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  • 1
    ISSN: 1432-072X
    Keywords: Rhodobacter capsulatus ; Protein export ; Porin ; Protein folding ; Phototrophic bacteria
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Export of porin to the outer membrane of the phototrophic purple bacterium Rhodobacter capsulatus was studied with the use of the uncoupler of the electron transport chain, carbonylcyanide-m-chlorophenylhydrazone (CCCP). The agent reversibly blocked the transport of porin across the cytoplasmic membrane. By means of radioactive labeling and immunoprecipitation, porin was found to occur in two forms: (i) the exported form that was extractable from the outer membrane without disrupting the cells, and (ii) a pre-form with a slightly higher apparent molecular mass which accumulated in the cells during the block of the export process. Proteolysis studies revealed that the preform was highly sensitive to added proteases, whereas the exported form was resistant.
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  • 2
    ISSN: 1432-072X
    Keywords: Cyanobacterium ; Fischerella ; 3-Hydroxy fatty acids ; Lipopolysaccharide ; Outer membrane ; Sheath
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A purified sheath fraction and an outer membrane fraction were obtained from the cyanobacterium Fischerella sp. PCC 7414. The sheath had a fine structure with osmiophilic fibers running in parallel to the cell surface in two distinct layers. The sheath fraction contained mainly neutral sugars (Glc, Man, Gal, Xyl, Fuc, 2-O-methylhexose), GlcN, uronic acids, and minor components such as amino acids, sulfate, phosphate, and fatty acids. The protein moiety was removable from the sheath fraction by treatment with boiling sodium dodecyl sulfate. The presence of three different 3-hydroxy fatty acids (3-OH-14:0, 3-OH-16:0, 3-OH-18:0) in addition to GlcN indicated the presence of lipopolysaccharide in the outer membrane. One major (Mr 50,000) and two minor (Mr 54,000 and 65,000) proteins were detected as constituents of the outer membrane.
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  • 3
    ISSN: 1432-072X
    Keywords: Fast growing rhizobia ; Lipid A ; 3-Hydroxy-octadecenoic acid ; 27-Hydroxyoctacosanoic acid ; Rhizobium meliloti
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The structure of the lipopolysaccharide from Rhizobium meliloti 10406, a derivative of the wild-type strain MVII-1, was examined. The compositional analysis of its polysaccharide moiety demonstrated lack of heptose(s), but high contents in glucose, galacturonic acid and 2-keto-3-deoxy-octonate (dOclA) as characteristic features. The lipid A moiety consisted of a β-1,6 linked glucosamine disaccharide carrying ester (at C-4′) and glycosidically (at C-1) linked phosphate residues, both present exclusively as monoester phosphates but not as phosphodiesters. Ester- and amidelinked 3-hydroxy fatty acids were mostly present as non-3-O-acylated residues. Laser desorption mass spectrometry (LD-MS) revealed heterogeneity in the fatty acid substitution, as was also indicated by the non-stoichiometric ratios obtained by quantitative fatty acid analysis. The predominating lipid A structure contained at the reducing glucosamine residue ester-linked 3-hydroxy-tetradecanoic acid (3-OH-14:0) and amide-linked 3-OH-18:0, or 3-OH-18:1, respectively. The distal (non-reducing) glucosamine carried ester-bound the recently discovered 27-hydroxyoctacosanoic acid and 3-OH-14:0 and, as amide-linked fatty acid, mostly 3-hydroxy-stearic acid (3-OH-18:0). The isolated lipopolysaccharide exhibited a high extent of lethal toxicity in galactosamine-treated mice, comparable to that of enterobacterial lipopolysaccharide. The structural relationship of LPS and lipid A of Rhizobium meliloti to other rhizobial lipopolysaccharides and lipid A's with respect to questions of taxonomy and of phylogenetic relationships will be discussed.
