Complete physical map of the rfb gene cluster encoding biosynthetic enzymes for the O antigen of Salmonella typhimurium LT2

Brahmbhatt, Himanshu; Wyk, Paul; Quigley, Neil B.; Reeves, Peter R.

doi:10.1128/jb.170.1.98-102.1988

Cited by 50 publications

(23 citation statements)

References 18 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…One of the current aims in perspective of 0-antigen genetics with a focus on the evolutionary origin of the 0-antigen variation and its role in pathogenesis. Initiation towards this goal began with the cloning and analysis of the Salmonella typhimurium rfb (0-antigen) cluster, which led to the elucidation of the entire sequence of this region and the identification of 12 of the 15 genes present in the cluster (6,15,46). Subsequently, the deoxy and dideoxy genes within the rfb clusters of S. typhimurium (group B, which contains abequose) were compared to those within Salmonella paratyphi (group A, which contains paratose), Salmonella typhi (group D, which contains tyvelose), and, most recently, Y pseudotuberculosis M85 (serogroup IIA, which contains abequose) (17,18).…”

Section: Discussionmentioning

confidence: 99%

“…It has been shown that only a limited number of species of gram-negative bacteria, all in the family Enterobacteraceae, are able to produce these unusual sugars, and, of the eight possible stereoisomers of 3,6-dideoxyhexoses, only five have been identified in nature. Although colitose (3,6-dideoxy-L-Xylo-hexose) and abequose (3,6-dideoxy-D-ylo-hexose, compound I [see Fig. 1]) can be detected in the lipopolysaccharide of some Escherichia coli serotypes and Citrobacter strains, respectively, the five 3,6-dideoxyhexoses known to occur naturally are present mainly in Salmonella and Yersinia spp.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Studies of the biosynthesis of 3,6-dideoxyhexoses: molecular cloning and characterization of the asc (ascarylose) region from Yersinia pseudotuberculosis serogroup VA

et al. 1994

View full text Add to dashboard Cite

The 3,6-dideoxyhexoses are found in the lipopolysaccharides of gram-negative bacteria, where they have been shown to be the dominant antigenic determinants. Of the five 3,6-dideoxyhexoses known to A large degree of bacterial immunological diversity in gram-negative species is attributed to the portion of the lipopolysaccharides known as the 0 antigen. Exposed on the cell envelope, the 0 antigen is the dominant surface entity which is polymorphic and provides the basis for the serological classification of the family Enterobacteriaceae (4,16,26). Among the large number of monosaccharides found as components of 0-specific polysaccharides, derivatives of the 3,6-dideoxyhexoses have drawn special attention because of their highly immunogenic characteristics (22,43,45). It has been shown that only a limited number of species of gram-negative bacteria, all in the family Enterobacteraceae, are able to produce these unusual sugars, and, of the eight possible stereoisomers of 3,6-dideoxyhexoses, only five have been identified in nature. Although colitose (3, and abequose (3,6-dideoxy-D-ylo-hexose, compound I [see Fig. 1]) can be detected in the lipopolysaccharide of some Escherichia coli serotypes and Citrobacter strains, respectively, the five 3,6-dideoxyhexoses known to occur naturally are present mainly in Salmonella and Yersinia spp. as major antigenic determinants (23). For example, abequose, colitose, tyvelose (3,6-dideoxy-D-arabino-hexose, compound II), and paratose (3,6-dideoxy-D-ribo-hexose, compound III) have been found in strains of Salmonella enterica (34), while all five, including ascarylose (3,6-dideOxy-L-arabino-hexose, compound * Corresponding author. Phone: (612) 626-7541. t Present address:

show abstract

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Studies of the biosynthesis of 3,6-dideoxyhexoses: molecular cloning and characterization of the asc (ascarylose) region from Yersinia pseudotuberculosis serogroup VA

et al. 1994

View full text Add to dashboard Cite

show abstract

mentioning

confidence: 99%

“…We have therefore investigating the feasability of enzymatic synthesis of the S. enterica 0-antigenspecific oligosaccharide using enzymes overproduced by cloned genes, for the production of nucleotide sugars and glycosyltransferase reactions. The genes coding for these enzymes belong to the rfb gene cluster [5]. The different genes dTDP-D -glucose In microorganisms, at least three different enzymes are involved in the conversion of dTDP-D-glucose to dTDP-Lrhamnose, the nucleotide sugar which assembles rhamnose to the tetrasaccharide repeating unit [6, 8 -121.…”

mentioning

confidence: 99%

Enzymatic synthesis and isolation of thymidine diphosphate‐6‐deoxy‐D‐xylo‐4‐hexulose and thymidine diphosphate‐L‐rhamnose

