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

Release of lipopolysaccharide from E. coli was studied in the presence of sub-minimal inhibitory concentrations of ethylenediaminetetraacetic acid (EDTA). In untreated cells no release was detected with 50 mM Mg2+ in the medium, but a steady release of over 50% of the synthesized lipopolysaccharide was observed with 0.1 mM Mg2+. EDTA at MIC/8 led to a 2- to 3-fold higher release, presumably by an adjustment of the concentration of unchelated Mg2+ to a value still sustaining normal growth but giving rise to a highly unstable outer membrane. No structural difference was observed between cell-bound and released lipopolysaccharide.
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PMID:Effects of sub-minimal inhibitory concentrations of EDTA on growth of Escherichia coli and the release of lipopolysaccharide. 818 24

L1 is a cell surface glycoprotein of the immunoglobulin superfamily which was initially shown to mediate adhesion between neural cells. Recently we have reported that L1 is expressed by bone marrow cells and the majority of mature lymphocytes (Kowitz et al., Eur. J. Immunol. 1992. 22: 1199-1205). To analyze the function of L1 on leukocytes we studied its regulation following cell activation. In vitro activation of B lymphocytes with lipopolysaccharide or T lymphocytes with phorbol 12-myristate 13-acetate/Ca2+ ionophore, concanavalin A or anti-CD3 monoclonal antibody as well as in vivo activation of V beta 8+ T cells with staphylococcal enterotoxin B (SEB) revealed a down-regulation of L1 within 48 h. A rapid loss of L1 expression was seen when mouse neutrophils were activated with PMA alone. This rapid loss paralleled the shedding of L-selectin. We also studied a possible role of L1 in the binding of leukocytes to endothelial cells. ESb-MP lymphoma cells with a high expression of L1 (L1hi) could bind to bend3 endothelioma cells without prior activation with inflammatory cytokines. The interaction was inhibited by anti-L1 antibodies. In contrast, ESb-MP cells with low L1 expression (L1lo) were only marginally bound. Latex beads coated with affinity-isolated L1 antigen were also able to bind to the endothelioma cells in a specific fashion. The binding of ESb-MP lymphoma cells required Ca2+ and Mg2+ ions and was sensitive to cold temperature. Since the endothelioma cells did not express L1 the binding mechanism studied here is distinct from the established L1-L1 homotypic interaction. It is possible that the novel L1-mediated adhesion pathway involves an unidentified ligand and could play a role in leukocyte migration.
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PMID:L1 adhesion molecule on mouse leukocytes: regulation and involvement in endothelial cell binding. 822 69

Although the antimicrobial activity of lactoferrin has been well described, its mechanism of action has been poorly characterized. Recent work has indicated that in addition to binding iron, human lactoferrin damages the outer membrane of gram-negative bacteria. In this study, we determined whether bovine lactoferrin and a pepsin-derived bovine lactoferrin peptide (lactoferricin) fragment have similar activities. We found that both 20 microM bovine lactoferrin and 20 microM lactoferricin release intrinsically labeled [3H]lipopolysaccharide ([3H]LPS) from three bacterial strains, Escherichia coli CL99 1-2, Salmonella typhimurium SL696, and Salmonella montevideo SL5222. Under most conditions, more LPS is released by the peptide fragment than by whole bovine lactoferrin. In the presence of either lactoferrin or lactoferricin there is increased killing of E. coli CL99 1-2 by lysozyme. Like human lactoferrin, bovine lactoferrin and lactoferricin have the ability to bind to free intrinsically labeled [3H]LPS molecules. In addition to these effects, whereas bovine lactoferrin was at most bacteriostatic, lactoferricin demonstrated consistent bactericidal activity against gram-negative bacteria. This bactericidal effect is modulated by the cations Ca2+, Mg2+, and Fe3+ but is independent of the osmolarity of the medium. Transmission electron microscopy of bacterial cells exposed to lactoferricin show the immediate development of electron-dense "membrane blisters." These experiments offer evidence that bovine lactoferrin and lactoferricin damage the outer membrane of gram-negative bacteria. Moreover, the peptide fragment lactoferricin has direct bactericidal activity. As lactoferrin is exposed to proteolytic factors in vivo which could cleave the lactoferricin fragment, the effects of this peptide are of both mechanistic and physiologic relevance.
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PMID:Antibacterial activity of lactoferrin and a pepsin-derived lactoferrin peptide fragment. 842 97