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  • 4
    ISSN: 1432-072X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary 1. The isolation and chemical characterization of the lipopolysaccharides (O-antigens) of 12 strains of the gram-negative photosynthetic bacterium Rhodopseudomonas palustris is described. Lipopolysaccharides (LPS) were extractable with phenol/water, however, the bulk of the LPS of all strains remained in the phenol phase and only trace amounts were found in the aqueous layer. The LPS was also extractable by a phenol/chloroform/petroleum ether mixture (PCP-method), recommended for lipophilic glycolipids. Neither incubation of living bacteria with EDTA nor with NaCl liberated appreciable amounts of LPS-protein-lipid conjugates from the cells. 2. All strains investigated were found to have galactose, mannose, heptose, 2-keto-3-deoxyoctonate (KDO), glucosamine, 6-deoxy-glucosamine (quinovosamine) and a recently identified sugar, a 2,3-diamino-2,3-dideoxyhexose, as common LPS constituents. The presence of additional sugars allowed the classification of the strains into three distinct chemotypes. Chemotype I contains 4-O-methyl-D-xylose, and several non-identified amphoteric amino sugars. Chemotype II contains 4-O-methyl-D-xylose, 3-O-methyl-6-deoxy-D-talose, 6-deoxy-talose, xylose and again some unidentified amphoteric amino sugars, which were different from those of chemotype I. In chemotype III xylose, glucose, rhamnose, galactosamine and 6-O-methyl-glucosamine were identified. The main fatty acid in the high molecular weight material from the phenol phase of phenol/water extracts of all strains is β-hydroxymyristic acid. In addition in all strains β-hydroxypalmitic, palmitic and stearic acids were found. It has still to be proven that all these fatty acids are LPS constituents. 3. Like enteric LPS the LPS of R. palustris can be split by mild acid hydrolysis in a lipid portion (lipid A) and the degraded polysaccharide. But contrary to enteric lipid A, the lipid A of R. palustris does not contain glucosamine, but has the 2,3-diamino-hexose as the only amino sugar constituent. The possible occurrence of a common R-core and the question if repeating units exist in the O-specific chains of R. palustris LPS are discussed. 4. In two strains small amounts of additional LPS, which differ in their chemical composition from the respective LPS of the phenol phase, were isolated from the aqueous phase.
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  • 5
    ISSN: 1432-072X
    Keywords: Rhodospirillum rubrum ; Rhodospirillum molischianum ; Rhodopila globiformis ; Lipopolysaccharide ; Lipid A ; 2,3-Diamino-glucose
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The cell wall lipopolysaccharides from three phototrophic species of the alpha1-group of Proteobacteria, Rhodospirillum rubrum, Rhodospirillum molischianum, and Rhodopila globiformis were isolated and chemically characterized. Sodium deoxycholate polyacrylamide gel electrophoresis patterns revealed that the lipopolysaccharides of all three species possess O-chains. They are composed of repeating units only in R. molischianum and R. globiformis. The presence of l-glycero-d-mannoheptose and 2-keto-3-deoxyoctonate indicated core structures in all three lipopolysaccharides. Glucosamine was found as backbone amino sugar in lipid A of R. molischianum and R. rubrum, while R. globiformis has 2,3-diaminoglucose as backbone amino sugar. The latter species also differed from the two former ones in its content of hydroxy fatty acids (3-OH-14:0, 3-OH-16:0 in R. rubrum and R. molischianum and 3-OH-14:0, 3-OH-18:0 and 3-OH-19:0 (possibly iso- or anteisobranched) in R. globiformis).
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  • 6
    ISSN: 1432-072X
    Keywords: Key wordsRhodospirillum salinarum ; Lipopolysaccharide ; Mixed lipid A ; 2 ; 3-Diamino-2 ; 3-dideoxy-d-glucose ; 4-O-Methyl- ; galacturonic acid ; Halophilic bacteria ; Lethal toxicity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The structural elucidation of lipid A of the cell wall lipopolysaccharide (LPS) of Rhodospirillum salinarum 40 by chemical methods and laser desorption mass spectrometry revealed the presence of a mixed lipid A composed of three different 1,4′bisphosphorylated β(1→6)-linked backbone hexosaminyl-hexosamine disaccharides, i.e. those composed of GlcN→GlcN, 2,3-diamino-2,3-dideoxy-d-Glc-(DAG)→DAG, and DAG→GlcN. Lipid A of R. salinarum contained preferentially 3-OH-18 : 0 and 3-OH-14 : 0 as amide-linked and cisΔ11–18 : 1 and c19 : 0 as ester-linked fatty acids. The mass spectra of the liberated acyl-oxyacyl residues proved the concomitant presence of 3-O-(cisΔ11–18 : 1)–18 : 0 and 3-O-(c19 : 0)-14 : 0 as the predominating diesters in this mixed lipid A. The glycosidically linked and the ester-linked phosphate groups of the backbone disaccharide were neither substituted by ethanolamine, phosphorylethanolamine, nor by 4-amino-4-deoxy-l-arabinose, in contrast to most of the enterobacterial lipid As. In the core oligosaccharide fraction, a HexA (1→4)HexA(1→5)Kdo-trisaccharide was identified by methylation analysis. The terminal HexA (hexuronic acid) is possibly 4-OMe-GalA, a component described here as an LPS constituent for the first time. LPS of R. salinarum showed a lethality in C57BL/10 ScSN (LPS-responder)-mice) of an order of 10–1–10–2 of that reported for Salmonella abortus equi LPS, and it was also capable of inducing TNFα and IL6 in macrophages of C57BL/10ScSN mice.
