UNIPROT:P43026 - FACTA Search

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

The mortality rate for septic patients with acute renal failure is extremely high. Since sepsis is often caused by lipopolysaccharide (LPS), a model of LPS challenge was used to study the development of kidney injury. Intravital video microscopy was utilized to investigate renal peritubular capillary blood flow in anesthetized male C57BL/6 mice at 0, 2, 6, 10, 18, 24, 36, and 48 h after LPS administration (10 mg/kg ip). As early as 2 h, capillary perfusion was dramatically compromised. Vessels with continuous flow were decreased from 89 +/- 4% in saline controls to 57 +/- 5% in LPS-treated mice (P < 0.01), and vessels with intermittent flow were increased from 6 +/- 2% to 31 +/- 5% (P < 0.01). At 2 h, mRNA for intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 were elevated 50- and 27-fold, respectively, suggesting that vascular inflammation is an early event that may contribute to capillary dysfunction. By 10 h, vessels with no flow increased from 5 +/- 2% in saline controls to 19 +/- 3% in LPS-treated mice (P < 0.05). By 48 h, capillary function was returning toward control levels. The decline in functional capillaries preceded the development of renal failure and was paralleled by induction of inducible nitric oxide synthase in the kidney. Using NAD(P)H autofluorescence as an indicator of cellular redox stress, we found that tubular cell stress was highly correlated with the percentage of dysfunctional capillaries (r(2) = 0.8951, P < 0.0001). These data show that peritubular capillary dysfunction is an early event that contributes to tubular stress and renal injury.
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PMID:Peritubular capillary dysfunction and renal tubular epithelial cell stress following lipopolysaccharide administration in mice. 1692 42

The mortality rate for septic patients with acute renal failure is approximately doubled compared with patients with sepsis alone. Unfortunately, the treatment for sepsis-induced renal failure has advanced little during the last several decades. Because sepsis is often caused by lipopolysaccharide (LPS), a mouse model of LPS challenge was used to study the development of kidney injury. We hypothesized that inducible nitric-oxide synthase (iNOS)-catalyzed nitric oxide production and that generation of reactive nitrogen species (RNS) might play a role in the microcirculatory defect and resulting tubular injury associated with LPS administration. Fluorescent intravital videomicroscopy was used to assess renal peritubular capillary perfusion and document RNS generation by renal tubules in real time. As early as 6 h after LPS administration (10 mg/kg i.p.), RNS generation (rhodamine fluorescence), redox stress [NAD(P)H autofluorescence], and the percentage of capillaries without flow were each significantly increased compared with saline-treated mice (p < 0.05). The generation of RNS was supported by the detection of nitrotyrosine-protein adducts in the kidney using immunohistochemistry. The iNOS inhibitor l-N(6)-(1-iminoethyl)-lysine (l-NIL; 3 mg/kg i.p.) completely blocked the increase in rhodamine fluorescence and NAD(P)H autofluorescence and prevented the capillary defects at 6 h after LPS administration. These results suggest that iNOS-derived RNS is an important contributor to the peritubular capillary perfusion defects and RNS generation that occur during sepsis and emphasize that pharmacological inhibition of iNOS may provide beneficial effects during sepsis by improving renal capillary perfusion and reducing RNS generation in the kidney.
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PMID:Effects of the inducible nitric-oxide synthase inhibitor L-N(6)-(1-iminoethyl)-lysine on microcirculation and reactive nitrogen species generation in the kidney following lipopolysaccharide administration in mice. 1720 3

Bacteria of genus Aeromonas, responsible for a variety of pathological conditions in humans and fish, are ubiquitous waterborne bacteria. Aeromonas produces several virulent factors including a complex of lipopolysaccharide and surface array protein, involved in colonization. UDP-galactose 4-epimerase (GalE) catalyzes the production of UDP-galactose, a precursor for lipopolysaccharide biosynthesis, and thus is an important drug target. GalE exhibits interspecies variation and heterogeneity at its structural and functional level and therefore, the differences between the GalE of the host and the pathogen can be exploited for drug designing. In the present study, we report biochemical and functional characterization of the recombinant GalE of Aeromonas hydrophila. Unlike GalE reported from all other species, the purified recombinant GalE of A. hydrophila was found to exist as a monomer. This is the first report of UDP-galactose 4-epimerase from any species being a monomer. The molecular mass of the 6xHis-rGalE was determined to be 38271.477 (m/z). The 6xHis-rGalE with a K(m) of 0.5 mM for UDP-galactose exhibited optimum activity at 37 degrees C and pH 8-9. Spectrofluorimetric and CD analysis confirmed that the thermal inactivation was due to structural changes and not due to the NAD-dissociation. A relatively more ordered structure of the enzyme at pH 8 and 9 as compared to that at pH 6 or 7 suggests a key role of the electrostatic interactions in maintaining its native tertiary structure.
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PMID:Biochemical and functional characterization of UDP-galactose 4-epimerase from Aeromonas hydrophila. 1755 60