This study describes activation of serum complement by lipoteichoic acid (LTA) from Streptococcus mutans OMZ 176, while in solution. Serum from 16 healthy students was taken. Test samples were incubated with increasing doses (1-5,000 micrograms/ml) of LTA or lipopolysaccharide (LPS) from Escherichia coli 0111:B4 for 1 h at 37 degrees C; then assayed for degradation of C3, C4 or factor B by crossed immunoelectrophoresis. Each preparation caused a significant (p < 0.05) dose-dependent conversion of C3. The response curves obtained were not statistically different. LPS was a stronger activator of the alternative pathway than LTA, as judged from analysis of C3 degradation in the presence of Mg2+/EGTA, and from their effects on factor B cleavage. LTA caused, however, pronounced alterations in the shape of C4 precipitation in the gels. Functional (hemolytic) assays showed that, when tested at 200 micrograms/ml, LTA and LPS triggered significant (p < 0.05) consumptions of both classical and alternative pathway proteins. LPS was a significantly (p < 0.05) stronger activator than LTA. Apparently, the C3 degradation found for this LTA involved the alternative pathway to a small extent; thus some other mechanism of fluid-phase C3 cleavage seemed also to be operative.
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PMID:Effects of a streptococcal lipoteichoic acid on complement activation in vitro. 845 83

Teicoplanin is a glycopeptide antibiotic which is ineffective against gram-negative bacteria because of its inability to penetrate the outer membrane. Removal of the sugar residues and attachment of polyamines to carbon 63 yielded two dibasic deglucoteicoplanin amides, MDL 62,766 (766) and MDL 62,934 (934), with moderate MICs for Escherichia coli (2 to 4 micrograms/ml) and Pseudomonas aeruginosa (8 to 32 micrograms/ml) compared with those of the monobasic teicoplanin aglycone (16 and > 1,024 micrograms/ml, respectively). MICs were increased 16- to 32-fold by Mg2+ supplementation of Luria broth but not by Na+ supplementation at an equivalent ionic strength. Both 766 and 934 were capable of binding to P. aeruginosa lipopolysaccharide (LPS) at Mg(2+)-binding sites, as assessed by dansyl polymyxin displacement experiments. Furthermore, both compounds increased E. coli and P. aeruginosa outer membrane permeability to the hydrophobic fluorescent probe 1-N-phenylnaphthylamine (NPN), whereas the parent compounds teicoplanin aglycone and teicoplanin and the beta-lactam ceftazidime were totally ineffective. Addition of 1 mM Mg2+ blocked the increase in outer membrane permeability. Compound 766 had a lower MIC than 934 for both bacteria tested, bound to LPS with a higher affinity, and permeabilized outer membranes to NPN at lower concentrations. We propose that both deglucoteicoplanin amides exhibit increased gram-negative activity by virtue of their ability to access the self-promoted uptake pathway across the outer membrane.
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PMID:Mechanism of uptake of deglucoteicoplanin amide derivatives across outer membranes of Escherichia coli and Pseudomonas aeruginosa. 846 Sep 14