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  • 7
    ISSN: 1432-072X
    Keywords: Rhodopseudomonas palustris ; O-Antigens ; Serology ; Lipopolysaccharides
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Description / Table of Contents: Zusammenfassung Die isolierten O-Antigen (Lipopolysaccharide) aus 12 Rhodopseudomonas palustris-Stämmen konnten drei verschiedenen Serotypen zugeordnet werden. Diese decken sich mit der kürzlich erarbeiteten Chemotypisierung dieser O-Antigene. Im passiven Hämagglutinationstest, der mit den Lipopolysacchariden der Stämme 2/2, K/1 (Chemotyp I), sowie 8/1, 1e5 (Chemotyp II) und 11/1, 15, 1a1, 42 (Chemotyp III) und den entsprechenden Antiseren gegen diese Stämme ausgeführt wurde, zeigten die O-Antigene nur innerhalb eines Chemotyps Kreuzreaktionen. Zwischen Lipopolysacchariden verschiedener Chemotypen wurden in keinem Fall Kreuzreaktionen beobachtet. Entsprechende Ergebnisse wurden durch Immunelektrophorese und im Agargel-Präcipitationstest nach Ouchterlony erhalten, bei welchen die Lipopolysaccharide der Stämme 8/1 (Chemotyp II) und 11/1 (Chemotyp III) mit den homologen und den gegen die übrigen Stämme gerichteten heterologen Antiseren getestet wurden. Durch Kreuzabsorptionen der Antiseren wurde für die dem Chemotyp III zugehörenden Stämme 11/1, 15 und 1a1 O-Identität nachgewiesen. Die weitgehende Parallelität von Chemo- und Serotyp hat somit nicht nur für Enterobacteriaceae, sondern auch für die taxonomisch weit entfernten Rhodospirillaceae Gültigkeit. Jedoch sind die Serotypen bzw. Chemotypen der untersuchten Stämme nicht als selbständige Taxa anzusehen. Mit Lipopolysacchariden aus anderen Rhodospirillaceae-Species (Rhodopseudomonas capsulata, Rhodopseudomonas viridis, Rhodopseudomonas gelatinosa, Rhodospirillum rubrum) wurde in keinem Fall Kreuzreaktion erhalten.
    Notes: Abstract Serological investigations carried out with isolated O-antigens (lipopolysaccharides) of 12 strains of Rhodopseudomonas palustris showed that these O-antigens can be arranged into 3 distinct serotypes. These are identical with the three chemotypes previously established. The passive hemagglutination test carried out with isolated lipopolysaccharides of the strains 2/2 and K/1 of chemotype I, strains 8/1 and 1 e5 of chemotype II and strains 11/1, 15, 1a1 and 42 of chemotype III and their respective rabbit antisera showed that crossreactions can only be observed in between the chemotypes. No cross-reactions were observed between O-antigens of different chemotypes. This result was essentially confirmed by gel-precipitation studies (according to Ouchterlony) and in immunoelectrophoresis using the alkali-treated lipopolysaccharides of strains 8/1 (chemotype II) and 11/1 (chemotype III) as test antigens and antisera against strains of chemotypes I–III. Cross absorptions carried out with strains 11/1, 15 and 1a1, all belonging to chemotype III, showed identity of their O-antigens. The earlier established rather far-reaching parallelity of chemo- and serotypes existing in Enterobacteriaceae is therefore also valid for the taxonomically remote Rhodospirillaceae. In contrast to Enterobacteriaceae the different serotypes of Rhodopseudomonas palustris are not distinct taxa. Additional studies carried out with isolated lipopolysaccharides of other species of Rhodospirillaceae (Rhodopseudomonas capsulata, viridis and gelatinosa and Rhodospirillum rubrum) failed to show any cross reactivities with the three serotypes of Rhodopseudomonas palustris.
