Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P43026 (lipopolysaccharide)
 62,215 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The enzyme 3-deoxy-D-manno-octulosonic acid (Kdo) transferase is encoded by the kdtA gene of Escherichia coli and plays a key role in lipopolysaccharide biosynthesis. It transfers Kdo from CMP-Kdo to lipid A or its tetraacyldisaccharide-1,4'-bisphosphate precursor, lipid IVA. Using a strain that overproduces the transferase approximately 500-fold, we have purified the enzyme to near homogeneity. The subunit molecular mass is approximately 43 kDa. Activity is stimulated by Triton X-100, is maximal at pH 7, but does not require Mg2+. The apparent Km values for lipid IVA and CMP-Kdo are 52 and 88 microM, respectively. Vmax is 15-18 mumol/min/mg when both substrates are added near saturation at pH 8. The purified enzyme transfers 2 Kdo residues to lipid A precursors or analogs bearing four to six fatty acyl chains and a 4'-monosphosphate moiety. Activity is inhibited by polymixin B and Re endotoxin. At low Kdo concentrations small amounts of the intermediate, (Kdo)1-IVA, accumulate. When this substance is isolated and incubated with purified enzyme in the presence of CMP-Kdo, it is converted to (Kdo)2-IVA. Formation of (Kdo)1-IVA is also observed when purified enzyme is incubated with (Kdo)2-IVA and 5 mM CMP, demonstrating that Kdo transfer is reversible. In summary, Kdo transferase consists of a single bifunctional polypeptide that incorporates the 2 innermost Kdo residues common to all lipopolysaccharide molecules in E. coli.
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PMID:Biosynthesis of endotoxins. Purification and catalytic properties of 3-deoxy-D-manno-octulosonic acid transferase from Escherichia coli. 157 28

The cDNA coding for xanthine dehydrogenase (XD) is isolated from mouse liver mRNA by cross-hybridization with a DNA fragment of the Drosophila melanogaster homologue. Two lambda bacteriophage overlapping clones represent the copy of a 4538-nucleotide-residue-long transcript with an open reading frame of 4005 nucleotide residues, coding for a putative polypeptide of 1335 amino acid residues. Comparison of the deduced amino acid sequence of the mouse XD with those of the Drosophila and the rat homologues shows a high conservation of this protein (55% identity between mouse and Drosophila, and 94% identity between mouse and rat). RNA blotting analysis demonstrates that interferon-alpha (IFN-alpha) and its inducers, i.e. poly(I).poly(C), bacterial lipopolysaccharide (LPS) and tilorone (2,7-bis-[2-(diethylamino)ethoxy]fluoren-9-one), increase the expression of XD mRNA in liver. Poly(I).poly(C) also induces XD mRNA in several other tissues in vivo. Protein synthesis de novo is not required for the elevation of XD mRNA after IFN-alpha treatment, since cycloheximide does not block the induction. The elevation of XD mRNA concentration is relatively fast and precedes the induction of both XD and xanthine oxidase (XO) enzymic activities.
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PMID:Molecular cloning of a cDNA coding for mouse liver xanthine dehydrogenase. Regulation of its transcript by interferons in vivo. 159 Jul 74

Helicobacter pylori is a gram-negative, curved, rod-shaped bacterium known to cause gastritis and to be an important factor in the pathogenesis of peptic ulcers. Serological testing has recently been proposed as an aid in diagnosis of H. pylori infections. In this study, we used the Western immunoblot technique to evaluate the possibility of using one or more of the antigens from H. pylori for this purpose. Thirteen major bands and about 30 minor bands could be identified by Western blotting when sera from 53 consecutive dyspeptic patients, 27 healthy children, and 25 blood donors were evaluated. Antibodies against most of the major bands were found significantly more frequently in patients with H. pylori infections than in patients without such infections. However, antibodies against a single polypeptide were not produced by all patients with H. pylori infection. Polypeptides with molecular masses of 120, 50, and between 19 and 36 kDa seem to be the most specific polypeptides for the diagnosis of H. pylori infections. This study showed only minor differences in antigenic composition between different clinical isolates of H. pylori, and serological cross-reactions with other bacteria were limited. Major serological cross-reactions were found only with Campylobacter jejuni and with bacterial lipopolysaccharide. However, one band at 60 kDa reacted with antiserum to the Legionella micdadei common antigen, which may indicate a cross-reaction with common antigen from several other bacteria. This study demonstrates that a number of bands may be useful as antigens in serological tests after isolation and purification.
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PMID:Immunoglobulin G antibodies to Helicobacter pylori in patients with dyspeptic symptoms investigated by the western immunoblot technique. 162 30

