Diversity of capsular polysaccharide synthesis gene clusters in Streptococcus pneumoniae

Echoplanar functional magnetic resonance imaging (fMRI) was used in normal human subjects to investigate the role of the amygdala in conditioned fear acquisition and extinction. A simple discrimination procedure was employed in which activation to a visual cue predicting shock (CS+) was compared with activation to another cue presented alone (CS-). CS+ and CS- trial types were intermixed in a pseudorandom order. Functional images were acquired with an asymmetric spin echo pulse sequence from three coronal slices centered on the amygdala. Activation of the amygdala/periamygdaloid cortex was observed during conditioned fear acquisition and extinction. The extent of activation during acquisition was significantly correlated with autonomic indices of conditioning in individual subjects. Consistent with a recent electrophysiological recording study in the rat (Quirk et al., 1997), the profile of the amygdala response was temporally graded, although this dynamic was only statistically reliable during extinction. These results provide further evidence for the conservation of amygdala function across species and implicate an amygdalar contribution to both acquisition and extinction processes during associative emotional learning tasks.     

Antibody responses to capsular polysaccharide backbone and O-acetate side groups of Streptococcus pneumoniae type 9V in humans and rhesus macaques

Streptococcus pneumoniae is responsible for high rates of pneumococcal bacteremia, meningitis, pneumonia, and acute otitis media worldwide. Protection from disease is conferred by antibodies specific for the polysaccharide (Ps) capsule of the bacteria. Of the four types of group 9 pneumococci, types 9N and 9V cause the most disease, and both types are included in the polyvalent pneumococcal vaccine. The type 9V capsule consists of repeating pentasaccharide units linearly arranged, with an average of 1 to 2 mol of O-acetate side chains per mol of repeat units, added in a complex pattern in which not all repeat units are alike. alpha-GlcA residues may be O-acetylated in the 2 (17%) or 3 (25%) position and beta-ManNAc residues may be O-acetylated in the 4 (6%) or 6 (55%) position. Under certain conditions, the O-acetate side chains are subject to oxidation, which results in subsequent de-O-acetylation of a significant number of the repeat units. This de-O-acetylation could adversely affect the efficacy of a vaccine containing the 9V Ps. A study was undertaken to compare the relative contributions of O-acetate and Ps backbone epitopes in the immune response to S. pneumoniae 9V type-specific Ps. In both an infant rhesus monkey model and humans, antibodies against the non-O-acetylated 9V backbone as well as against O-acetylated 9V Ps were detected. Functional (opsonophagocytic) activity was observed in antisera in which the predominant species of antibody recognized de-O-acetylated 9V Ps. We concluded that the O-acetate side groups, while recognized, are not essential to the ability of the 9V Ps to induce functional antibody responses.     

Streptococcus pneumoniae type 14 polysaccharide-conjugate vaccines: length stabilization of opsonophagocytic conformational polysaccharide epitopes

A simple and convenient method was developed for the preparation of Streptococcus pneumoniae type 14 polysaccharide (Pn14PS)-tetanus toxoid (TT) conjugate vaccines, using terminally linked Pn14PS fragments of different lengths. Native Pn14PS was simultaneously depolymerized and activated for conjugation by partial N-deacetylation followed by nitrous acid deamination which yielded fragments (1.4 to 150.0 kDa) having a free aldehyde at the reducing end. These were then conjugated to TT through their terminal aldehydic groups, using the reductive amination procedure. All of the above conjugates, when injected in rabbits, induced anti-Pn14PS antibodies, whereas the native Pn14PS did not. The amounts of anti-Pn14PS antibodies elicited by these conjugates, as determined by enzyme-linked immunosorbent assay, followed a trend with conjugates containing the highest-molecular-weight Pn14PS eliciting the highest titers. The same trend was also observed in the ability of the antibodies to opsonize and kill live type 14 pneumococci, although the increase in opsonophagocytic activity was more pronounced and did not correlate linearly with increases in antibody titer. Competitive inhibition of the binding of different conjugate antisera to the native Pn14PS, using Pn14PS fragments as inhibitors, established that the conjugates induced antibodies with specificities for different lengths of Pn14PS beginning at 2 repeating units (RU). It was also established, both immunologically and antigenically, that at least 4 RU of Pn14PS were required to form an extended conformational epitope and that approximately 22 RU of Pn14PS were required to duplicate the same epitope on the same saccharide chain. The conformational epitope was found to be essential for the induction of antibodies with high opsonophagocytic activity and that augmentation of opsonophagocytic activity was also dependent on further chain extension.     

