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Query: UNIPROT:P43026 (lipopolysaccharide)
 62,215 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In group C1 salmonellae, rfe and rff genes linked to the ilv locus specify the synthesis of a glycolipid called the enterobacterial common antigen. In contrast, in group B salmonellae the synthesis requires in addition some of the genes in the rfb cluster, the main genetic determinant of the O side chains of lipopolysaccharide. In an effort to define the biochemical functions of these rfb genes, we looked in Salmonella typhimurium LT2 (group B) for rfb mutants in which the synthesis of both enterobacterial common antigen and the O side chains would be blocked in a manner suppressible by the wild-type rfe cluster of S. montevideo, of group C1. We found one mutant with these characteristics. This rfb mutation affected the activity of dTDPglucose pyrophosphorylase (glucose-1-phosphate thymidylyltransferase, EC 2.7.7.24). Whereas the rfe cluster of S. montevideo contained a gene producing this enzyme activity, there was no evidence for the presence of such a gene in the rfe cluster of group B strains. These results also showed that the synthesis of dTDP-glucose is necessary for the biosynthesis of enterobacterial common antigen; this conclusion fits with the recent demonstration of 4-acetamido-4,6-dideoxy-D-galactose as a component of enterobacterial common antigen (Lugowski et al., Carbohydr. Res. 118:173-181, 1983), because the biosynthesis of the donor of this sugar, dTDP-4-acetamido-4,6-dideoxy-D-galactose, requires dTDPglucose pyrophosphorylase.
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PMID:Biosynthesis of enterobacterial common antigen requires dTDPglucose pyrophosphorylase determined by a Salmonella typhimurium rfb gene and a Salmonella montevideo rfe gene. 243 Sep 41

The membrane fraction from a mutant of Salmonella anatum deficient in UDPgalactose-4-epimerase, utilized synthetic ficaprenyl alpha-D-galactosyl diphosphate as a substrate in the biosynthesis of the O-polysaccharide portion of lipopolysaccharide which has a mannosylrhamnosylgalactose repeating sequence. The galactosyl lipid was prepared by chemical synthesis from D-galactose and ficaprenol extracted from Ficus elastica. Membrane preparations catalyzed the transfer of rhamnose from TDP-rhamnose onto membrane-bound ficaprenyl galactosyl diphosphate forming rhamnosylgalactosyl ficaprenyl diphosphate; the reaction was dependent on the prior insertion of the synthetic glycosyl-lipid into the membrane, and was proportional to incubation time up to 4 min at 29 degrees C. When both TDP-rhamnose and GDP-mannose were added, the product formed was O-polysaccharide. These results indicate that the chemically synthesized ficaprenyl galactosyl diphosphate can be an active substrate for the in vitro synthesis of the Salmonella O-polysaccharide.
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PMID:Chemically synthesized galactosyl ficaprenyl diphosphate as an intermediate in the biosynthesis of the Salmonella O-antigenic polysaccharide. 618 14

The incorporation of rhamnose and glucose into the core part of the lipopolysaccharide (LPS) of Pseudomonas aeruginosa was studied using enzyme preparations from strain PAC1R and LPS-defective mutants derived from it. Crude membrane preparations from the LPS-defective mutant PAC556 transferred rhamnose from dTDP-L-[14C]rhamnose to material insoluble in trichloracetic acid. The preparations contained both transferase enzyme and acceptor, the former being destroyed by heating. Between 60 and 70% of the radioactive rhamnose transferred to the membranes was extractable by aqueous phenol and non-diffusible. The material extracted did not move in any of the chromatography solvents tested and contained rhamnose as the sole radioactive component. Soluble dTDP-L-rhamnose-LPS rhamnosyltransferase was obtained from the parent strain PAC1R by ammonium sulphate precipitation of a 105000 g supernatant fraction from broken bacteria. It was most active at pH 8 with 5 mM-MgCl2 and required heat-treated membranes of PAC556 as acceptor. This mutant, whose LPS lacks both O-antigenic side-chains and rhamnose in the core, was shown to lack either the epimerase or the NADP-dependent oxidoreductase used to synthesize dTDPrhamnose. After preincubation with soluble transferase and UDPglucose, heated membranes of mutant strains PAC611, PAC612 and PAC605 could also act as acceptors for rhamnose. These mutants all lacked some or all of the glucose as well as the rhamnose from the core of their LPS and the experiments thus provided confirmation that rhamnose was the terminal hexose of the core in P. aeruginosa PAC1.
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PMID:Biosynthesis of the core part of the lipopolysaccharide of Pseudomonas aeruginosa. 677 64