Transposon insertion, followed by screening, has allowed the identification of a set of genes, called htr, whose products are required for Escherichia coli growth at elevated temperatures. The htrB gene has been shown to map at 23.5 min on the E. coli genetic map. It codes for a very basic, hydrophobic, 35,000-Mr polypeptide, possessing a putative membrane-spanning domain. At the non-permissive temperature, htrB mutant bacteria stop dividing, followed by the formation of bulges and eventual lysis. The htrC gene maps at 90 min, is under sigma 32 regulation and codes for a 21, 130-Mr polypeptide. At 43 degrees C, htrC mutant bacteria gradually lyse, whereas at intermediate temperatures they filament extensively. Finally, the htrM gene maps at 81 min, is under sigma 32 regulation and codes for a 35,000-Mr polypeptide. The HtrM null phenotype included inability to grow above 42 degrees C, extreme mucoidness and sensitivity to bile salts, even at the permissive temperatures. The htrM gene is identical to the rfaD gene, whose product is required for the biosynthesis of the lipopolysaccharide precursor ADP-L-glycero-D-mannoheptose (Pegues et al., J. Bact., 1990, 172, 4652-4660).
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PMID:Complex phenotypes of null mutations in the htr genes, whose products are essential for Escherichia coli growth at elevated temperatures. 165 98

Interleukin 1 beta (IL1 beta) is an inducible polypeptide with many roles in host defence and homoeostasis. It has also been implicated as a mediator of infectious, inflammatory and autoimmune diseases, and the kinetics of its production are relevant to an understanding of the pathogenesis of these conditions. We report here the time-course of IL1 beta production in human adherent monocytes. Both IL1 beta protein and mRNA were measured following cell activation with bacterial endotoxin (lipopolysaccharide; LPS), and pro-inflammatory crystals of monosodium urate (MSU), which cause arthritis and kidney disease. We also tested other crystal types associated with arthritis, namely hydroxylapatite and calcium pyrophosphate dihydrate. IL1 was absent from unstimulated cells, but IL1 beta mRNA accumulated rapidly after LPS or MSU stimulation and was associated with the later appearance of intracellular IL1 beta protein which was subsequently released from the cells (60% at 9 h). The other crystals failed to induce significant IL1 production. Our findings support the view that production of IL1 beta in human mononuclear cells is based on rapid translation of an inducible pool of mRNA and that no pre-formed mRNA or intracellular protein exists in normal blood monocytes. Further, although IL1 beta is translated without a conventional leader sequence, it is translocated extracellularly with the kinetics of a secretory protein.
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PMID:Kinetics of IL1 beta mRNA and protein accumulation in human mononuclear cells. 166 80

The major outer membrane protein (Omp34) of Acidovorax delafieldii (formerly Pseudomonas delafieldii) was purified to homogeneity and was characterized biochemically and functionally. The polypeptide has an apparent molecular weight (Mr) of 34,000, and it forms stable oligomers at pH 9.0 in the presence of 10% octylpolyoxyethylene or 2% lithium dodecyl sulfate below 70 degrees C. The intact protein has a characteristic secondary structure composition, as revealed by Fourier transforming infrared spectroscopy (about 60% beta sheet). These features and the amino acid composition are typical for porins. The purified Omp34 is associated with 1 to 2 mol of lipopolysaccharide per mol of the monomer. Pore-forming activity was demonstrated with lipid bilayer experiments. Single-channel and selectivity measurements showed that the protein forms highly anion-selective channels. The unusual dependence of the single-channel conductance on salt concentration suggests that the porin complexes bear positive surface charges, accumulating negatively charged counterions at the pore mouth.
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PMID:The major outer membrane protein of Acidovorax delafieldii is an anion-selective porin. 171 13