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  • 8
    ISSN: 1432-072X
    Keywords: Microcystis aeruginosa ; Chemotype ; Lipopolysaccharide ; Polysaccharide ; Lipid A ; Cell envelope ; Cell wall
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Lipopolysaccharides were obtained from three strains (PCC 7806, PCC 7820 and UV-017) of the waterbloom-forming cyanobacterium Microcystis aeruginosa. 3-Hydroxy fatty acids (3-OH-14:0, 3-OH-16:0, 3-OH-18:0) in addition to other fatty acids were identified in all three lipopolysaccharides. Glucosamine, the only amino sugar found, presumably represents the backbone amino sugar of the phosphate-free lipid A moiety. Heptoses were absent and 2-keto-3-deoxyoctonate was not detected in all three lipopolysaccharides. Strain-specificity was revealed in the complex composition of the polysaccharide moieties. Strains PCC 7806 and UV-017 were of the same chemotype, it differs from that of strain PCC 7820. Polysaccharides with strain-specific chemical compositions different from those of the respective lipopolysaccharides were obtained from each strain. The polysaccharides are likely to be ascribed to external cell envelope layers, their sugar specificity was in parallel with the O-chain chemotypes of the lipopolysaccharides of the three strains.
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  • 9
    ISSN: 1432-072X
    Keywords: Sphaerotilus natans ; Lipopolysaccharide ; Lipid A ; Laser desorption mass spectrometry ; DOC-PAGE ; 3-Hydroxycapric acid ; Proteobacteria
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The lipopolysaccharide of Sphaerotilus natans afforded a ladder-like pattern of bands in sodium deoxycholate-polyacrylamide gel electrophoresis, indicating the presence of a S-form lipopolysaccharide. The chemical analysis showed neutral sugars (rhamnose, glucose, l-glycero-d-manno-heptose), 3-deoxy-octulosonic acid (Kdo), amino compounds (glucosamine, glucosamine phosphate, ethanolamine and ethanolamine phosphate), and phosphorus. The lipid A fraction contained saturated and unsaturated capric, lauric, and myristic acids, and 3-hydroxy capric acid (3-OH-10:0). Its chemical structure was consisting of a glucosamine disaccharide, glycosidically substituted by a phosphomonoester, and substituted at C-4′ by a pyrophosphodiester esterified with ethanolamine. The amino groups of both glucosamines are acylated by 3-hydroxy capric acids and these in turn are substituted by saturated and unsaturated capric, lauric, and myristic acids. Hydroxyl groups of the backbone disaccharide at C-3 and C-3′ were also esterified by 3-hydroxy capric acid, those at C-4 and C-6 were unsubstituted. The latter provides the attachment site for Kdo.
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  • 10
    ISSN: 1432-072X
    Keywords: Carotenoids ; Cell wall ; Cyanobacterium ; Peptidoglycan-associated proteins ; Polypeptide pattern ; Synechocystis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cell walls free of cytoplasmic- and thylakoid membranes were isolated from Synechocystis PCC 6714 by sucrose density gradient centrifugation and extraction with Triton X-100. The Triton-insoluble cell wall fraction retained the multilayered fine structure. Peptidoglycan, proteins, polysaccharides, lipopolysaccharides, lipids and carotenoids were found as constituents of the cell wall. Polypeptide and lipid patterns of cell walls were completely different from that of the cytoplasmic/thylakoid membrane fraction. The purified cell walls contained about twelve outer membrane proteins. The two major polypeptides (Mr 67,000 and 61,000) were found to be associated with the peptidoglycan by ionic interactions. Myxoxanthophyll (major carotenoid), related carotenoid-glycosides and zeaxanthin were the predominating carotenoids of the cell wall of Synechocystis PCC 6714 over echinenone and β-carotene. A polar unknown carotenoid was observed, the absorption spectrum of which resembled that of myxoxanthophyll. It was exclusively found in cell walls, but not in the cytoplasmic/thylakoid membrane fraction.
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