The structural polymorphism of free lipid A and deep rough mutant lipopolysaccharide (LPS Re) from Salmonella minnesota strain R595 and Escherichia coli strain F515 was characterized by Fourier transform infrared (IR) spectroscopy. For this, the beta <--> alpha phase states and the three-dimensional supramolecular structures, the latter deduced from small-angle synchrotron radiation x-ray diffraction, were investigated at different water contents, Mg2+ concentrations, and temperatures. The analysis of the IR data for vibrations originating from the hydrophobic moiety shows that the beta <--> alpha acyl chain melting is strongly expressed only for the stretching and scissoring modes of the methylene groups. Vibrational groups originating from the interface region sense the acyl chain melting well (ester carbonyl bands) or only weakly (amide bands), and those resulting from the pure polar moiety not at all. From the x-ray data, the existence of lamellar (L), different cubic, and, for lipid A and LPS R595, also inverted hexagonal (HII) structures could be proven in the temperature range 20-80 degrees C with cubic <--> cubic and cubic <--> HII transitions for the Mg(2+)-free and L <--> HII transitions for the Mg(2+)-containing samples. These structural transitions can be characterized most readily by specific changes of the vibrational bands resulting from the interface region: the ester carbonyl and the amide bands. The magnitude of the changes corresponds to that of the structural rearrangement, i.e., is highest for the L <--> HII, lower for the cubic <--> HII, and lowest for the cubic <--> cubic transitions. The structural transitions are only marginally expressed for vibrational bands of the hydrophobic moiety. Similarly, the band contours of vibrations from the hydrophilic region are no indicators of the structural reorientations except for the carboxylate bands of LPS Re. Particularly the stretching vibrations of the phosphate groups are nearly completely invariant; the absolute values of their half bandwidths, however, differ significantly for lipid A and LPS Re, which seems to be of biological relevance. The ability of IR spectroscopy to detect supramolecular changes also beyond the measurability by x-ray diffraction, i.e., at water contents > 95 to 99.5%, is demonstrated.
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PMID:Fourier transform infrared spectroscopy characterization of the lamellar and nonlamellar structures of free lipid A and Re lipopolysaccharides from Salmonella minnesota and Escherichia coli. 849 79

1. Inositol hexakisphosphate (InsP6) is a ubiquitous and abundant cytosolic inositol phosphate that has been reported to prime human neutrophils for enhanced agonist-stimulated superoxide anion generation. This led to the proposal that the release of InsP6 from necrotic cells may augment the functional responsiveness of neutrophils at an inflammatory focus. The aim of this study was to examine whether the functional effects of InsP6 in neutrophils are receptor-mediated and establish the magnitude of this priming effect relative to other better characterized priming agents. 2. Analysis of [3H]-InsP6 binding to human neutrophil membranes in 20 mM Tris, 20 mM NaCl, 100 mM KCl, 5 mM EDTA (pH 7.7) buffer using 0.1 mg ml-1 membrane protein and 2.5 nM [3H]-InsP6 (90 min, 4 degrees C), demonstrated specific low affinity [3H]-InsP6 binding that was non-saturable up to a radioligand concentration of 10 nM. 3. [3H]-InsP6 displacement by InsP6 gave a Hill coefficient of 0.55 and best fitted a two-site logistic model (53% KD 150 nM, 47% KD 5 microM). [3H]-InsP6 binding also displayed low (3 fold) selectivity for InsP6 over Ins(1,3,4,5,6)P5. 4. The specific [3H]-InsP6 binding displayed a pH optimum of 8, was abolished by pre-boiling the membranes, and was enhanced by Ca2+, Mg2+ and Na+. 5. In incubations with intact neutrophils, where high levels of specific [3H]-LTB4 binding was observed, no [3H]-InsP6 binding could be identified. 6. Preincubation of neutrophils with 100 microM InsP6 had no effect on resting cell morphology, but caused a minor and transient (maximal at 30 s) enhancement of (0.1 nM) fMLP-induced shape change (% cells shape changed: fMLP 53 +/- 3%, fMLP+InsP6 66 +/- 4%). Similarly, InsP6 (100 microM, 30 s) had no effect on basal superoxide anion generation and, compared to lipopolysaccharide (LPS, 100 ng ml-1, 60 min), tumour necrosis factor-alpha (TNF alpha, 200 u ml-1, 30 min) or platelet-activating factor (PAF, 100 nM, 5 min) caused only a small enhancement of 100 nM fMLP-stimulated superoxide anion generation (fold-increase in superoxide anion generation over fMLP alone: InsP6 1.8 +/- 0.3, LPS 6.8 +/- 0.6, TNF alpha 5.2 +/- 0.7, PAF 5.8 +/- 0.6). 7. While these data support the presence of a specific, albeit low affinity, [3H]-InsP6 binding site in human neutrophil membrane preparations, the lack of binding to intact cells implies that the functional effects of InsP6 (ie. enhanced fMLP-stimulated superoxide anion generation and shape change) are not receptor-mediated.
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PMID:Characterization of inositol hexakisphosphate (InsP6)-mediated priming in human neutrophils: lack of extracellular [3H]-InsP6 receptors. 885 21