The nuclear enzyme poly(ADP-ribose) polymerase-1 (PARP-1) which was initially known for its role in the repair of oxidative stress-induced DNA damage, has also been reported to play a mediating role in the inflammatory response. Studies with PARP-1 knockout models have shown that PARP-1 is a co-activator of Nuclear Factor-kappa B (NF-kappaB), although this appears not to require its enzyme activity. In addition, drug-induced inhibition of the enzyme activity of PARP-1 was observed to reduce the production of pro-inflammatory mediators. In this study, the flavonoid compound flavone was demonstrated to significantly inhibit the enzyme activity of PARP-1. Further evaluation of flavone in N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-treated human pulmonary epithelial and vascular endothelial cells revealed that both the decrease in NAD(+) levels, as well as the formation of PAR-polymers was dose-dependently attenuated by flavone. In addition, flavone was found to reduce the lipopolysaccharide (LPS)-induced interleukin (IL)-8 production in pulmonary epithelial cells, which was confirmed by transcription analysis. Furthermore, the transcription Inhibitor kappa B alpha (of IkappaBalpha) was significantly increased by flavone. The results of the present study indicate that the flavonoid flavone could be a potential candidate for application in treatment of chronic inflammatory diseases. PARP-1 inhibition could have beneficial effects in such diseases as Chronic Obstructive Pulmonary Disease (COPD) and diabetes, by preservation of cellular NAD(+) levels and attenuating inflammatory conditions.
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PMID:Flavone as PARP-1 inhibitor: its effect on lipopolysaccharide induced gene-expression. 1764 14

The pathogenic bacteria Bordetella parapertussis and Bordetella bronchiseptica express a lipopolysaccharide O antigen containing a polymer of 2,3-diacetamido-2,3-dideoxy-l-galacturonic acid. The O-antigen cluster contains three neighbouring genes that encode proteins belonging to the short-chain dehydrogenase/reductase (SDR) family, wbmF, wbmG and wbmH, and we aimed to elucidate their individual functions. Mutation and complementation implicate each gene in O-antigen expression but, as their putative sugar nucleotide substrates are not currently available, biochemical characterisation of WbmF, WbmG and WbmH is impractical at the present time. SDR family members catalyse a wide range of chemical reactions including oxidation, reduction and epimerisation. Because they typically share low sequence conservation, however, catalytic function cannot be predicted from sequence analysis alone. In this context, structural characterisation of the native proteins, co-crystals and small-molecule soaks enables differentiation of the functions of WbmF, WbmG and WbmH. These proteins exhibit typical SDR architecture and coordinate NAD. In the substrate-binding domain, all three enzymes bind uridyl nucleotides. WbmG contains a typical SDR catalytic TYK triad, which is required for oxidoreductase function, but the active site is devoid of additional acid-base functionality. Similarly, WbmH possesses a TYK triad, but an otherwise feature-poor active site. Consequently, 3,5-epimerase function can probably be ruled out for these enzymes. The WbmF active site contains conserved 3,5-epimerase features, namely, a positionally conserved cysteine (Cys133) and basic side chain (His90 or Asn213), but lacks the serine/threonine component of the SDR triad and therefore may not act as an oxidoreductase. The data suggest a pathway for synthesis of the O-antigen precursor UDP-2,3-diacetamido-2,3-dideoxy-l-galacturonic acid and illustrate the usefulness of structural data in predicting protein function.
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PMID:Predicting protein function from structure--the roles of short-chain dehydrogenase/reductase enzymes in Bordetella O-antigen biosynthesis. 1795 Jul 51

