SEARCH · PubMed Health

Results for “HEPTOSES”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 145 records · Page 8Linked to original sources

PubMed2004

Characterization of the alpha-1,2-N-acetylglucosaminyltransferase of Neisseria gonorrhoeae, a key control point in lipooligosaccharide biosynthesis.

The biosynthesis of the lipooligosaccharide (LOS) in Neisseria meningitidis has a control point that regulates the extension of the alpha-chain on heptose (I) of the LOS. The gene that encodes the protein responsible for this control had been identified elsewhere, but the enzyme encoded by the gene was not characterized. We have now shown that this same control mechanism operates in the related species, Neisseria gonorrhoeae, using a gene knockout and subsequent characterization of the LOS species produced. We also cloned and expressed the enzyme from both of these pathogens. Using a synthetic acceptor substrate, we have shown unequivocally that the enzyme is an alpha-1,2-N-acetylglucosaminyltransferase. Experiments with both the core oligosaccharide and the synthetic acceptors suggests that the addition of the alpha-1,2-N-acetylglucosamine moiety on the heptose (II) residue precedes the addition of the ethanolamine phosphate at the O3 position on this heptose (II), and that in the absence of the alpha-1,2-N-acetylglucosamine moiety leads to the addition of an extra ethanolamine phosphate on the heptose (II) residue. Our data do not support the hypothesis that ethanolamine phosphate at O3 of heptose (II) is added and is then required for the addition of the N-acetylglucosamine at O2 by the LgtK enzyme. This enzyme represents a control point in the biosynthesis of the LOS of this pathogen and is a potential target for therapeutic intervention.

Carbohydrate Conformation↗

PubMed2004

Three genes, lgtF, lic2C and lpsA, have a primary role in determining the pattern of oligosaccharide extension from the inner core of Haemophilus influenzae LPS.

Lipopolysaccharide (LPS) is a virulence determinant of Haemophilus influenzae and exhibits substantial heterogeneity in structure within and between strains. Key factors contributing to this heterogeneity are the genes required to add the first glycose to each of the three heptose residues of the LPS inner core. In each case this addition can facilitate further oligosaccharide extension. lgtF is invariably present in strains and the product has a function in adding the glucose to the first heptose. lic2C is present in half the strains and was found to add a glucose to the second heptose. Insertion of lic2C into a strain that does not naturally contain it resulted in hexose incorporation from the second heptose of the LPS. The product of the lpsA gene can add a glucose or galactose to the third heptose. By allelic replacement of lpsA between strains it is shown that the sequence of the gene can be the sole determinant of this specificity. Thus, lgtF, lic2C and lpsA make significant but very distinct contributions to the conservation and variable patterns of oligosaccharide extensions seen in H. influenzae LPS.

Bacterial Proteins↗

PubMed1984

Structure of the lipopolysaccharide core isolated from a human strain of Aeromonas hydrophila.

Lipopolysaccharide was isolated from the cell-walls of a human strain of Aeromonas hydrophila by the aqueous phenol method in 0.58% yield (based on dry weight of bacteria). The lipopolysaccharide consisted of SR-polysaccharide, core-oligosaccharide and lipid A; there was no O-specific polysaccharide. The core had the composition D-galactose, D-glucose, D-glycero-D-manno-heptose, L-glycero-D-manno-heptose and D-glucosamine in a molar ratio of 1:1:2:4:1. Glucosamine was linked to an L-glycero-D-manno-heptose residue by a bond which was resistant to hydrolysis. The D-glucosamine-(1----7)-LD-heptose disaccharide was isolated and identified by the mass spectrum of its methylated alditol and the heptose residue not observed under normal hydrolysis conditions was easily determined after deamination of the complete core. Methylation analysis, chemical degradation, periodate and chromium trioxide oxidations and nuclear magnetic resonance (13C and 1H NMR) spectroscopy were used to identify the structure of the core oligosaccharide as: (formula: see text)

Aeromonas↗

PubMed1996

Lipopolysaccharide core glycosylation in Rhizobium leguminosarum. An unusual mannosyl transferase resembling the heptosyl transferase I of Escherichia coli.

