1-OMe, Glc alpha 1-3Rha alpha 1-2Rha alpha 1-OMe, Rha alpha 1-2(Glc alpha 1-3)Rha alpha 1-OMe, GlcNAc beta 1-2(Glc alpha 1-3)Rha alpha 1-OMe, and GlcNAc beta 1-2(Glc alpha 1-3) Rha alpha 1-2Rha alpha 1-OMe, using passive haemagglutination reaction demonstrated the tetrasaccharide to possess the highest activity in V; 7,8-anti-7,8 immune system. Among the oligosaccharides under study, only Rha alpha 1-2(Glc alpha 1-3)Rha alpha 1-OMe exhibited moderate anti-V activity.     

Structure of O-specific polysaccharide from Yersinia pseudotuberculosis of serotype VI lipopolysaccharide

Using methylation studies, partial hydrolysis and 13C NMR spectroscopy data, the following structure of O-specific polysaccharide from lipopolysaccharide of Yersinia pseudotuberculosis VI serovar has been proposed: (Formula: see text).