Functional analysis of glycosyltransferases encoded by the capsular polysaccharide biosynthesis locus of Streptococcus pneumoniae serotype 14

Bacteria belonging to the species Streptococcus pneumoniae vary in their capsule. Presently, 90 capsular serotypes are known, all possessing their own specific polysaccharide structure. Little is known about the biosynthesis of these capsular polysaccharides. The cps locus of S. pneumoniae serotype 14 was cloned. So far, 7 open reading frames have been sequenced, cps14B to cps14H. The gene products are similar to proteins involved in bacterial polysaccharide biosynthesis, both of Gram-negative and -positive micro-organisms. Gene-specific mutants were created for cps14D to cps14H by insertional mutagenesis. All mutants no longer agglutinated with a monoclonal antibody against type 14 capsule polysaccharides. The biosynthetic function of cps14E and cps14G was determined by analysis of the intermediates in the synthesis of the oligosaccharide subunit, formed in membrane preparations of the wild-type and mutant strains and in membrane preparations of Escherichia coli expressing the pneumococcal glycosyltransferases. The enzyme encoded by cps14E is a glucosyl-1-phosphate transferase that links glucose to a lipid carrier, the first step in the biosynthesis of the type 14 repeating unit. The gene product of cps14G encodes a beta-1,4-galactosyltransferase, the enzyme responsible for the second step in the subunit synthesis, the transfer of galactose to lipid-linked glucose.     

Molecular characterization of the complete 23F capsular polysaccharide locus of Streptococcus pneumoniae

The complete DNA sequence of the capsular locus 23F of Streptococcus pneumoniae is presented. The 18.6-kb cps23f locus is composed of 18 open reading frames flanked at the 5' and 3' ends by the genes dexB and aliA, an arrangement similar to those of some of the other identified cps loci.     

Synthesis of four spacer-containing 'tetrasaccharides' that represent four possible repeating units of the capsular polysaccharide of Streptococcus pneumoniae type 6B

In the framework of studies towards oligosaccharide-conjugate based vaccines against Streptococcus pneumoniae, the synthesis is reported of four spacer-containing tetrasaccharides that each can be conceived as representing a repeating unit of the capsular polysaccharide of S. pneumoniae serotype 6B, namely, 3-aminopropyl D-ribityl-(5-->hydrogen phosphate-->2)-alpha-D-galactopyranosyl-(1-->3) -alpha-D-glucopyranosyl-(1-->3)-alpha-L-rhamnopyranoside, 3-aminopropyl alpha-L-rhamnopyranosyl-(1-->4)-D-ribityl-5(-->hydrogen phosphate-->2)-alpha-D-galactopyranosyl-(1-->3)-alpha-D-glucopyranoside, 3-aminopropyl alpha-D-glucopyranosyl-(1-->3)-alpha-L-rhamnopyranosyl-(1-->4) -D-ribityl-(5-->hydrogen phosphate-->2) -alpha-D-galactopyranoside, and alpha-D-galactopyranosyl-(1-->3)-alpha-D-glucopyranosyl-(1-->3)-alpha-L -rhamnopyranosyl-(1-->4)-5-O-(3-aminopropyl hydrogen phosphate)-D-ribitol. Phosphorylations were carried out using the H-phosphonate method.     