The lipopolysaccharide structure of the nitrogen-fixing bacterium Rhizobium leguminosarum differs from that of Escherichia coli in several ways, one of which is the sugar composition of the core. The E. coli inner core consists of 3-deoxy-D-manno-octulosonic acid (Kdo) and L-glycero-D-manno-heptose (heptose), while the inner core of R. leguminosarum contains 2-keto-3-deoxy-D-manno-octulosonic acid (Kdo), mannose, galactose, and galacturonic acid. The two Kdo residues and their linkages appear to be identical in both species. The linkages of heptose in E. coli and of mannose in R. leguminosarum to Kdo are both alpha1-5. We now characterize a membrane-associated glycosyl transferase in R. leguminosarum extracts that incorporates mannose into nascent lipopolysaccharide, using Kdo2-lipid IVA as the acceptor and GDP-mannose (or synthetic ADP-mannose) as the donor. The mannosyl transferase is associated with the inner membrane. The apparent Km values for GDP-mannose and Kdo2-lipid IVA are 4.3 microM and 7.1 microM, respectively, in the presence of excess co-substrate. Extracts of E. coli do not catalyze GDP-mannose-dependent glycosylation of Kdo2-lipid IVA, but they are active when ADP-mannose is substituted for GDP-mannose. Given the structural similarity of ADP-mannose to ADP-heptose, we examined the possibility that heptosyl transferase I of E. coli (the product of the rfaC gene) catalyzes mannose transfer from ADP-mannose to Kdo2-lipid IVA. Extracts of E. coli mutants defective in the rfaC gene are unable carry out ADP-mannose-dependent glycosylation of Kdo2-lipid IVA. Plasmids bearing rfaC+ not only restore the missing activity but also direct its overexpression. Our assay using ADP-mannose as a substitute for ADP-heptose (which is not readily available) should facilitate the purification and characterization of heptosyl transferase I of E. coli. The GDP-mannose-dependent enzyme of R. leguminosarum may represent a functional equivalent of E. coli RfaC.

Escherichia coli↗

PubMed1971

Lipopolysaccharide and aldoheptose biosynthesis in transketolase mutants of Salmonella typhimurium.

Genetic and biochemical evidence that sedoheptulose-7-phosphate is an obligatory precursor of the L-glycero-D-mannoheptose residues of the lipopolysaccharide of Salmonella was obtained by isolation and characterization of transketolase-negative mutants of Salmonella typhimurium. These mutants, which are defective in synthesis of sedoheptulose-7-phosphate, were found to produce an incomplete heptose-deficient lipopolysaccharide, and were also sensitive to bile salts, a characteristic property of heptose-deficient mutants. Phenotypic repair of the defect in lipopolysaccharide synthesis was obtained by addition of exogenous sedoheptulose-7-phosphate to growing cultures of the mutant strains. Characterization of revertants isolated either as transketolase-positive or heptose-positive provided further evidence that the heptose deficiency resulted from mutation at the transketolase locus. On the basis of these findings a possible pathway for conversion of sedoheptulose-7-phosphate to L-glycero-D-mannoheptose is proposed.

Carbon Isotopes↗

PubMed2001

Molecular cloning and functional expression of the rfaE gene required for lipopolysaccharide biosynthesis in Salmonella typhimurium.

