Salmonella, O-specific polysaccharides

The structures of polyprenyl diphosphate-linked intermediates of Salmonella O-specific-polysaccharide biosynthesis were confirmed by chemical synthesis of their analogs derived from the plant polyprenols ficaprenol and moraprenol (structurally related to bacterial polyprenol57) with the following study of their behavior as substrates of enzymic reactions. Synthetic polyprenyl a-D-galactopyranosyl diphosphate291,292 was found to serve as an effective acceptor for the transfer of L-rhamnosyl groups.293"295 Two synthetic, isomeric disaccharide derivatives,292 13 and296 14, were tested as acceptors for enzymic D-mannosyl transfer from GDP-Man, but only the former was found to be an efficient substrate.294... 

The factors that determine the degree of polymerization of the polysaccharides remain unclear. It seems that the process is not under strict control, as significant heterogeneity in chain length is observed for Salmonella O-specific polysaccharides.302 304 In some cases, the degree of polymerization was found to depend on the conditions of cultivation of the micro-... 

The block mechanism of chain assembly is characteristic for polymeric chains of the UGT type (see Salmonella O-specific polysaccharides 10-12 and 18) and the UG type (see capsular polysaccharides 25, 27, and 33), with UDP-activated sugars serving as initiators of chain growth. It seems rather safe to suggest that the biosynthesis of other polymers of these types occurs through a block mechanism as well. 

V. N. Shibaev, L. L. Danilov, T. N. Druzhinina, L. M. Gogilashvili, S. D. Maltsev, and N. K. Kochetkov, Enzymatic synthesis of Salmonella O-specific polysaccharide analogs from modified polyprenyl pyrophosphate sugar acceptors, FEBS Lett., 139 (1982) 177-180. 

CDP-6-deoxy-D-xy/o-hexos-4-ulose (7b) serves as a precursor in the biosynthesis of CDP-3,6-dideoxyhexoses28,168 having the d-ribo (paratose, 3,6-dideoxy- D-glucose ), o-xylo (abequose, 3,6-dideoxy- D-galactose ), and d-arabino (tyvelose, 3,6-dideoxy- D-mannose ) configurations. These monosaccharides are characteristic components of O-specific polysaccharides from Salmonella and Yersinia pseudotuberculosis. 

Approximately 400 different glycosyltransferases are necessary in order to ensure the synthesis of those bacterial polysaccharides whose structures have thus far been elucidated. This estimate is based on the results of an analysis of the structures, made in order to ascertain how many different disaccharide fragments are present. An example of such an analysis is shown in Table V for the disaccharide sequences L-rhamnopyranosyl-D-galacto-pyranose, D-mannopyranosyl-L-rhamnopyranose, and D-galactopyranosyl-D-mannopyranose that are characteristic for the O-specific polysaccharides of Salmonella serogroups A, B, D, and E, the objects of many biosynthetic studies. Full details of similar analyses for other disaccharide sequences will be published elsewhere, as the resulting Tables are too voluminous for inclusion in this Chapter, but the most interesting results are summarized in Tables VI and VII. 

Incorporation of abequosyl side-chains in O-specific polysaccharides (12) of Salmonella serogroup B occurs at the stage of repeating-unit assembly. The polyprenyl trisaccharide diphosphate 16a serves as an acceptor for abequosyl transfer from its CDP derivative.83,307 A similar reaction was demonstrated with an enzyme preparation from Citrobacter.290 Polymerases from S. typhimurium83,308 and S. bredeney286 were able to act on the derivatives of linear trisaccharide 16a, but, in the former case, the efficiency of... 

Another example of O-specific polysaccharides that are assembled through the block mechanism are the polymers from Salmonella serogroups C2 and C3. Their polysaccharide chains (18) are composed97 of linear, tetrasaccharide repeating-units having abequosyl and D-glucosyl branches,... 

Preparation of modified, bacterial polysaccharides having monosaccharide analogs inserted into the polymeric chain is of interest for study of the structure-properties relationship in these biopolymers. Incorporation of chemically prepared, modified, biosynthetic precursors of the polymers in enzymic reactions seems a promising approach for achieving this aim. Such an approach, which may be termed chemical-enzymic synthesis, has now been studied by our group,439-441 using O-specific polysaccharides (10-12) of Salmonella serogroups B and E as an example. 

The chemical-enzymic approach to the synthesis of modified polysaccharides presents a good prospect for the preparation of small quantities of these polymers, which may prove very useful for immunochemical studies. The approach is certainly not limited by the specific case of Salmonella polysaccharides 10-12, and may well be extended to other polymers. The first results from this group322 show that several analogs of O-specific polysaccharides (18) of Salmonella serogroups C2 and C3 may be prepared through this approach. 