Archaea and eukaryotes share a dolichol phosphate-dependent system for protein N-glycosylation. In both domains, the acetamido sugar N-acetylglucosamine (GlcNAc) forms part of the core oligosaccharide. However, the archaeal Methanococcales produce GlcNAc using the bacterial biosynthetic pathway. Key enzymes in this pathway belong to large families of proteins with diverse functions; therefore, the archaeal enzymes could not be identified solely using comparative sequence analysis. Genes encoding acetamido sugar-biosynthetic proteins were identified in Methanococcus maripaludis using phylogenetic and gene cluster analyses. Proteins expressed in Escherichia coli were purified and assayed for the predicted activities. The MMP1680 protein encodes a universally conserved glucosamine-6-phosphate synthase. The MMP1077 phosphomutase converted alpha-D-glucosamine-6-phosphate to alpha-D-glucosamine-1-phosphate, although this protein is more closely related to archaeal pentose and glucose phosphomutases than to bacterial glucosamine phosphomutases. The thermostable MJ1101 protein catalyzed both the acetylation of glucosamine-1-phosphate and the uridylyltransferase reaction with UTP to produce UDP-GlcNAc. The MMP0705 protein catalyzed the C-2 epimerization of UDP-GlcNAc, and the MMP0706 protein used NAD(+) to oxidize UDP-N-acetylmannosamine, forming UDP-N-acetylmannosaminuronate (ManNAcA). These two proteins are similar to enzymes used for proteobacterial lipopolysaccharide biosynthesis and gram-positive bacterial capsule production, suggesting a common evolutionary origin and a widespread distribution of ManNAcA. UDP-GlcNAc and UDP-ManNAcA biosynthesis evolved early in the euryarchaeal lineage, because most of their genomes contain orthologs of the five genes characterized here. These UDP-acetamido sugars are predicted to be precursors for flagellin and S-layer protein modifications and for the biosynthesis of methanogenic coenzyme B.
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PMID:Acetamido sugar biosynthesis in the Euryarchaea. 1826 21

Dysregulation of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) has been implicated in the pathogenesis of alcoholic liver injury. Sirtuin 1 (SIRT1) is an NAD(+)-dependent class III protein deacetylase that is known to be involved in regulating production of proinflammatory cytokines including TNF-alpha. In the present study, we examined the role of SIRT1 signaling in TNF-alpha generation stimulated by either lipopolysaccharide (LPS), acetaldehyde (AcH), or acetate (two major metabolites of ethanol) in two cultured macrophage cell lines. In both rat Kupffer cell line 1 (RKC1) and murine RAW 264.7 macrophages, treatment with either LPS, AcH, or acetate caused significant decreases in SIRT1 transcription, translation, and activation, which essentially demonstrated an inverse relationship with TNF-alpha levels. LPS, AcH, and acetate each provoked the release of TNF-alpha from RKC1 cells, whereas coincubation with resveratrol (a potent SIRT1 agonist) inhibited this effect. Conversely, addition of sirtinol (a known SIRT1 inhibitor) or knocking down SIRT1 by the small silencing SIRT1 plasmid (SIRT1shRNA) augmented TNF-alpha release, suggesting that impairment of SIRT1 may contribute to TNF-alpha secretion. Further mechanistic studies revealed that inhibition of SIRT1 by LPS, AcH, or acetate was associated with a marked increase in the acetylation of the RelA/p65 subunit of nuclear transcription factor (NF-kappaB) and promotion of NF-kappaB transcriptional activity. Taken together, our findings suggest that SIRT1-NF-kappaB signaling is involved in regulating LPS- and metabolites-of-ethanol-mediated TNF-alpha production in rat Kupffer cells and in murine macrophages. Our study provides new insights into understanding the molecular mechanisms underlying the development of alcoholic steatohepatitis.
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PMID:Role of SIRT1 in regulation of LPS- or two ethanol metabolites-induced TNF-alpha production in cultured macrophage cell lines. 1929 82