The rfaE (WaaE) gene of Salmonella typhimurium is known to be located at 76min on the genetic map outside of the rfa gene cluster encoding core oligosaccharide biosynthesis of lipopolysaccharide(LPS). The rfaE mutant synthesizes heptose-deficient LPS; its LPS consists of only lipid A and 3-deoxy-D-manno-octulosonic acid (KDO), and the rfaE gene is believed to be involved in the formation of ADP-L-glycero-D-manno-heptose. Mutants, which make incomplete LPS, are known as rough mutants. Salmonella typhimurium deep-rough mutants affected in the heptose region of the inner core often show reduced growth rate, sensitivity to high temperature and hypersensitivity to hydrophobic antibiotics. We have cloned the rfaE gene of S. typhimurium. The chromosomal region carrying this gene was isolated by screening a genomic library of S. typhimurium using the complementation of S. typhimurium rfaE mutant. The 2.6-Kb insert in the plasmid pHEPs appears to carry a functional rfaE gene. SL1102 (rfaE543) makes heptose-deficient LPS and has a deep rough phenotype, but pHEPs complement the rfaE543 mutation to give the smooth phenotype. The sensitivity of SL1102 to bacteriophages (P22.c2, Felix-O, Br60) which use LPS as their receptor for adsorption is changed to that of wild-type strain. The permeability barrier of SL1102 to hydrophobic antibiotics (novobiocin) is restored to that of wild-type. LPS produced by SL1102 (rfaE543) carrying pHEPs makes LPS indistinguishable from that of smooth strains. The rfaE gene encoded a polypeptide of 477 amino acid residues highly homologous to the S. enterica rfaE protein (98% identity), E. coli (93% identity), Yersenia pestis (85% identity), Haemophilus influenzae (70% identity) and Helicobacter pyroli (41% identity) with a molecular weight 53 kDa.

Amino Acid Sequence↗

PubMed1977

Characterization of lipid A and polysaccharide moieties of the lipopolysaccharides from Vibrio cholerae.

Lipid A and polysaccharide moieties obtained by mild acid hydrolysis of the lipopolysaccharides from Vibrio cholerae 569 B (Inaba) and Vibrio el-tor (Inaba) were characterized. Heterogeneity of lipid A fractions was indicated by t.l.c. and by gel filtration of the de-O-acylated products from mild alkaline methanolysis of the lipids. Presumably lipid A contains a glucosamine backbone, and the fatty acids are probably bound to the hydroxyl and amino groups of glucosamine residues. Approximately equal amounts of fatty acids C16:0, C18:1 and 3-hydroxylauric acid were involved in ester linkages, but 3-hydroxymyristic acid was the only amide-linked fatty acid. Sephadex chromatography of the polysaccharide moiety showed the presence of a high-molecular-weight heptose-free fraction and a low-molecular-weight heptose-containing fraction. Haemagglutination-inhibition assays of these fractions showed the heptose-free fraction to be an O-specific side-chain polysaccharide, whereas the heptose-containing fraction was the core polysaccharide region of the lipopolysaccharides. Identical results were obtained for both organisms.

Chromatography, Gel↗

PubMed1999

Isolation and structural analysis of phosphorylated oligosaccharides obtained from Escherichia coli J-5 lipopolysaccharide.

The chemical structure of the phosphorylated lipopolysaccharide (LPS) of Escherichia coli J-5 was investigated because it is of biomedical interest in the context of septic shock, a syndrome often encountered in nosocomial infections with gram-negative pathogens. The successive de-O-acylation and de-N-acylation of J-5 LPS yielded phosphorylated oligosaccharides which represent the complete carbohydrate backbone. Five compounds were separated by high-performance anion-exchange chromatography and analysed by one-dimensional and two-dimensional homonuclear and heteronuclear 1H-NMR, 13C-NMR and 31P-NMR spectroscopy. The main product was a nonasaccharide of the structure alpha-D-Glcp-(1-->3)-[alpha-D-GlcpN- (1-->7)-alpha-L,D-Hepp-(1-->7)]-alpha-L,D-Hepp-(1-->3)-alpha -L, D-Hepp-4P-(1-->5)-[alpha-Kdop-(2-->4)]-alpha-Kdop-(2-- >6)-beta-D-GlcpN-4p- (1-->6)-alpha-D-GlcN-1P wherein all sugars are present as D-pyranoses. Hep and Kdo represent L-glycero-D-manno-heptose and 3-deoxy-D-manno-oct-2-ulosonic acid, respectively. In addition, two octasaccharides and two heptasaccharides were isolated that were partial structures of the nonasaccharide. In one octasaccharide the terminal alpha-D-GlcpN was missing and an additional phosphate group linked to O4 of the branched heptose was present, whereas in the other octasaccharide the side-chain Kdo was missing. In both heptasaccharides the side-chain alpha-D-GlcpN-(1-->7)-L-alpha-D-Hepp-disaccharide was absent; they differed in their phosphate substitution. Whereas both heptasaccharides contained two phosphates in the lipid-A backbone (beta-1,6-linked GlcpN-disaccharide at the reducing end) and one phosphate group at O4 of the first heptose, only one of them was additionally substituted with phosphate at O4 of the second heptose.