The outer membranes (OMs) from serovars a, b, and c of Treponema denticola, originally isolated from periodontal patients, were prepared. Dialysis of the OMs against 20 mM MgCl2 yielded the aggregable (A) and the nonaggregable (NA) moieties of the OMs. The absence of muramic acid, adenosine triphosphatase, hexokinase, and nucleic acid as well as electron microscopy indicated that the OM preparations were homogeneous. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the A and NA moieties of the OMs showed approximately 25 Coomassie brilliant blue R-250 stain-positive bands or 47 silver-stained polypeptides. The relative molecular masses ranged between 14 and 97 kDa. The electrophoretic polypeptide profiles of the A and NA moieties shared many similarities among serovars a, b, and c. However, they exhibited variation in the overall pattern, intensity, or location of the polypeptide stained zones. This was especially true for serovar b. Two-dimensional electrophoretic studies showed an excess of 100 silver-stained spots with isoelectric points of 4.6 to 7.0 and relative molecular masses in the 14- to 97-kDa range. The OMs contained simple proteins, glycoproteins, and lipoproteins. The NA moieties of the OMs contained 4 to 6, 10 to 12, and 4 to 6 glycopeptides as well as two, seven, and two lipoprotein bands for serovars a, b, and c, respectively. The A moieties of the OMs showed 7 to 9, 11 to 13 and 5 to 6 glycopeptides as well as four, five, and three lipoprotein bands for serovars a, b, and c, respectively. Lipopolysaccharide was detected in the OMs of the three serovars following removal of proteins with proteinase K, pronase and silver staining of sodium dodecyl sulfate-polyacrylamide gels, or removal of lipopolysaccharide from the OMs by hot phenol extraction. The 66- and 53-kDa bands were present in serovars b and c, while a band with a relative molecular mass of 45 kDa was present only in serovar c. Endotoxin-like activity was also shown in the OMs of the three serovars by the Limulus amebocyte clotting assay and the chick embryo lethality test. This is the first report on selected biochemical properties of the OM macromolecules of three known serovars of T. denticola.
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PMID:Biochemical properties of the outer membrane of Treponema denticola. 171 83

The conductance properties of three members of the porin family which form channels across the outer membrane of Gram-negative bacteria were compared. With their endogenous lipopolysaccharide (LPS) bound, the closely related porins F and C from Escherichia coli reveal significantly different conductance steps and closing potentials, with values of 0.82 nS (nanosiemens) and 89 mV for F-type channels, and 0.49 nS and 158 mV for C-type pores (1 M NaCl), respectively. On the basis of their closing potentials, the two channel types can be distinguished unequivocally. If reconstituted in asolectin and extraneous LPS, porin C forms F-type in addition to C-type channels. Substitution of asolectin by mitochondrial lipids yields the native C-type pores only. Both channel types can be induced to assume the mutually other channel configuration by variation of ionic strength. A multiplicity of channel subtypes is observed by variation of the pH of the medium. The three channels within a trimer are, however, consistently of the same type. Since structural studies have revealed a single channel per monomer, the several conductance steps observed are likely to reflect distinct configurations of the same channel. Best channel recoveries were observed if endogenous LPS remained associated to porin during purification. Significant yields could nevertheless be obtained also if LPS was removed from porin and replaced with various precursors or chemically synthesized analogues. As function requires the presence of glycolipids, yet crystallization is perturbed by heterodisperse endogenous LPS, the smallest monodisperse analogues yielding good channel recovery were determined. The minimal synthetic moiety is a monoglucosaminetetraacyl compound. The characteristics of porin B from E. coli BE are shown to be indistinguishable from those of porin F. The conductance properties of this porin, refolded from random coil configuration, are indistinguishable from those exhibited by native protein. The formation of channels is thus encoded by the sequence of the mature polypeptide alone.
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PMID:Plasticity of Escherichia coli porin channels. Dependence of their conductance on strain and lipid environment. 172 4

