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

Outer membrane fractions (OMs) of nine Campylobacter (C.) jejuni and two C. coli strains belonging to different serovars, from human and various animal origins, were extracted by treatment with sodium N-lauryl sarcosinate. Using n-octyl-beta-D-glucopyranoside a 42-kDa protein and a flagella-enriched fraction were obtained. The capacity of the crude bacterial OM preparations, the purified 42-kDa protein and the flagella to bind to membranes of the human embryonic intestinal cell line INT 407 was tested by an enzyme-linked immunosorbent assay. The crude OM and the 42-kDa-enriched fraction were found to bind very well to the cell membranes, whereas the flagella preparation showed only a weak binding. Using monoclonal antibodies (mAbs) with HS 2-lipopolysaccharide (LPS) specificity, binding of crude HS 2 strain OM preparations to cell membranes was detected in a significant range, whereas with flagellin-specific mAbs binding of OMs and flagella to cell membranes was only detected to a very low extent. Binding of OMs to cell membranes was inhibited by preincubation of OMs with serovar-specific mouse hyperimmune serum, whereas on preincubation with mAbs directed against LPS or flagella binding was practically not inhibited. OMs extracted after pretreatment of the bacteria with proteinase K showed an altered SDS-PAGE pattern especially for the 42-kDa protein subunit and and their capacity to bind to cell membranes was significantly reduced. The binding was also reduced by preincubation of the OMs with L-fucose or D-mannose.
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PMID:In vitro binding of Campylobacter jejuni/coli outer membrane preparations to INT 407 cell membranes. 154 70

Artificial Salmonella serogroup B, D or Cl-specific glycolipids were prepared by covalently linking oligosaccharides corresponding to two O-antigen repeating units, obtained by phage enzyme hydrolysis of native O-antigenic polysaccharides, to octyl residues. Sheep erythrocytes coated with the artificial glycolipids were studied for their ability to consume C3, when incubated in C4- deficient guinea pig serum. Salmonella C1 (0-6,7) glycolipid-coated erythrocytes consumed C3 40% more efficiently than Salmonella D (0-9,12) glycolipid-coated erythrocytes, and 10-times more efficiently than Salmonella B (0-4,12) glycolipid-coated erythrocytes. These results resemble C3 consumption by Salmonella C1, D, and B cells and by sheep erythrocytes coated with purified lipopolysaccharides of these O-specificities. The results prove directly that in a particulate system C3 activation via the alternative pathway depends on the structural properties of the O-antigenic side chain. Structures as small as octasaccharides, or as two O-antigenic repeating units, are sufficient for triggering C3 activation, but the magnitude of activation depends on the nature of the monosaccharides. Apparently, neither the core oligosaccharide nor Lipid A of lipopolysaccharide are required for C3 activation via the alternative pathway.
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PMID:Salmonella O antigen-specific oligosaccharide-octyl conjugates activate complement via the alternative pathway at different rates depending on the structure of the O antigen. 169 20

Bacterial lipid macroamphiphiles extracted with phenol/water can be purified in one step by hydrophobic interaction chromatography. Lipids and the major part of protein are separated from macroamphiphiles during phenol/water extraction. Coextracted nucleic acids, polysaccharides, and residual protein are effectively removed by column chromatography on octyl-Sepharose whereby macroamphiphiles are primarily adsorbed and later eluted with a buffered propanol gradient. The procedure is applicable to macroamphiphiles with various lipid structures as was demonstrated using the diacylglycerol-containing lipoglycan of Micrococcus luteus, the lipid A-containing lipopolysaccharide of Salmonella typhimurium, and the diglyceryl tetraether lipoglycans of Thermoplasma acidophilum and Thermoplasma volcanicum. On elution from octyl-Sepharose, separation into molecular species of different compositions was observed with the lipopolysaccharide of S. typhimurium and the lipoglycan of T. volcanicum. It was also shown that, after phenol/water extraction, membrane lipids are completely recoverable from the phenol layer, which makes it possible to isolate lipids along with macroamphiphiles from the same sample of bacteria.
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PMID:One-step purification of bacterial lipid macroamphiphiles by hydrophobic interaction chromatography. 186 38