Carbohydrate Sequence↗

PubMed1989

Structure and heterogeneity of the oligosaccharides from the lipopolysaccharides of a pyocin-resistant Neisseria gonorrhoeae.

The compositions and partial structures of the oligosaccharides from the lipopolysaccharides (LPS) of a pyocin-resistant Neisseria gonorrhoeae (strain JW31R) have been determined by liquid secondary ion mass spectrometry (LSIMS), tandem mass spectrometry, and methylation analysis. Four major structures were identified with Mr 2123, 2000, 1961, and 1838, as well as seven species of lower abundance of Mr 1758-1272. The largest of the major oligosaccharides (Mr, 2122) consists of 3-deoxymanno-2-ketooctulosonic acid (KDO)-Hep2GalNAcGlcNAcGal4Glc2 (Hep, heptose) and phosphoethanolamine (PEA). The smaller oligosaccharides are truncated versions of this larger oligosaccharide. The oligosaccharides consist of a common triantennary structure containing KDO at the reducing terminus attached to a heptose disaccharide. A hexose (Hex)2-3 branch is attached to the heptose linked directly to KDO and a GalNAc-Hex3, GlcNAc, and PEA are separately attached to the second heptose. These oligosaccharides are the first structures to be determined for a gonococcal LPS and should further our understanding of the structural and antigenic diversity of these glycolipids.

Bacteriocins↗

PubMed1986

Comparison of bacterial lipopolysaccharides by high-performance liquid chromatography.

A comparison of lipid-free polysaccharides from gram-negative bacteria was rapidly accomplished by using high-performance liquid chromatography of underivatized hydrolysates. Examination of a number of such products revealed that, contrary to earlier reports, Xanthomonas campestris lipopolysaccharide contained heptose, together with rhamnose and galactose, but not mannose. The polymers from the methanotrophs "Methylomonas albus" and "Methylosinus trichosporium" contained heptose and glucose, and that from a "Klebsiella aerogenes" strain contained heptose, glucose, and galactose. The absence of heptose from the lipopolysaccharide of Myxococcus xanthus was confirmed.

Journal Article↗

PubMed1990

Characterization of anti-core glycolipid monoclonal antibodies with chemically defined lipopolysaccharides.

Five anti-core glycolipid monoclonal antibodies (MAb) (four against Escherichia coli J5 lipopolysaccharide [LPS] and one against the Re core glycolipid of Salmonella typhimurium) were characterized using LPS from several rough and smooth strains and derivatives of E. coli J5 LPS, obtained by N acetylation and hydrolysis. The MAb against E. coli J5 were not only weakly cross-reactive with clinical isolates, whereas the anti-Re MAb was highly cross-reactive. The MAb differed in their reaction pattern with E. coli J5 LPS. MAb 4-7B5 (immunoglobulin M) and MAb 4-6A1 (immunoglobulin G1) cross-reacted with LPS of Salmonella minnesota R5 and S. typhimurium Ra and Rc and little with Re and lipid A. The dominant binding site of these MAb was located in the glucose-heptose-heptose region and was independent of phosphate substitution. The MAb 4-9A1 reacted with the terminal part of the core region (glucose-heptose) and was dependent on phosphate substitution of the LPS. The MAb BA7 (immunoglobulin G3) was E. coli J5 LPS specific and reacted with the glucosaminyl-heptose disaccharide. Antibody 8-2C1 was directed against the common parts of LPS, 3-deoxy-D-manno-octulosonic acid, and lipid A, which are not (or only weakly) recognized by the four anti-J5 LPS MAb. Thus, MAb that are not cross-reactive can be directed against at least three different antigenic determinants present on the core oligosaccharide of E. coli J5 LPS.