An extracellular surface-active agent, PM-factor, was obtained by high-speed centrifugation from the culture broth of Pseudomonas marginalis PD-14B. PM-factor exhibited emulsifying activity on a broad spectrum of hydrocarbon liquids, including aromatics, aliphatics, crude oil, and creosote. The factor appeared as ball-shaped particles of varying diameter when examined by electron microscopy (0.16-1.4 microns). Gel filtration chromatography demonstrated a high molecular mass of the factor (> 10(6) Da). The ultraviolet absorption spectrum manifested a peak in the region 200 nm rather in the region 260-280 nm. Amino acid analysis showed a very low amount of aromatic amino acids residues in the protein moiety of PM-factor. The presence of 3-deoxy-D-mannooctulosonic acid, heptose, hexosamine, phosphorus, and 3-hydroxy fatty acid indicated that PM-factor contained lipopolysaccharide. The emulsifying activity of PM-factor was inhibited strongly by mercuric chloride and moderately by EDTA. Polymyxin B, Ca2+, and Mg2+ markedly stimulated the factors emulsifying activity. Roles of the bioemulsifier in the functioning of P. marginalis as a plant pathogen and in bioremediation are discussed.
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PMID:Physicochemical properties of PM-factor, a surface-active agent produced by Pseudomonas marginalis. 886 31

Bacterial lipopolysaccharide (LPS)-induced exocytosis is one of the primary immune responses of the Limulus granulocyte (GR). Exocytosis can be mediated by guanine nucleotide-binding protein (G-protein)-linked surface receptors that activate phospholipase C (PLC) to produce inositol 1,4,5-triphosphate (IP3) and diacylglycerol (DAG). IP3 mobilizes intracellular Ca2+ ([Ca2+]i), which can lead to exocytosis. We used activators and inhibitors of known signal transduction pathways to investigate the signaling pathway responsible for LPS-induced exocytosis in the GR. These compounds have been shown to similarly effect pathways in vertebrate and invertebrate systems and this assumption is made here. Pretreatment of GRs with cholera and pertussis toxins, which modulate G-proteins, and U73122, which inhibits PLC, inhibited LPS-induced exocytosis, but pretreatment with the tyrosine kinase inhibitor herbimycin did not. In contrast, exocytosis was induced with fluoride (a G-protein activator) and thapsigargin with Mg2+ (an inhibitor of endomembranous Ca(2+)-ATPase). Exocytosis was not induced by phorbol ester, which mimics DAG to activate protein kinase C (PKC) and it was not effected by ethanol or chelerythrine, which inhibit phospholipase D and PKC, respectively. Microinjection of GRs with different concentrations of IP3, an IP3 analog (DL-2,3,6,trideoxy-myo-inositol 1,4,5-triphosphate), Mg2+, or Ca2+ induced different percentages of exocytosis in individual cells, while HEPES buffer did not. Microfluorometric analysis of intracellular Mg2+ ([Mg2+]i) and [Ca2+]i, using the dyes Mag Fura-2AM and Calcium Green 5N, respectively, revealed [Mg2+]i and [Ca2+]i fluxes during LPS-induced exocytosis. This study suggests that LPS induces exocytosis in the Limulus GR through activation of G-protein-coupled receptors, which stimulate the IP3 signaling pathway to induce both [Ca2+]i and [Mg2+]i fluxes to facilitate vesicular and plasma membrane fusion. This is the first demonstration of the signal transduction pathway responsible for the primary immune response of the GR.
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PMID:Signal transduction during exocytosis in Limulus polyphemus granulocytes. 901 85