The B-band O-antigen of the lipopolysaccharide found in the opportunistic pathogen Pseudomonas aeruginosa PAO1 (serotype O5) comprises a repeating trisaccharide unit that is critical for virulence and protection from host defense systems. One of the carbohydrates in this repeating unit, the rare diacetylated aminuronic acid derivative 2,3-diacetamido-2,3-dideoxy-beta-d-mannuronic acid (ManNAc(3NAc)A), is thought to be produced by five enzymes (WbpA, WbpB, WbpE, WbpD, and WbpI) in a stepwise manner starting from UDP-GlcNAc. Although the genes responsible for the biosynthesis of this sugar are known, only two of the five encoded proteins (WbpA and WbpI) have been thoroughly investigated. In this report, we describe the cloning, overexpression, purification, and biochemical characterization of the three central enzymes in this pathway, WbpB, WbpE, and WbpD. Using a combination of capillary electrophoresis, RP-HPLC, and NMR spectroscopy, we show that WbpB and WbpE are a dehydrogenase/aminotransferase pair that converts UDP-GlcNAcA to UDP-GlcNAc(3NH(2))A in a coupled reaction via a unique NAD(+) recycling pathway. In addition, we confirm that WbpD catalyzes the acetylation of UDP-GlcNAc(3NH(2))A to give UDP-GlcNAc(3NAc)A. Notably, WbpA, WbpB, WbpE, WbpD, and WbpI can be combined in vitro to generate UDP-ManNAc(3NAc)A in a single reaction vessel, thereby providing supplies of this complex glycosyl donor for future studies of lipopolysaccharide assembly. This work completes the biochemical characterization of the enzymes in this pathway and provides novel targets for potential therapeutics to combat infections with drug resistant P. aeruginosa strains.
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PMID:Biosynthesis of UDP-GlcNAc(3NAc)A by WbpB, WbpE, and WbpD: enzymes in the Wbp pathway responsible for O-antigen assembly in Pseudomonas aeruginosa PAO1. 1934 2

Extracellular nicotinamide adenine dinucleotide (NAD(+)) is known to increase the intracellular calcium concentration [Ca(2+)](i) in different cell types and by various mechanisms. Here we show that NAD(+) triggers a transient rise in [Ca(2+)](i) in human monocytes activated with lipopolysaccharide (LPS), which is caused by a release of Ca(2+) from IP(3)-responsive intracellular stores and an influx of extracellular Ca(2+). By the use of P2 receptor-selective agonists and antagonists we demonstrate that P2 receptors play a role in the NAD(+)-induced calcium response in activated monocytes. Of the two subclasses of P2 receptors (P2X and P2Y) the P2Y receptors were considered the most likely candidates, since they share calcium signaling properties with NAD(+). The identification of P2Y(1) and P2Y(11) as receptor subtypes responsible for the NAD(+)-triggered increase in [Ca(2+)](i) was supported by several lines of evidence. First, specific P2Y(1) and P2Y(11) receptor antagonists inhibited the NAD(+)-induced increase in [Ca(2+)](i). Second, NAD(+) was shown to potently induce calcium signals in cells transfected with either subtype, whereas untransfected cells were unresponsive. Third, NAD(+) caused an increase in [cAMP](i), prevented by the P2Y(11) receptor-specific antagonist NF157.
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PMID:Extracellular NAD(+) induces a rise in [Ca(2+)](i) in activated human monocytes via engagement of P2Y(1) and P2Y(11) receptors. 1974 17

In the present study, we aimed to identify the synergistic effects of concurrent treatment of low concentrations of cilostazol and probucol to inhibit the oxidative stress with suppression of inflammatory markers in the cultured human coronary artery endothelial cells (HCAECs). Combination of cilostazol (0.3~3 microM) with probucol (0.03~0.3 microM) significantly suppressed TNF-alpha-stimulated NAD(P)H-dependent superoxide, lipopolysaccharide (LPS)-induced intracellular reactive oxygen species (ROS) production and TNF-alpha release in comparison with probucol or cilostazol alone. The combination of cilostazol (0.3~3 microM) with probucol (0.1~0.3 microM) inhibited the expression of vascular cell adhesion molecule-1 (VCAM-1) and monocyte chemoattractant protein-1 (MCP-1) more significantly than did the monotherapy with either probucol or cilostazol. In line with these results, combination therapy significantly suppressed monocyte adhesion to endothelial cells. Taken together, it is suggested that the synergistic effectiveness of the combination therapy with cilostazol and probucol may provide a beneficial therapeutic window in preventing atherosclerosis and protecting from cerebral ischemic injury.
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PMID:Synergistic efficacy of concurrent treatment with cilostazol and probucol on the suppression of reactive oxygen species and inflammatory markers in cultured human coronary artery endothelial cells. 1996 51

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