Antibodies, Monoclonal↗

PubMed1997

Characterization of a transposon Tn916-generated mutant of Haemophilus ducreyi 35000 defective in lipooligosaccharide biosynthesis.

To define the role of the surface lipooligosaccharide (LOS) of Haemophilus ducreyi in the pathogenesis of chancroid, Tn916 mutants of H. ducreyi 35000 defective in expression of the murine monoclonal antibody (MAb) 3F11 epitope on H. ducreyi LOS were identified by immunologic screening. One mutant, designated 1381, has an LOS which lacks the MAb 3F11 epitope and migrates with an increased mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The gene disrupted by the Tn916 element in strain 1381 was identified by cloning the sequences flanking the Tn916 element. The sequences were then used to probe a lambda DASHII genomic library. In strain 1381, Tn916 interrupts a gene which encodes an open reading frame (ORF) with an Mr of 40,246. This ORF has homology to the product of the rfaK gene of Escherichia coli. The major LOS glycoform produced by strain 1381 was analyzed by using a combination of mass spectrometry, linkage and composition analysis, and 1H nuclear magnetic resonance spectroscopy. The major LOS species was found to terminate in a single glucose attached to the heptose (L-glycero-D-manno-heptose, or Hep) trisaccharide core. In the wild-type strain 35000, glucose serves as the acceptor for the addition of the D-glycero-D-manno-heptose (or DDHep), which extends to form the mature branch of the H. ducreyi LOS. This mature oligosaccharide is in turn partially capped by the addition of sialic acid (NeuAc), i.e., NeuAc2 alpha-->3Gal beta1-->4GlcNAc beta1-->3Gal beta1-->4DDHep alpha1-->6Glc beta1 (W. Melaugh et al., Biochemistry 33:13070-13078, 1994). Since this LOS terminates prior to the addition of the branch DD-heptose, this gene is likely to encode the D-glycero-D-manno-heptosyltransferase. Strain 1381 exhibits a significant reduction in adherence to and invasion of primary human keratinocytes. This defect was complemented by the cloned heptosyltransferase gene, indicating that the terminal portion of the LOS oligosaccharide plays an important role in adherence to human keratinocytes.

Bacterial Adhesion↗

PubMed1976

Influence of temperature on growth of lipopolysaccharide-deficient (rough) mutants of Salmonella typhimurium and Salmonella minnesota.

Smooth strains of Salmonella typhimurium and S. minnesota, and chemotypes Ra, Rb, and Rc, which are deficient in lipopolysaccharide components of the somatic side chains and outer core region, grow normally on nutrient agar and nutrient broth up to 45 degrees C. However, most mutants with defects in the heptose region of the LPS (chemotypes Rd2 and Re) do not grow on this medium at 42 degrees C or above; a few grow at 42 degrees C but not at 45 degrees C. In liquid medium (nutrient broth, or phosphate minimal medium), growth, measured as turbidity or as colony-forming units, stops 60 to 90 min after shift from 30 to 42 degrees C; DNA and protein synthesis cease at the same time. Growth does not reoccur at 42 degrees C; protein synthesis and growth reinitiate upon shift to 30 or 37 degrees C. Growth cessation does not alter cell morphology in the phase-contrast microscope. Growth of heptose-deficient strains at 42 degrees C in nutrient broth is restored by MgCl2 (0.5 mM), NaCl (50 mM), or sucrose (100 mM). Sensitivity to smooth-specific and rough-specific phages, and analysis of LPS composition, indicate that heptose-deficient mutants grown at temperatures from 30 to 45 degrees C, and in the presence or absence of high salt, do not contain heptose or O-specific sugars in their LPS.

Bacterial Proteins↗

PubMed1992

Partial characterization of the major lipooligosaccharide from a strain of Haemophilus ducreyi, the causative agent of chancroid, a genital ulcer disease.