Tumour necrosis factor alpha (TNF alpha) is a cytokine with a wide range of effects on both lymphoid and non-lymphoid cell types. By hybridization with a human TNF alpha cDNA probe the corresponding ovine cDNA was isolated from a lipopolysaccharide (LPS) stimulated alveolar macrophage cDNA library. The sequence of the cDNA clone showed that ovine TNF alpha encodes a polypeptide of 234 amino acids that, based on analysis of human TNF alpha, is processed to a protein of 157 amino acids. The nucleotide and amino acid sequences showed a high degree of homology to the equivalent human and mouse molecules. In a mammalian COS cell expression system the ovine cDNA was found to encode a protein which was able to lyse actinomycin-D treated WEHI-164 cells and induce COS cells to produce and secrete interleukin 6 (IL-6). Further experiments demonstrated the importance of sequences within the 3' untranslated region of the cDNA in determining the level of expression of ovine TNF alpha. Northern blot analysis was used to analyse the kinetics of induction of ovine TNF alpha mRNA in alveolar macrophages stimulated with a variety of mitogens. Addition of LPS increased mRNA encoding TNF alpha at 1 h and 5 h but not 24 h post stimulation. In contrast, addition of phorbol myristic acid (PMA) led to increased TNF alpha mRNA at 5 h while the combination of PMA and ionomycin increased the level of specific mRNA detected at 1 h, 5 h and 24 h. From genomic analysis ovine TNF alpha appears to exist as a single copy.
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PMID:Molecular cloning, expression and characterization of ovine TNF alpha. 178 96

The T-even type Escherichia coli phage Ox2 recognizes the outer membrane protein OmpA as a receptor. This recognition is accomplished by the 266 residue protein 38, which is located at the free ends of the virion's long tail fibers. Host-range mutants had been isolated in three consecutive steps: Ox2----Ox2h5----Ox2h10----Ox2h12, with Ox2h12 recognizing the outer membrane protein OmpC efficiently and having lost some affinity for OmpA. Protein 38 consists, in comparison with these proteins of other phages, of two constant and one contiguous array of four hypervariable regions; the alterations leading to Ox2h12 were all found within the latter area. Starting with Ox2h12, further host-range mutants could be isolated on strains resistant to the respective phage: Ox2h12----h12h1----h12h1.1----h12h1.11----h12 h1.111. It was found that Ox2h12h1.1 (and a derivative of Ox2h10, h10h4) probably uses, instead of OmpA or OmpC, yet another outer membrane protein, designated OmpX. Ox2h12h1.11 was obtained on a strain lacking OmpA, -C and -X. This phage could not grow on a mutant of E. coli B, possessing a lipopolysaccharide (LPS) with a defective core oligosaccharide; Ox2h12h1.111 was obtained from this strain. It turned out that the latter two mutants used LPS as a receptor, most likely via its glucose residues. Selection for resistance to them in E. coli B (ompA+, ompC-, ompX-) yielded exclusively LPS mutants, and in another strain, possessing OmpA, C and X, the majority of resistant mutants were of this type. Isolated LPS inactivated the mutant phages very well and was inactive towards Ox2h12. By recombining the genes of mutant phages into the genome of parental phages it could be shown that the phenotypes were associated with gene 38. All mutant alterations (mostly single amino acid substitutions) were found within the hypervariable regions of protein 38. In particular, a substitution leading to Ox2h12h1.11 (Arg170----Ser) had occurred at the same site that led to Ox2h10 (His170----Arg), which binds to OmpC in addition to OmpA. It is concluded that not only can protein 38 gain the ability to switch from a protein to a carbohydrate as a receptor but can do so using the same domain of the polypeptide.
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PMID:Single mutations in a gene for a tail fiber component of an Escherichia coli phage can cause an extension from a protein to a carbohydrate as a receptor. 182 15


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