Association of intrastrain phase variation in quantity of capsular polysaccharide and teichoic acid with the virulence of Streptococcus pneumoniae

The pneumococcus undergoes spontaneous phase variation between an opaque and a transparent colony form. In an animal model of systemic infection following intraperitoneal inoculation of mice, the opaque phenotype was significantly more virulent than the transparent for each of 3 strains examined. The opaque phenotype was associated with 1.2- to 5.6-fold greater amounts of capsular polysaccharide compared with the transparent using a sandwich ELISA. A similar technique comparing the amount of total teichoic acid showed that the transparent phenotype had 2.1- to 3.8-fold more immunodetectable teichoic acid. This difference was confirmed by comparing the incorporation of [3H]choline into teichoic acid. Cell fractionation revealed that variation in quantity of incorporated choline was due to differences in cell wall-associated teichoic acid. Results suggest that the pneumococcus phase varies between a virulent form with more capsular polysaccharide and less teichoic acid and an avirulent form with less capsular polysaccharide and more teichoic acid.     

The structure of the capsular polysaccharide from Klebsiella type 52, using the computerised approach CASPER and NMR spectroscopy

In swine, distinct centromeric satellite DNA families have been described that correspond to either all the metacentric chromosomes except the Y (Mc1) or all the acrocentric chromosomes (Ac2). Using primed in situ (PRINS) labeling, we show here that primers derived from various sequences specifically label the centromeres of different subgroups of chromosomes. Among five primers derived from centromeric sequences of acrocentric chromosomes reported to be very homogeneous, four recognize all the acrocentric chromosomes, whereas one labels prominently chromosome 17. For the metacentric chromosomes, six primers have been derived from several divergent sequences. Among these primers, two recognize all the metacentric chromosomes except 5, 10, and 12. Three other primers label small subsets of metacentric chromosomes, including the X and one or two additional chromosomes. The last primer is specific to chromosome 1. These preliminary results suggest that it should be possible to define specific primers for almost every swine chromosome. Already, some of the primers reported here permit a distinction between swine chromosomes difficult to differentiate without banding, such as the X chromosome and chromosome 9. Therefore, the PRINS technique using centromeric motifs constitutes an additional tool for cytogenetic studies in swine.     

Genetic analysis of type 5 capsular polysaccharide expression by Staphylococcus aureus

Capsules are produced by over 90% of Staphylococcus aureus strains, and approximately 25% of clinical isolates express type 5 capsular polysaccharide (CP5). We mutagenized the type 5 strain Reynolds with Tn918 to target genes involved in CP5 expression. From a capsule-deficient mutant, we cloned into a cosmid vector an approximately 26-kb EcoRI fragment containing the transposon insertion. In the absence of tetracycline selection, Tn918 was spontaneously excised, thereby resulting in a plasmid containing 9.4 kb of S. aureus DNA flanking the Tn918 insertion site. The 9.4-kb DNA fragment was used to screen a cosmid library prepared from the wild-type strain. Positive colonies were identified by colony hybridization, and a restriction map of one clone (pJCL19 with an approximately 34-kb insert) carrying the putative capsule gene region was constructed. Fragments of pJCL19 were used to probe genomic DNA digests from S. aureus strains of different capsular serotypes. Fragments on the ends of the cloned DNA hybridized to fragments of similar sizes in most of the strains examined. Blots hybridized to two fragments flanking the central region of the cloned DNA showed restriction fragment length polymorphism. A centrally located DNA fragment hybridized only to DNA from capsular types 2, 4, and 5. DNA from pJCL19 was subcloned to a shuttle vector for complementation studies. A 6.2-kb EcoRI-ClaI fragment complemented CP5 expression in a capsule-negative mutant derived by mutagenesis with ethyl methanesulfonate. These experiments provide the necessary groundwork for identifying genes involved in CP5 expression by S. aureus.     