The first preliminary structure of a surface lipooligosaccharide from Haemophilus ducreyi has been determined. The major oligosaccharide was released by mild acid hydrolysis and analyzed by liquid secondary ion and tandem mass spectrometry. The mass spectral data combined with composition and methylation analysis yielded the most probable structure; Gal1----4GlcNAc1----3Gal1----4Hep1----6Glc1----( Hep1----2Hep1----)3,4Hep1---- KDO, where the reducing terminal 3-deoxy-D-manno-octulosonic acid (or KDO) exists in an anhydro form. This anhydro species results from the elimination of a phosphate from C-4 of KDO during mild acid hydrolysis. The core heptose trisaccharide consists of L-glycero-D-manno-heptose, but analysis of the peracetylated sugars indicated that the 1,4-linked heptose is likely D-glycero-D-manno-heptose. The monoclonal antibody 3F11 generated against Neisseria gonorrhoeae also binds to this lipooligosaccharide and suggests that the terminal trisaccharide is Gal beta 1----4GlcNAc beta 1----3Gal beta 1----, an epitope found in the glycose moiety of the human erythrocyte glycosphingolipid lactoneotetraglycosylceramide. Mass spectrometric and composition analysis of the lipid A moiety shows that it is similar to the lipid A of Haemophilus influenzae strain I-69 Rd-/b+ proposed by Helander et al. (Helander, I. M., Lindner, B., Brade, H., Altmann, K., Lindberg, A. A., Rietschel, E. T., and Zähringer, U. (1988) Eur. J. Biochem. 177, 483-492). Electrospray mass spectrometric analysis of the intact O-deacylated lipooligosaccharides gave an average Mr of 2710, and supported an overall structure consisting of the above nonasaccharide linked directly to a diphosphorylated lipid A moiety through the single KDO which is phosphorylated. This structure should provide a framework to investigate the roles of lipooligosaccharides in the host immunochemical response and pathology of H. ducreyi infection, a leading cause of genital ulcer disease.

Carbohydrate Sequence↗

PubMed2005

A phosphoethanolamine transferase specific for the outer 3-deoxy-D-manno-octulosonic acid residue of Escherichia coli lipopolysaccharide. Identification of the eptB gene and Ca2+ hypersensitivity of an eptB deletion mutant.

Addition of a phosphoethanolamine (pEtN) moiety to the outer 3-deoxy-D-manno-octulosonic acid (Kdo) residue of lipopolysaccharide (LPS) in WBB06, a heptose-deficient Escherichia coli mutant, occurs when cells are grown in 5-50 mM CaCl2 (Kanipes, M. I., Lin, S., Cotter, R. J., and Raetz, C. R. H. (2001) J. Biol. Chem. 276, 1156-1163). A Ca2+-induced, membrane-bound enzyme was responsible for the transfer of the pEtN unit to the Kdo domain. We now report the identification of the gene encoding the pEtN transferase. E. coli yhjW was cloned and overexpressed, because it is homologous to a putative pEtN transferase implicated in the modification of the beta-chain heptose residue of Neisseria meningitidis lipo-oligosaccharide (Mackinnon, F. G., Cox, A. D., Plested, J. S., Tang, C. M., Makepeace, K., Coull, P. A., Wright, J. C., Chalmers, R., Hood, D. W., Richards, J. C., and Moxon, E. R. (2002) Mol. Microbiol. 43, 931-943). In vitro assays with Kdo2-4'-[32P]lipid A as the acceptor showed that YhjW (renamed EptB) utilizes phosphatidylethanolamine in the presence of Ca2+ to transfer the pEtN group. Stoichiometric amounts of diacylglycerol were generated during the EptB-catalyzed transfer of pEtN to Kdo2-lipid A. EptB is an inner membrane protein of 574 amino acid residues with five predicted trans-membrane segments within its N-terminal region. An in-frame replacement of eptB with a kanamycin resistance cassette rendered E. coli WBB06 (but not wild-type W3110) hypersensitive to CaCl2 at 5 mM or higher. Ca2+ hypersensitivity was suppressed by excess Mg2+ in the medium or by restoring the LPS core of WBB06. The latter was achieved by reintroducing the waaC and waaF genes, which encode LPS heptosyl transferases I and II, respectively. Our data demonstrate that pEtN modification of the outer Kdo protected cells containing heptose-deficient LPS from damage by high concentrations of Ca2+. Based on its sequence similarity to EptA(PmrC), we propose that the active site of EptB faces the periplasmic surface of the inner membrane.