1. Nucleotide-induced currents in untreated (proliferating) and lipopolysaccharide (LPS; 100 ng ml(-1)) treated (non-proliferating) rat microglial cells were recorded by the whole-cell patch-clamp technique. Most experiments were carried out on non-proliferating microglial cells. ATP (100 nM-1 mM), ADP (10 nM-10 mM) and UTP (1 microM-100 mM), but not uridine (100 microM-10 mM) produced a slow outward current at a holding potential of 0 mV. The effect of UTP (1 mM) did not depend on the presence of extracellular Mg2+ (1 mM). The outward current response to UTP (1 mM) was similar in non-proliferating and proliferating microglia. 2. In non-proliferating microglial cells, the ATP (10 microM)-induced outward current was antagonized by suramin (300 microM) or reactive blue 2 (50 microM), whereas 8-(p-sulphophenyl)-theophylline (8-SPT; 100 microM) was inactive. By contrast, the current induced by UTP (1 mM) was increased by suramin (300 microM) and was not altered by reactive blue 2 (50 microM) or 8-SPT (100 microM). 3. The current response to UTP (1 mM) disappeared when K+ was replaced in the pipette solution by an equimolar concentration of Cs+ (150 mM). However, the effect of UTP (1 mM) did not change when most Cl- was replaced with an equimolar concentration of gluconate (145 mM). The application of 4-aminopyridine (1 mM) or Cs+ (1 mM) to the bath solution failed to alter the UTP (1 mM)-induced current. UTP (1 mM) had almost no effect in a nominally Ca2+-free bath medium, or in the presence of charybdotoxin (0.1 microM); the inclusion of U-73122 (5 microM) or heparin (5 mg ml(-1)) into the pipette solution also blocked the responses to UTP (1 mM). By contrast, the effect of ATP (10 microM) persisted under these conditions. 4. I-V relations were determined by delivering fast voltage ramps before and during the application of UTP (1 mM). In the presence of extracellular Cs+ (1 mM) and 4-aminopyridine (1 mM) the UTP-evoked current crossed the zero current level near -75 mV. Omission of Ca2+ from the Cs+ (1 mM)- and 4-aminopyridine (1 mM)-containing bath medium or replacement of K+ by Cs+ (150 mM) in the pipette solution abolished the UTP current. 5. Replacement of GTP (200 microM) by GDP-beta-S (200 microM) in the pipette solution abolished the current evoked by UTP (1 mM). 6. When the pipette solution contained Cs+ (150 mM) instead of K+ and in addition inositol 1,4,5,-trisphosphate (InsP3; 10 microM), an inward current absolutely dependent on extracellular Ca2+ was activated after the establishment of whole-cell recording conditions. This current had a typical delay, a rather slow time course and did not reverse its amplitude up to 100 mV, as measured by fast voltage ramps. 7. A rise of the internal free Ca2+ concentration from 0.01 to 0.5 microM on excised inside-out membrane patches produced single channel activity with a reversal potential of 0 mV in a symmetrical K+ solution. The reversal potential was shifted to negative values, when the extracellular K+ concentration was decreased from 144 to 32 mM. By contrast, a decrease of the extracellular Cl- concentration from 164 to 38 mM did not change the reversal potential. 8. Purine and pyrimidine nucleotides act at separate receptors in rat microglial cells. Pyrimidinoceptors activate via a G protein the enzyme phospholipase C with the subsequent release of InsP3. The depletion of the intracellular Ca2+ pool appears to initiate a capacitative entry of Ca+ from the extracellular space. This Ca2+ then activates a Ca2+-dependent K+ current.
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PMID:Coexistence of purino- and pyrimidinoceptors on activated rat microglial cells. 924 43


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