The aqueous solution structure of a lipoteichoic acid from Streptococcus pneumoniae strain R6 containing 2,4-diamino-2,4,6-trideoxy-galactose: evidence for conformational mobility of the galactopyranose ring

The 2D-NOESY spectra for the per-N-acetylated and the native lipoteichoic acid (LTA) oligomer from Streptococcus pneumoniae strain R6 clearly indicate a difference in conformation of the 2,4,6-trideoxy-galactopyranose ring. Whereas the 2,4-N-acetylated Gal24N adopts the usual 4C1 chair conformation, the native 2-N-acetyl-4-amino Gal24N exhibits conformational mobility with comparable populations in the 4C1 chair and 5S1 skew conformations, as determined using MD simulation for the partial trisaccharide Me-beta-D-Glc6P-(1-->3)-alpha-D-Gal24N-[6-PC]-(1-->4)-alpha- D-galNAc and from the intra-ring NOE effects. 31P-NMR spectra point to a strong electrostatic or hydrogen-bonding interaction between the free 4-NH2 group on the Gal24N and the negatively charged diester phosphate group between adjacent pentasaccharide repeating-units [Ribitol-(5-->6)-beta-D-Glc6P]. Molecular modelling and MD simulation experiments confirmed that such an interaction was feasible with the Gal24N galactopyranose ring in the inverted B1.4 or skew 5S1 conformation.     

The chemical structures of the capsular polysaccharides from Streptococcus pneumoniae types 32F and 32A

The structures of the capsular polysaccharides from Streptococcus pneumoniae types 32F and 32A have been determined by means of NMR spectroscopy as the principal method. It is concluded that both polysaccharides are composed of tetrasaccharide repeating units with a phosphorylcholine (PCho) group linked to the 3-position of the 4-substituted beta-L-rhamnose (Rha) residue. Both polysaccharides are substituted with one O-acetyl group at the 2-position of the same beta-L-rhamnose residue. In addition, the type-32A polysaccharide is substituted with another O-acetyl group at the 4-position of the 2,3-disubstituted alpha-D-glucose residue, i.e. the branch-point residue. An unusual detail in the structure is that the side chain is composed of a rhamnosyl phosphate. [chemical structure: see text] In the type-32F polysaccharide R=H, and in the type-32A polysaccharide R=Ac. The structure of C-polysaccharide found in our preparations of type-32F and type-32A capsular polysaccharides is in agreement with that published previously for the pneumococcal common antigen C-polysaccharide [Fischer, W., Behr, T., Hartmann, R., Peter-Katalinic, J. & Egge, H. (1993) Eur. J. Biochem. 215, 851-857; Kulakowska, M., Brisson, J.-R., Griffith, D. W., Young, N. M. & Jennings, H. J. (1993) Can. J. Chem. 71, 644-648].     

Oligoclonality of serum immunoglobulin G antibody responses to Streptococcus pneumoniae capsular polysaccharide serotypes 6B, 14, and 23F

Serum antibodies (Abs) specific for the capsular polysaccharides of Streptococcus pneumoniae provide protection against invasive pneumococcal disease. Previous studies indicate that Abs to pneumococcal polysaccharide (PPS) serotypes 1 and 6B have limited clonal diversity. To determine if restricted diversity was a feature common to other PPS specificities, we examined the light (L)-chain expression and isoelectric heterogeneity of type 6B, 14, and 23F Abs elicited in 15 adults following PPS vaccination. At the population level, both PPS-6B and PPS-14 Abs expressed kappa and lambda chains, although 6B Abs more frequently expressed lambda chains lambda and 14 Abs more frequently expressed kappa chains. In individual sera, Abs were generally skewed towards either kappa or lambda expression. 23F-specific Abs had predominantly kappa chains. Isoelectric focusing analyses showed that sera contained one or at most a few immunoglobulin G Ab spectrotypes to all three respective capsular serotypes, a result indicative of oligoclonality. A sequence analysis of a purified PPS-14-specific Ab having a single spectrotype gave uniform amino-terminal sequences for both the heavy chain (V(H)III subgroup) and the L chain (kappaIII-A27 V region). From these results we conclude that within individual adults, serum Ab responses to PPS serotypes 6B, 14, and 23F derive from a small number of dominant B-cell clones, and consequently variable-region expression is probably individually limited as well. Oligoclonality appears to be a general characteristic of human PPS-specific Ab repertoires, and we suggest that this property could lead to individual differences in Ab fine specificity and/or functional activity against encapsulated pneumococci.     