Biological Transport↗

PubMed1987

Chemical composition and immunological characterization of polysaccharide antigen from Eubacterium saburreum T18.

Two types of polysaccharide were obtained from the oral microorganism Eubacterium saburreum T18 by formamide extraction and subsequent gel filtration and ion-exchange chromatography. One polysaccharide, which was composed of D-glycero-D-galacto-heptose, had antigenic activity in an immunoprecipitation reaction with rabbit anti-T18 serum due to immunoglobulin M antibodies. The second polysaccharide was composed of D-glycero-D-manno-heptose and L-rhamnose, but it did not have immunoprecipitation activity. These polysaccharide antigens were not alkali labile and differed from E. saburreum L44 and T27 antigens, which were composed of D-glycero-D-galacto-heptose.

Antibodies, Bacterial↗

PubMed1975

Blocking of bacteriophages phi W and phi 5 with lipopolysaccharides from Escherichia coli K-12 mutants.

In the preceding paper we presented a formula for the composition of lipopolysaccharides (LPS) from Escherichia coli K-12. This formula contains four regions defined from analyses of LPS from four key strains, the parent and mutants which had lost one, two, or three regions of their carbohydrates. Support for the formula was derived from the susceptibility of the key mutants to several bacteriophages. One of these, phage phi W, was found specific for strains which had lost region 4. In this paper we described inactivation in vitro of phage phi W and its host-range mutant phi 5, using LPS devoid of regions 2 to 4. The blocking of phi W was found to require about 0.15 M concentrations of monovalent cations and to be inhibited by low concentrations of calcium and magnesium ions. One particle of phage phi W required 2 times 10-16 g of LPS devoid of region 4 for stoichiometric inactivation. Phage phi 5 was blocked by both heptose-less LPS (devoid of regions 2 to 4) and glucose-less LPS (devoid of regions 3 to 4) but was unaffected by LPS devoid of region 4. LPS from a heptose-less mutant of Salmonella minnesota showed the same inactivation ability as did LPS from heptose-less strains of E. coli K-12. Lipid A was prepared from LPS containing all four regions. Such lipid A was found to inactivate phi 5, whereas both the polysaccharide moiety as well as the intact LPS were without effect. It is suggested that lipid A is part of the receptor site for phage phi 5.

Adsorption↗

PubMed1965

BIOCHEMICAL STUDIES OF THE SMOOTH-ROUGH MUTATION IN SALMONELLA MINNESOTA.

Lüderitz, O. (Max-Planck-Institut für Immunbiologie, Frieburg, Germany), H. J. Risse, H. Schulte-Holthausen, J. L. Strominger, I. W. Sutherland, and O. Westphal. Biochemical studies of the smooth-rough mutation in Salmonella minnesota. J. Bacteriol. 89:343-354. 1965.-A comparative study of the O antigen from the smooth strain of Salmonella minnesota and of the two R antigens derived from two rough forms of S. minnesota (strains R 60 and R 345) has been carried out. The O-specific polysaccharide of the smooth form is composed of heptose, galactose, glucose, glucosamine, galactosamine, and ketodeoxyoctanoate (KDO). R 60 polysaccharide contains KDO, heptose, galactose, glucose, and glucosamine, whereas the R 345 polysaccharide contains only KDO, heptose, galactose, and glucose. Serologically, R 345 and R 60 polysaccharides belong to serogroups R I and R II, respectively. Enzymatic studies revealed that the acetylgalactosamine-synthesizing enzyme, uridine diphosphate-N-acetylglucosamine-4-epimerase, is present in wild-type and R 345 cells but is absent from R 60 cells. Two distinct polysaccharides were obtained from the R 345 cells: a polysaccharide derived from the R antigen (lipopolysaccharide) containing no galactosamine and exerting R specificity, and a soluble polysaccharide containing galactosamine and exerting O specificity. The structure of O and R antigens is discussed, together with the general significance of the results for the biosynthesis of the O antigens of the genus Salmonella.

Carbohydrates↗