By hydrophobic interaction chromatography on octyl-Sepharose, lipopolysaccharide (LPS) of Escherichia coli Re mutant and of wild-type smooth-form (S-form) Salmonella typhimurium and Salmonella abortus equi is fractionated according to increasing amount of fatty acids. Thereby a fractionation of S-form LPS according to the length of the O-polysaccharide chain also occurs, because with increasing of fatty acids there is a decrease in the mean length of the O-polysaccharide chain from approximately 30 to 4 repeating units. Molecular species of Re-mutant LPS contain four 3-hydroxytetradecanoyl residues in addition to which dodecanoic, tetradecanoic and possibly hexadecanoic acid, appear in this sequence. Among the molecular species of S-form LPS, dodecanoic, tetradecanoic and hexadecanoic acids appear in the same order, but in contrast to Re-mutant LPS a significant fraction of S-form LPS contains less than four 3-hydroxytetradecanoyl residues. Hydrophobic interaction chromatography also proved an effective one-step purification procedure of LPS as was shown with a crude preparation from S-form S. typhimurium.
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PMID:Purification and fractionation of lipopolysaccharide from gram-negative bacteria by hydrophobic interaction chromatography. 226 90

A large-scale purification scheme was developed for lipopolysaccharide-free protein P, the phosphate-starvation-inducible outer-membrane porin from Pseudomonas aeruginosa. This highly purified protein P was used to successfully form hexagonal crystals in the presence of n-octyl-beta-glucopyranoside. Amino-acid analysis indicated that protein P had a similar composition to other bacterial outer membrane proteins, containing a high percentage (50%) of hydrophilic residues. The amino-terminal sequence of this protein, although not homologous to either outer membrane protein, PhoE or OmpF, of Escherichia coli, was found to have an analogous protein-folding pattern. Protein P in the native trimer form was capable of maintaining a stable functional trimer after proteinase cleavage. This suggested the existence of a strongly associated tertiary and quaternary structure. Circular dichroism studies confirmed these results in that a large proportion of the protein structure was determined to be beta-sheet and resistant to acid pH and heating in 0.1% sodium dodecyl sulphate.
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PMID:Large-scale purification and biochemical characterization of crystallization-grade porin protein P from Pseudomonas aeruginosa. 245 38

Thermal and low pH stabilities of matrix porin (Omp F) solubilized in the micellar solutions of ionic (SDS) and nonionic detergents were investigated by the methods of circular dichroism, intrinsic fluorescence, light scattering and sedimentation velocity. The stability of porin structure in solution is much higher in the presence of beta-octyl glucoside than with SDS. In the presence of SDS, sharp transitions were detected by all parameters measured, above 55 degrees C at neutral pH and below pH 4.5 at 20 degrees C. These transitions involve at least three concomitant processes: unfolding of protein, dissociation of trimers to monomers and the disruption of the protein-detergent micellar complexes, all events being irreversible in the presence of SDS. The nonionic detergent, beta-octyl glucoside, increases the stability of porin in acidic conditions, since neither dissociation nor denaturation was observed in the pH region between 7.5 and 2.0. However, at pH less than 3.5, small, reversible changes in protein structure became evident. The thermal stability of porin is also increased by beta-octyl glucoside as evidenced by a transition temperature 15-20 degrees C higher as compared to SDS. A considerable degree of native porin structure was regained after heat treatment in the presence of beta-octyl glucoside, though the reconstituted trimers were not identical to the native ones. The addition of lipopolysaccharide and divalent cations (Ca2+, Mg2+) to the experimental system did not improve the thermal reversibility.
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PMID:Effect of temperature and low pH on structure and stability of matrix porin in micellar detergent solutions. 300 79