Antigenic determinants of Staphylococcus aureus type 5 and type 8 capsular polysaccharide vaccines

Bacterial capsular polysaccharides (CP) are carbohydrate polymers comprised of repeating saccharide units. Several of these CP have side chains attached to their backbone structures. The side chains may include O-acetyl, phosphate, sialic acid, and other moieties. Those moieties represent the immunodominant epitopes and the most functional ones. The clinically significant Staphylococcus aureus type 5 CP (CP 5) and type 8 CP (CP 8) are comprised of a trisaccharide repeat unit with one O-acetyl group attached to each repeat unit. The immunogenicity of these CP and the functionality of antibodies to the backbone and the O-acetyl moieties were investigated. Immunization with the native CP conjugates (CP with 75% O-acetylation) elicited a high proportion of antibodies directed against the O-acetyl moiety. Nonetheless, all of the vaccinees produced antibodies to the backbone moieties as well. Conjugate vaccines made of de-O-acetylated CP elicited backbone antibodies only. Antibodies to both backbone and O-acetyl groups were found to be opsonic against S. aureus strains which varied in their O-acetyl content. Absorption studies with O-acetylated and de-O-acetylated CP showed that (i) native CP conjugates generated antibodies to both backbone and O-acetyl groups and (ii) O-acetylated isolates were opsonized by both populations of antibodies while the non-O-acetylated strains were predominantly opsonized by the backbone antibodies. These results suggest that S. aureus CP conjugate vaccines elicit multiple populations of antibodies with diverse specificities. Moreover, the antibodies of different specificities (backbone or O-acetyl) are all functional and efficient against the variations in bacterial CP that may occur among clinically significant S. aureus pathogenic isolates.     

1H-15N NMR signal assignment and secondary structure of bacteriorhodopsin (1-231) in solution

15N-Labeled de-(232-248)-bacteriorhodopsin [BR(1-231)] was solubilized in 1:1 chloroform-methanol solvent mixture that contained 1.0 M 2HCO2N2H4 and mimic membrane medium. Resonances in the 1H-15N heteronuclear multiple-quantum coherence (HMQC) spectrum of BR (1-231) were assigned using the data of two- and three-dimensional NMR experiments. Of 117 cross-peaks present in the 1H-15N HMQC spectrum, 98 were assigned to residues in 1-75 and 193-231 segments of the protein. Almost all cross-peaks that correspond to the 76-192 segment were absent in the HMQC spectrum (except for six cross-peaks from the side chains and 14 cross-peaks from the backbone). Deuterium exchange rates of amide protons and cross-peaks of nuclear Overhauser effect helped to localize helices A (residues 8-30), B (residues 40-65), and G (residues 198-226). The periodicity in the rates of deuterium exchange of NH protons of helices A, B, and G was explained by the compact arrangement of these helices in the protein globule. The broadening of signals from six residues in helix G, which, according to the electron cryomicroscopy model of bacteriorhodopsin, is in contact with the NMR-unobservable bundle of helices CDEF, indicates specific interactions of the helices in BR(1-231). These data suggest that BR(1-231) solubilized in an organic medium has a spatial structure similar to that in the electron cryomicroscopy model of BR.     