Figure 2. Specific immunoabsorbents for Salmonella O and R form antibodies. Key PS, O-specific polysaccharide, core, core fragment and GlcNHr glucosamine and fatty acids. Specific immunoabsorbents for lipid A antibodies have also been prepared (C. Galanos and D. Nerkar, unpublished results Luderitz et a ., 1982).

Synthesis of f -D-(l,6) linked disaccharides of N-fatty acylated 2-amino-2-deoxy-D-glucose an approach to the lipid A component of the bacterial lipopolysaccharide. Carbohydr. Res., 88, C10-C13 Kochetkov, N.K., Dmitriev, B.A., Malysheva, N.N., Chernyak, A.Ya., Klimov, E.M., Bayramova, N.E., and Torgov, V.I. (1975). Synthesis of O-p-D-mannopyranosy l-( 1 - 0-0-ot -L-rhamnopyranosy l-( 1 -3)-D-galacto-pyranose, the trisaccharide repeating-unit of the O-specific polysaccharide from Salmonella anatum. Carbohydr. Res. 15, 283-290... 

Synthesis of the tetrasaccharide repeating-unit of the O-specific polysaccharide from Salmonella senftenberg. Carbohydr. Res. 5 1, 269-27 1... 

V Shibaev Biosynthesis of salmonella O-antigenic polysaccharide specificity of glycosyl transferases... 

D. C. Watson, J. B. Robbins, and S. C. Szu, Protection of mice against Salmonella typhimurium with an O-specific polysaccharide-protein conjugate vaccine, Infect Immun., 60 (1992) 4679-4686. 

S. N. Senchenkova, A. S. Shashkov, Y. A. Knirel, E. Schwarzmiiller, andH. Mayer, Structure of the O-specific polysaccharide of Salmonella enterica ssp. Arizonae 050 (Arizona 9a,9b),... 

Bovine Serum Albumin Conjugates of Mono- and Di-saccharides Corresponding to O-Specific Polysaccharides of Salmonella ... 

Olsthoom, M.M., Petersen, B.O., Duus, J., Haverkamp, J., Thomas-Oates, J.E., Bock, K., Holst, O. The structure of the linkage between the O-specific polysaccharide and the core region of the lipopolysaccharide from Salmonella enterica serovar Typhimurium revisited. Eur J Biochem 267 (2000) 2014-2027. 

The repeating unit of the O-specific polysaccharide side-chains of the Salmonella typhimurium antigenic outer bacterial cell wall is as follows ... 

Silver oxide has also been successfully used for the formation of p-mannosyl-rhamnose linkages, which are present in the O-specific polysaccharide of Salmonella [17]. Kochetkov and co-workers effectively coupled identical donor 4 with the rhamnose acceptor 6 by activation with silver oxide to exclusively afford the P-Man-(1 — 4)-a-Rha disaccharide 7 in an impressive 91% yield (Scheme 8.2) [25]. 

Component of the O-specific polysaccharide of Salmonella kentucky strain 1S98. 

Among the tetra- and penta-saccharides to be synthesized were (53)/ (54) (the repeating-unit of the O-specific polysaccharide from Salmonella senftenberg), a derivative of (55), (56) (in connection with immunological studies of the blood-group substances), and (57) (one of the core oligosaccharides of human blood-group substances). ... 

The synthesis of 0-j3-D-mannopyranosyl-(l -> 4)-0-a-L-rhamnopyranosyl-(1 - 3)-[0-a-D-glucopyranosyl-(l -> 6)]-o-galactose, the repeating unit of the O-specific polysaccharide chain of the lipopolysaccharide from Salmonella... 

Figure 1 represents the general structure of Salmonella lipopolysaccharides. They contain an external polysaccharide, the O-antigenic chain, and an innermost component, termed lipid A. O-chain and lipid A are linked to each other by an oligosaccharide referred to as the core. O-Specific Chains. As indicated in Figure 1, O chains are in general made up of repeating units of di-, tri-, or higher oligosaccharides. In rare cases the O-chain is a homopolysaccharide. The structure of the O-chain is unique to each bacterial serotype great diversity is encountered in the structures of O-chains.

Figure 3. Structures of the O-antigenic polysaccharide chain in Salmonella bacteria of serogroups C2 and C3 and a synthetic disaccharide glycoconjugate (O-antigen 8-specific). Results of immunofluorescence studies (IFL) of these structures.

Figure 4. Structures of the O-antigenic polysaccharide chains in Salmonella bacteria ofserogroup E and a synthetic trisaccharide gly coconjugate (O-antigen 3-specific). Results of ELISA and immunofluorescence (IFL) studies. Key m, S. anatum (03, 10) , S. senftenburg (01, 3, 19) , BSA a, S. typhimurium (04, 5,...

TABLE I. Salmonella typhimurium Infection in Calves Delayed Skin Reactivity with Specificity for O-Antigenic Polysaccharide Chain... 

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