A method of reducing endotoxin contamination in protein-containing solutions is described here using a combination of polymyxin B-Sepharose 4B (PB-Seph 4B) affinity binding and endotoxin-protein dissociation with the dialyzable surfactant, octyl-beta-D-glucopyranoside (OBDG). Using the limulus amoebocyte lysate (LAL) assay to detect endotoxin, greater than 1000-fold reduction of endotoxin reactivity could be accomplished from a contaminated commercial preparation of bovine catalase. Importantly, this occurred with only a 24% protein loss and an 11% loss of catalase enzymatic activity after treatment. The treated catalase appeared to be largely endotoxin-free since it no longer elicited a pyrogenic response in rabbits or primed for intravascular coagulation of the generalized Shwartzman reaction. Of interest, OBDG treatment of Salmonella minnesota Re595 lipopolysaccharide enhanced its ability to bind to serum high density lipoproteins which might contribute to decreased in vivo toxicity. In quantitative studies using radiolabeled endotoxin, the OBDG was shown to be capable of dissociating protein-bound endotoxin thereby facilitating its binding to the PB-Seph 4B adduct. The technique was also useful in removing radiolabeled endotoxin added to human IgG. The methodology described here would be expected to have general usefulness in reducing endotoxin contamination of macromolecular solutions that can bind and retain endotoxin.
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PMID:A new method for reduction of endotoxin contamination from protein solutions. 369 9

During intraperiplasmic growth of Bdellovibrio bacteriovorus 109J, the substrate cell surface becomes more hydrophobic. This was shown (i) by comparing the sensitivity to hydrophobic antibiotics of wild-type and lipopolysaccharide mutant strains of Salmonella typhimurium to that of the bdellovibrio growing on these strains and (ii) by measuring the binding efficiency of these strains, Escherichia coli, and their derived bdelloplasts to octyl Sepharose. The kinetics of increase in surface hydrophobicity was similar to the kinetics of the conversion of the substrate cell peptidoglycan to a lysozyme-resistant form (M. Thomashow and S. Rittenberg, J. Bacteriol. 135:1008-1014, 1978), and hydrophobicity reached a maximum at about 60 min in a synchronous culture. The change in hydrophobicity was inhibited by chloramphenicol, suggesting that bdellovibrio protein synthesis was required. Control experiments revealed that the free-swimming bdellovibrio had a more hydrophobic surface than the deep rough mutants of S. typhimurium.
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PMID:Change in the surface hydrophobicity of substrate cells during bdelloplast formation by Bdellovibrio bacteriovorus 109J. 636 84

T6-phage inactivation by homogeneous, lipopolysaccharide-free Tsx protein showed a clear dependence on added lipopolysaccharide, when assayed in the presence of a non-ionic detergent, octyl-polyoxyethylene (octyl-POE). Under these conditions, the protein exists in a monomeric state.
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PMID:Lipopolysaccharide requirement of phage T6 inactivation in Escherichia coli K12. 704 10

During the course of gene therapy experiments in rodents, using intramuscular injections of plasmid DNA derived from Escherichia coli, we noted dose-related toxicity. This observation prompted a search for possible contaminants of DNA samples. We used the highly specific and sensitive limulus amoebocyte lysate assay (LAL), to monitor endotoxin bioactivity in DNA samples, and found plasmid DNA derived from standard E. coli bacterial strains, using traditional DNA isolation protocols, to be heavily contaminated with endotoxin, or lipopolysaccharide (LPA). Standard DNA isolation procedures resulted in the copurification of up to 500 micrograms/ml of LPS. LPS is a potent inducer of cytokines and other inflammatory mediators, and may complicate the use of naked DNA in gene therapy. The copurification of endotoxin with plasmid DNA also has important implications for in vitro transfection studies and microinjection of DNA into embryos. A simple and efficient protocol to reduce LPS contamination of plasmid DNA was developed. The conversion of intact bacteria to spheroplasts prior to the isolation of plasmid DNA, incubation with lysozyme, treatment with the detergent n-octyl-beta-D-thioglucopyranoside (OSPG) and polymyxin-B (PMB) chromatography, allowed the isolation of plasmid DNA containing less than 50 ng/ml LPS. This represents a 10,000-fold reduction in LPS contamination, compared to conventional methods of plasmid DNA purification, avoids potentially toxic reagents such as ethidium bromide, and produces a higher yield of plasmid DNA.
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PMID:Bacterial lipopolysaccharide copurifies with plasmid DNA: implications for animal models and human gene therapy. 777 15


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