The three-dimensional NMR solution structure of alpha-cobratoxin at pH 7.5 and conformational differences with the NMR solution structure at pH 3.2

The 3D solution structure of alpha-cobratoxin, a neurotoxin purified from the Naja naja siamensis snake venom, has been determined by Nuclear Magnetic Resonance spectroscopy, in conjunction with distance geometry and restrained molecular dynamics, at pH 7.5. A total of 490 distance restraints were obtained from NOE intensities and 25 phi dihedral angle restraints deduced from J-coupling data. The generated structures are well defined with root mean square deviations from a geometrical mean structure of 0.107 +/- 0.036 nm for the backbone atoms and 0.128 +/- 0.073 nm for the side-chain atoms (considering residues 1 to 66 minus 26 to 35). A comparison between the generated structures at pH 7.5 and the mean NMR solution structure at pH 3.2 revealed that the 3D structure of alpha-cobratoxin is more compact at neutral pH. This major difference is mainly due to the pH-dependent conformational variations of three residues His18, Thr44 and Thr59.     

Structure determination of the capsular polysaccharide from Vibrio vulnificus strain 6353

Although significant progress has been made towards the understanding of cellular and molecular mechanisms underlying the pathogenetic pathways of transplant arteriosclerosis, its knowledge is still not comprehensive. Nevertheless, experimental and clinical studies have enabled us to discover some of the complex processes involved in the progression and evolution of transplant arteriosclerosis. Despite the advances in transplantation immunology and atherosclerosis research, transplant arteriosclerosis still remains a major cause of allograft failure. A curative treatment, in order to inhibit or at least modify the development of transplant arteriosclerosis, is urgently needed. This review article highlights some of the more recent aspects of cellular and molecular pathology of transplant arteriosclerosis that may add to our current and future diagnostic and curative interventions.     

Structural organization of the major autolysin from Streptococcus pneumoniae

LytA amidase is the best known bacterial autolysin. It breaks down the N-acetylmuramoyl-L-alanine bonds in the peptidoglycan backbone of Streptococcus pneumoniae and requires the presence of choline residues in the cell-wall teichoic acids for activity. Genetic experiments have supported the hypothesis that its 36-kDa chain has evolved by the fusion of two independent modules: the NH2-terminal module, responsible for the catalytic activity, and the COOH-terminal module, involved in the attachment to the cell wall. The structural organization of LytA amidase and of its isolated COOH-terminal module (C-LytA) and the variations induced by choline binding have been examined by differential scanning calorimetry and analytical ultracentrifugation. Deconvolution of calorimetric curves have revealed a folding of the polypeptide chain in several independent or quasi-independent cooperative domains. Elementary transitions in C-LytA are close but not identical to those assigned to the COOH-terminal module in the complete amidase, particularly in the absence of choline. These results indicate that the NH2-terminal region of the protein is important for attaining the native tertiary fold of the COOH terminus. Analytical ultracentrifugation studies have shown that LytA exhibits a monomer <--> dimer association equilibrium, through the COOH-terminal part of the molecule. Dimerization is regulated by choline interaction and involves the preferential binding of two molecules of choline per dimer. Sedimentation velocity experiments give frictional ratios of 1.1 for C-LytA monomer and 1.4 for C-LytA and LytA dimers; values that deviated from that of globular rigid particles. When considered together, present results give evidence that LytA amidase might be described as an elongated molecule consisting of at least four domains per subunit (two per module) designated here in as N1, N2, C1, and C2. Intersubunit cooperative interactions through the C2 domain in LytA dimer occur under all experimental conditions, while C-LytA requires the saturation of low affinity choline binding sites. The relevance of the structural features deduced here for LytA amidase is examined in connection with its biological function.     

Identification of a gene essential for O-acetylation of the Staphylococcus aureus type 5 capsular polysaccharide