The rough lipopolysaccharide (LPS) of commonly used strains of Escherichia coli K-12 has two distinctly different band patterns when analyzed by high-resolution polyacrylamide gel electrophoresis. The LPS of ancestral strains such as W1485F- consists primarily of a single broad gel band. In contrast, the LPS of strains derived from strain Y10 such as AB1133 or C600 gives three sharp gel bands. Complementation studies using DNA fragments from the rfb gene cluster of Shigella dysenteriae 1 indicated that the difference between the two gel patterns is due to a mutation in the gene encoding the TDP-rhamnose synthetase, the final enzyme involved in TDP-rhamnose biosynthesis. This mutation arose during the construction of strain Y10, and not in strain 679-680 as previously thought. The requirement for the rfaS gene for synthesis of the broad major band seen in W1485F- LPS and the shift in gel migration of a component of this band when an rfaQ mutation was introduced indicated that this broad band contained the unique form of rough E. coli LPS which has been termed lipooligosaccharide. This finding indicates that lipooligosaccharide is likely to contain rhamnose and suggests a model in which one of the functions of partial substituents such as rhamnose may be to direct core synthesis into different pathways to produce alternative forms of LPS.
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PMID:Genes for TDP-rhamnose synthesis affect the pattern of lipopolysaccharide heterogeneity in Escherichia coli K-12. 751 88

The O-repeating unit of the Escherichia coli O7-specific lipopolysaccharide is made of galactose, mannose, rhamnose, 4-acetamido-4,6-dideoxyglucose, and N-acetyglucosamine. We have recently characterized the genes involved in the biosynthesis of the sugar precursor GDP-mannose occurring in the E. coli O7:K1 strain VW187 (C. L. Marolda and M. A. Valvano, J. Bacteriol. 175:148-158, 1993). In the present study, we identified and sequenced the rfbBDAC genes encoding the enzymes for the biosynthesis of another precursor, dTDP-rhamnose. These genes are localized on the upstream end of the rfbEcO7 region, and they are strongly conserved compared with similar genes found in various enteric and nonenteric bacteria. Upstream of rfbB we identified a DNA segment containing the rfb promoter and a highly conserved untranslated leader sequence also present in the promoter regions of other surface polysaccharide gene clusters. Also, we have determined that rfbB and rfbA have homologs, rffG (o355) and rffH (o292), respectively, located on the rff cluster, which is involved in the synthesis of enterobacterial common antigen. We provide biochemical evidence that rffG and rffH encode dTDP-glucose dehydratase and glucose-1-phosphate thymidylyltransferase activities, respectively, and we also show that rffG complemented the rfbB defect in the O7+ cosmid pJHCV32. We also demonstrate that rffG is distinct from rffE and map the rffE gene to the second gene of the rff cluster.
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PMID:Genetic analysis of the dTDP-rhamnose biosynthesis region of the Escherichia coli VW187 (O7:K1) rfb gene cluster: identification of functional homologs of rfbB and rfbA in the rff cluster and correct location of the rffE gene. 755 40

The Yersinia enterocolitica O:3 lipopolysaccharide O-antigen is a homopolymer of 6-deoxy-L-altrose. The cloned rfb region was sequenced, and 10 open reading frames were identified. Transposon mutagenesis, deletion analysis and transcomplementation experiments showed that eight of the genes, organized into two operons, rfbABC and rfbDEFGH, are essential for O-antigen synthesis. Functional tandem promoters were identified upstream of both operons. Of the deduced polypeptides RfbA, RfbF and RfbG were similar to Salmonella proteins involved in the dTDP-L-rhamnose biosynthesis. Rhamnose and 6-deoxy-L-altrose are C3-epimers suggesting that analogous pathways function in their biosynthesis. RfbD and RfbE were similar to capsular polysaccharide export proteins, e.g. KpsM and KpsT of Escherichia coli. This and transposon mutagenesis showed that RfbD and RfbE function as O-antigen exporters.
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PMID:Genetic organization and sequence of the rfb gene cluster of Yersinia enterocolitica serotype O:3: similarities to the dTDP-L-rhamnose biosynthesis pathway of Salmonella and to the bacterial polysaccharide transport systems. 769 17

A plasmid that included both an 8.9 kb chromosomal DNA insert containing genes from the rfb cluster of Shigella dysenteriae 1 and a smaller insert containing the rfp gene from a S. dysenteriae 1 multicopy plasmid resulted in efficient expression of O antigen in an rfb-deleted strain of Escherichia coli K-12. Eight genes were identified in the rfb fragment: the rfbB-CAD cluster which encodes dTDP-rhamnose synthesis, rfbX which encodes a hydrophobic protein involved in assembly of the O antigen, rfc which encodes the O antigen polymerase, and two sugar transferase genes. The production of an O antigen also required the E. coli K-12 rfe gene, which is known to encode a transferase which adds N-acetylglucosamine phosphate to the carrier lipid undecaprenol phosphate. Thus Rfe protein appears to function as an analogue of the Salmonella RfbP protein to provide the first sugar of the O unit. Functional analysis of the other genes was facilitated by the fact that partial O units of one, two or three sugars were efficiently transferred to the lipopolysaccharide core. This analysis indicated that the plasmid-encoded Rfp protein is the transferase that adds the second sugar of the O unit while the two rfb transferases add the distal sugars to make an O antigen whose structure is (Rha-Rha-Gal-GlcNAc)n. The use of the rfe gene product as the transferase that adds the first sugar of an O unit is a novel mechanism which may be used for the synthesis of other enteric O antigens.
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PMID:Function of the rfb gene cluster and the rfe gene in the synthesis of O antigen by Shigella dysenteriae 1. 769 19