Marumo

Lindqvist

Verma

et al. 1992

European Journal of Biochemistry

View full text Add to dashboard Cite

A two-step enzymatic synthesis of dTDP-L-rhamnose is developed using enzymes from sonicated extracts of cultures of Escherichia coli K12 strains harboring plasmids containing different parts of the rjb gene cluster of Salmonella enterica LT2. The intermediate dTDP-6-deoxy-~-xylo-4-hexulose was isolated after a 1 -h reaction, using only dTDP-D-glucose and dTDP-D-glucose 4,6-dehydratase, followed by protein precipitation and desalting by gel chromatography (yield 89Y0). In a two-step reaction using dTDP-D-glucose and dTDP-D-glucose 4,6-dehydratase in the first step, and with NADPH, dTDP-6-deoxy-~-xylo-4-hexulose 3,5-epimerase and NADPH : dTDP-6-deoxy-~-l.vxo-4-hexulose-4-reductase in the second hour of incubation, the dTDP-D-glucose was fully converted to dTDP-L-rhamnose. The hexoses of both products were identified by mass spectroscopy. The molar yield of dTDP-L-rhamnose, after protein precipitation, anion-exchange chromatography and desalting by gel chromatography, was ~W O , corresponding to more than 150 mg, starting from 250 mg of dTDP-D-glucose. When stored lyophilysed under nitrogen, these products were found to be stable for several months. Both dTDP-6-deoxy-~-xylo-4-hexulose and dTDP-L-rhamnose have light absorption maxima at 267 nm, with molar absorption coefficients close to that of dTMP. However, the absorption coefficient of dTDP-6-deoxy-~-.xylo-4-hexulose at the absorption maximum of 320 nm (specific for sugars containing keto groups) was found to be approximately 20% higher than values presented earlier.Furthermore, an HPLC technique is presented for determining the net activity of dTDP-6-deoxy-~-xy/o-4-hexulose 3,5-epimerase and NADPH : dTDP-6-deoxy-~-lyxo-4-hexulose-4-reductase, based on separation of dTDP-6-deoxy-~-xylo-4-hexulose and dTDP-L-rhamnose. The HPLC technique is also suitable for determination of all the nucleotide components involved in the synthesis.The 0 antigen part of the lipopolysaccharide of Salmonella enterica sero-group B strains has a saccharide chain built of site interaction. Therefore, isolated saccharides are important for elucidation of host-defence mechanisms.In recent years advances have been made in the chemical synthesis of saccharides [3]. However, saccharide synthesis is still a relatively complicated procedure, in particular when saccharides larger than the tetrasaccharides shown above are to be synthesized and synthesis is required in large quantities. Such saccharides may be required for the production of synthetic vaccines. S. entericu 0-antigen-specific oligosaccharides have also been produced by enzymatic degradation of the polysaccharide, utilizing bacteriophage glycosidases [4]. So far this procedure has yielded oligosaccharides (monotetramers, ditetramers, tritetramers, and tetramers of a tetrasaccharide) which have terminal sugars different from those of the natural biological repeating unit. We have therefore investigating the feasability of enzymatic synthesis of the S. enterica 0-antigenspecific oligosaccharide using enzymes overproduced by cloned gene...

show abstract

“…A partial gene order for the rJb cluster of strain LT2 was established by using a series of mutants with deletions extending from his into rjb (23,29), and we have recently cloned the whole region from strains LT2 (group B, serovar typhimurium), IMVS1316 (group A, serovar paratyphi A), and Ty2 (group D, serovar typhi) and located the known genes, approximately, on a restriction map of the region (2,3,44). In this study, we located and sequenced the rJbJ gene and showed that when cloned into group A or group D strains, it confers on them the ability to make abequose, which is then incorporated into 0 antigen.…”

mentioning

confidence: 99%

Identification and sequence of the gene for abequose synthase, which confers antigenic specificity on group B salmonellae: homology with galactose epimerase

Wyk

Reeves

1989

J Bacteriol

Self Cite

View full text Add to dashboard Cite

The 0 antigen of Salmonella group B strains contains the sugar abequose, whereas those from group A and D strains contain paratose or tyvelose in its place. This is the essential difference between these Salmonella groups. Only the final step in the biosynthesis of abequose differs from that of paratose, and the abequose confers on group B strains their specific 04 antigen. The gene, rJbJ, encoding the enzyme abequose synthase for this last specific step has been cloned, identified, and sequenced and has been shown to function in group A and D strains to make them 04+. This one gene thus differentiates group B from group A or group D salmonellae. The enzyme abequose synthase appears to be related to galactose epimerase, and the significance of this is discussed. The rjbJ gene and adjacent DNA is of much lower G+C content than is usual for salmonellae, indicating that the region did not originate in a salmonella but was transferred from outside.We are studying the genetics of a major polymorphism in bacteria and present here the genetic base for the 04 antigenic epitope, which characterizes one form of the 0 antigen polymorphism.The 0 antigen is a polysaccharide which covers much of the surface of gram-negative bacteria, although it may be masked by a capsule external to the cell proper. The 0 antigen is typically a polymer with a repeating oligosaccharide of 3 to 6 sugar residues, which is linked through an oligosaccharide core to lipid A, the whole comprising the lipopolysaccharide (LPS), which is the major lipid of the outer leaflet of the outer membrane characteristic of gramnegative bacteria. The lipid A and core do not vary greatly within a genus and are thought to be invariable within salmonellae. The 0 antigen, however, is extremely polymorphic and in salmonellae, about 40 major forms are currently recognized. It should be noted that the genus Salmonella comprises a single species, S. enterica; unfortunately, the many serovars are generally given full species rank, although DNA hybridization and other factors show that the general level of variation between these serovars is that expected for a single bacterial species (4,22

show abstract

Complete physical map of the rfb gene cluster encoding biosynthetic enzymes for the O antigen of Salmonella typhimurium LT2

Cited by 50 publications

References 18 publications

Studies of the biosynthesis of 3,6-dideoxyhexoses: molecular cloning and characterization of the asc (ascarylose) region from Yersinia pseudotuberculosis serogroup VA

Studies of the biosynthesis of 3,6-dideoxyhexoses: molecular cloning and characterization of the asc (ascarylose) region from Yersinia pseudotuberculosis serogroup VA

Enzymatic synthesis and isolation of thymidine diphosphate‐6‐deoxy‐D‐xylo‐4‐hexulose and thymidine diphosphate‐L‐rhamnose

Identification and sequence of the gene for abequose synthase, which confers antigenic specificity on group B salmonellae: homology with galactose epimerase

Contact Info

Product

Resources

About