It has been hypothesized that the developmental toxicity of certain compounds is, in part, due to maternal toxicity resulting in alterations in zinc (Zn) metabolism that affects the developing conceptus. In the present work the effects of developmentally toxic doses of 2-ethylhexanoic acid (EHXA), 2-ethylhexanol (EHXO), and valproic acid (VPA) on Zn metabolism were investigated in the pregnant rat. In experiment 1, dams were intubated with EHXA (3.13, 6.25, 9.38 or 12.5 mmol/kg), EHXO (6.25, 9.38 or 12.5 mmol/kg), VPA (1.56, 3.13, 6.25 or 9.38 mmol/kg), or corn oil (control; 1.0 ml/kg) at 14:00 h on gestation day (GD) 11.5, intubated with 32 microCi 65Zn at 22:00 h, and then killed at 08:00 h on GD 12.5. At the higher dose levels of EHXA and EHXO, and at all dosages of VPA, the percentage of 65Zn retained in maternal liver was higher, while that in the embryos was lower, than in controls. Chemical-associated changes in 65Zn distribution were associated with increased maternal liver metallothionein (MT) concentrations. In experiment 2, dams were fed diets containing 1, 25 or 97 microg Zn/g from GD 0-16 and intubated with 3.5 mmol EHXA or 1.0 ml corn oil/kg/d from GD 8-15. Dams were killed on GD 16 or 19. High incidences of encephalocele and tail defects were noted in the GD 16 fetuses of EHXA-treated dams fed either the low or adequate Zn diet, the highest incidences being in the low Zn group. On GD 19 the incidence of tail defects tended to be higher in the EHXA groups than in oil-treated controls, the highest incidence occurring in the low Zn EHXA group. Encephalocele was only observed in the low Zn EHXA-treated group. Fetal weight and crown-rump lengths were decreased by EHXA treatment and low dietary Zn. The incidence of rib anomalies was higher in the EHXA-exposed groups than in their respective oil controls. In experiment 3, GD 10.5 embryos collected from control dams were cultured for 48 h in serum from control or EHXA-treated male rats fed 4.5 or 25.0 microg Zn/g diets. Embryos cultured in either EHXA or low Zn sera exhibited delayed development; the addition of Zn to these sera eliminated their developmental toxicity. These results support the hypothesis that certain chemicals which induce maternal toxicity act, in part, to influence embryonic Zn metabolism and trigger abnormal development. Importantly, the teratogenic effects of these chemicals can be modulated by dietary Zn intake.     

An antigenic polysaccharide from Campylobacter coli serotype O:30. Structure of a teichoic acid-like antigenic polysaccharide associated with the lipopolysaccharide

A water-soluble antigenic polysaccharide of high M(r) associated with the lipopolysaccharide has been isolated from phenol-water extraction of cells of Campylobacter coli serotype O:30. The polysaccharide and oligosaccharide degradation products formed on O-dephosphorylation and by periodate oxidation followed by reduction have been investigated by one- and two-dimensional 1H, 13C, and 31P NMR. It is concluded that the antigenic polysaccharide has a teichoic acid-like structure with a poly-Ribitol phosphate, [5-Ribitol-1-P]n, backbone with side chains at O-2 of O-(6-deoxy-beta-D-talo-heptopyranosyl)-(1-->4)-(2-acetylamino-2-deoxy-beta-D- glucopyranosyl) units. The structure is unusual in Gram-negative bacteria and is unique in possessing 6-deoxy-D-talo-heptose as a constituent sugar. Evidence for the relationship of the antigenic polysaccharide to the lipopolysaccharide of low M(r) is discussed.     

Structure of the capsular polysaccharide from the Klebsiella K8 reference strain 1015

The structure of the capsular polysaccharide from the Klebsiella K8 reference strain 1015 has been elucidated. The structure was deduced from sugar analysis, different methylation analyses, a uronic acid degradation, and NMR spectroscopy. It is concluded that the polysaccharide is composed of pentasaccharide repeating units with the structure: [formula: see text] The structure differs from that of the previously published structure of the capsular polysaccharide from Klebsiella K8, which originates from another strain and has the following structure: [formula: see text] The serological similarity between the two strains is most likely derived from a common tetrasaccharide which is substituted in different ways in the two strains. Since the strain in the present investigation originates from the Klebsiella K reference strain collection of the International Escherichia and Klebsiella Centre, Copenhagen, Denmark, it is suggested that it should keep the designation K8. The other polysaccharide with Klebsiella K8 specificity should be renamed as K8,52,59 based on the cross-reactivity of the strain (I. Orskov, unpublished).