Neisseria gonorrhoeae synthesizes a rough lipopolysaccharide that does not contain any of the repetitive units characteristic of the smooth lipopolysaccharide of members of the family Enterobacteriaceae. Three gonococcal homologs of Salmonella serovar typhimurium genes involved in the synthesis of the rhamnose component of the repetitive subunits have been isolated. Gonococcal homologs for rfbB, rfbA, and rfbD were found downstream of the galE gene in a region of the chromosome which shows overall homology with the meningococcal capsule gene complex region D. Sequence alignment demonstrated that the gonococcal gene products have 69, 65, and 54% amino acid identity with the Salmonella proteins RfbB, RfbA, and RfbD. The gonococcal RfbB and RfbA amino acid sequences share even more identical residues (73 and 65%, respectively) with the amino acid sequences derived from Escherichia coli genes o355 and o292, respectively. These genes are clustered with the genes involved in the biosynthesis of enterobacterial common antigen, and o355 is listed in the GenBank and Swiss Protein data banks as rffE (encoding UDP-GlcNAc-2-epimerase). However, complementation studies demonstrated that o355 does not encode the enzyme UDP-GlcNAc-2-epimerase. Gonococcal strains constructed with null mutations in the rfbBAD genes were unchanged in lipopolysaccharide phenotype and in the synthesis of gonococcal carbohydrate-containing C antigen. We were unable to detect any changes in gonococcal phenotype with respect to lipopolysaccharide sialylation, monoclonal-antibody binding, serum sensitivity, or interaction with eukaryotic cells in vitro. We conclude that the absence of a homolog for rfbC precludes the existence of a functional dTDP-rhamnose biosynthesis pathway in the gonococcal strains examined and that these genes are only maintained in N. gonorrhoeae either because of the presence of the galE gene or because of another as yet unrecognized function.
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PMID:The identification of cryptic rhamnose biosynthesis genes in Neisseria gonorrhoeae and their relationship to lipopolysaccharide biosynthesis. 796 52

By mutational analysis it was found that a 3.9-kb SmaI-XhoII DNA fragment of Xanthomonas campestris pv. campestris is involved in lipopolysaccharide (LPS) biosynthesis. LPS samples isolated from different mutants carrying mutations in the 3.9-kb SmaI-XhoII DNA fragment exhibited banding patterns in silver-stained sodium dodecyl sulfate-polyacrylamide gels markedly different from that of the wild-type LPS. Moreover, comparison of the monosaccharide composition obtained by high-performance anion-exchange chromatography with pulsed amperometric detection of LPS purified from wild-type Xanthomonas campestris pv. campestris B100 and from mutants with mutations in the 3.9-kb SmaI-XhoII DNA fragment revealed a lack of rhamnose moieties in the mutant LPS. Sequence analysis of this DNA fragment revealed four open reading frames (ORFs), designated ORF302, ORF183, ORF295, and ORF351. The deduced amino acid sequences of these ORFs showed a high degree of homology to the deduced amino acid sequences of the rfbC, rfbD, rfbA, and rfbB genes of Salmonella typhimurium LT2, which have been shown to encode a set of enzymes responsible for conversion of glucose 1-phosphate to dTDP-rhamnose.
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PMID:A 3.9-kb DNA region of Xanthomonas campestris pv. campestris that is necessary for lipopolysaccharide production encodes a set of enzymes involved in the synthesis of dTDP-rhamnose. 825 67

We have cloned a new gene locus that comprises three genes concerned with the biosynthesis of the serotype c-specific polysaccharide antigen in Streptococcus mutans. The genes encode proteins exhibiting significant homology to the rfbA, rfbB, and rfbD gene products that are involved in the anabolism of dTDP-L-rhamnose from D-glucose-1-phosphate. This anabolism pathway pertains to biosynthesis of the O antigen of lipopolysaccharide in gram-negative bacteria. The cell extract of Escherichia coli expressing each of the cloned genes of S. mutans exhibited enzymatic activity corresponding to the homologous counterpart of the rfb gene products. Rhamnose was not detected in the cell wall preparation purified from the mutant in which each of the three cloned genes was insertionally inactivated. Rabbit antiserum against S. mutans serotype c-specific antigen did not react with the autoclaved extracts from these mutants. These results indicate that the gene products identified in the present study are involved in the dTDP-L-rhamnose synthesis pathway and that the pathway relates to the biosynthesis of the serotype-specific polysaccharide antigen of S. mutans. Southern hybridization analysis revealed that genes homologous to the cloned genes involved in the dTDP-L-rhamnose synthesis pathway were widely distributed in a variety of streptococci. This is the first report of the biological function of the dTDP-rhamnose pathway in streptococci.
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PMID:Biological function of the dTDP-rhamnose synthesis pathway in Streptococcus mutans. 902 94


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