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Other Gram-negative Bacteria (other + gram-negative_bacteria)
Selected AbstractsMultiple polypeptide forms observed in two-dimensional gels of Methylococcus capsulatus (Bath) polypeptides are generated during the separation procedureELECTROPHORESIS, Issue 4 2003Frode S. Berven Abstract We have examined two-dimensional electrophoresis (2-DE) gel maps of polypeptides from the Gram-negative bacterium Methylococcus capsulatus (Bath) and found the same widespread trains of spots as often reported in 2-DE gels of polypeptides of other Gram-negative bacteria. Some of the trains of polypeptides, both from the outer membrane and soluble protein fraction, were shown to be generated during the separation procedure of 2-DE, and not by covalent post-translational modifications. The trains were found to be regenerated when rerunning individual polypeptide spots. The polypeptides analysed giving this type of trains were all found to be classified as stable polypeptides according to the instability index of Guruprasad et al. (Protein Eng. 1990, 4, 155,161). The phenomenon most likely reflects conformational equilibria of polypeptides arising from the experimental conditions used, and is a clear drawback of the standard 2-DE procedure, making the gel picture unnecessarily complex to analyse. [source] The periplasmic peptidyl prolyl cis,trans isomerases PpiD and SurA have partially overlapping substrate specificitiesFEBS JOURNAL, Issue 13 2008Krista H. Stymest One of the rate-limiting steps in protein folding has been shown to be the cis,trans isomerization of proline residues, catalysed by a range of peptidyl prolyl cis,trans isomerases (PPIases). In the periplasmic space of Escherichia coli and other Gram-negative bacteria, two PPIases, SurA and PpiD, have been identified, which show high sequence similarity to the catalytic domain of the small PPIase parvulin. This observation raises a question regarding the biological significance of two apparently similar enzymes present in the same cellular compartment: do they interact with different substrates or do they catalyse different reactions? The substrate-binding motif of PpiD has not been characterized so far, and no biochemical data were available on how this folding catalyst recognizes and interacts with substrates. To characterize the interaction between model peptides and the periplasmic PPIase PpiD from E. coli, we employed a chemical crosslinking strategy that has been used previously to elucidate the interaction of substrates with SurA. We found that PpiD interacted with a range of model peptides independently of whether they contained proline residues or not. We further demonstrate here that PpiD and SurA interact with similar model peptides, and therefore must have partially overlapping substrate specificities. However, the binding motif of PpiD appears to be less specific than that of SurA, indicating that the two PPIases might interact with different substrates. We therefore propose that, although PpiD and SurA have partially overlapping substrate specificities, they fulfil different functions in the cell. [source] Ubiquinone biosynthesis in microorganismsFEMS MICROBIOLOGY LETTERS, Issue 2 2001R Meganathan Abstract The quinoid nucleus of the benzoquinone, ubiquinone (coenzyme Q; Q), is derived from the shikimate pathway in bacteria and eukaryotic microorganisms. Ubiquinone is not considered a vitamin since mammals synthesize it from the essential amino acid tyrosine. Escherichia coli and other Gram-negative bacteria derive the 4-hydroxybenzoate required for the biosynthesis of Q directly from chorismate. The yeast, Saccharomyces cerevisiae, can either form 4-hydroxybenzoate from chorismate or tyrosine. However, unlike mammals, S. cerevisiae synthesizes tyrosine in vivo by the shikimate pathway. While the reactions of the pathway leading from 4-hydroxybenzoate to Q are the same in both organisms the order in which they occur differs. The 4-hydroxybenzoate undergoes a prenylation, a decarboxylation and three hydroxylations alternating with three methylation reactions, resulting in the formation of Q. The methyl groups for the methylation reactions are derived from S -adenosylmethionine. While the prenyl side chain is formed by the 2- C -methyl- d -erythritol 4-phosphate (non-mevalonate) pathway in E. coli, it is formed by the mevalonate pathway in the yeast. [source] Detection of quinolone-resistance genes in Photobacterium damselae subsp. piscicida strains by targeting-induced local lesions in genomesJOURNAL OF FISH DISEASES, Issue 8 2005M-J Kim Abstract Quinolone-resistant strains of the fish-pathogenic bacterium, Photobacterium damselae subsp. piscicida are distributed widely in cultured yellowtail, Seriola quinqueradiata (Temminck & Schlegel), in Japan. The quinolone resistance-determining region (QRDR) was amplified with degenerate primers, followed by cassette ligation-mediated PCR. Open reading frames encoding proteins of 875 and 755 amino acid residues were detected in the gyrA and parC genes, respectively. Resistant strains of P. damselae subsp. piscicida carried a point mutation only in the gyrA QRDR leading to a Ser-to-Ile substitution at residue position 83. No amino acid alterations were discovered in the ParC sequence. A mutation in the gyrA gene was also detected in nalidixic acid-resistant mutants of strain SP96002 obtained from agar medium containing increased levels of quinolone. These results suggest that GyrA, as in other Gram-negative bacteria, is a target of quinolone in P. damselae subsp. piscicida. Furthermore, we attempted to detect a point mutation using targeting-induced local lesions in genomes (TILLING), which is a general strategy used for the detection of a variety of induced point mutations and naturally occurring polymorphisms. We developed a new detection method for the rapid and large-scale identification of quinolone-resistant strains of P. damselae subsp. piscicida using TILLING. [source] Novel ultrastructures of Treponema primitia and their implications for motilityMOLECULAR MICROBIOLOGY, Issue 6 2008Gavin E. Murphy Summary Members of the bacterial phylum Spirochaetes are generally helical cells propelled by periplasmic flagella. The spirochete Treponema primitia is interesting because of its mutualistic role in the termite gut, where it is believed to cooperate with protozoa that break down cellulose and produce H2 as a by-product. Here we report the ultrastructure of T. primitia as obtained by electron cryotomography of intact, frozen-hydrated cells. Several previously unrecognized external structures were revealed, including bowl-like objects decorating the outer membrane, arcades of hook-shaped proteins winding along the exterior and tufts of fibrils extending from the cell tips. Inside the periplasm, cone-like structures were found at each pole. Instead of the single peptidoglycan layer typical of other Gram-negative bacteria, two distinct periplasmic layers were observed. These layers formed a central open space that contained two flagella situated adjacent to each other. In some areas, the inner membrane formed flattened invaginations that protruded into the cytoplasm. High-speed light microscopic images of swimming T. primitia cells showed that cell bodies remained rigid and moved in a helical rather than planar motion. Together, these findings support the ,rolling cylinder' model for T. primitia motility that posits rotation of the protoplasmic cylinder within the outer sheath. [source] Characterization of HasB, a Serratia marcescens TonB-like protein specifically involved in the haemophore-dependent haem acquisition systemMOLECULAR MICROBIOLOGY, Issue 4 2001Annick Paquelin In Gram-negative bacteria, the TonB,ExbB,ExbD inner membrane multiprotein complex is required for active transport of diverse molecules through the outer membrane. We present evidence that Serratia marcescens, like several other Gram-negative bacteria, has two TonB proteins: the previously characterized TonBSM, and also HasB, a newly identified component of the has operon that encodes a haemophore-dependent haem acquisition system. This system involves a soluble extracellular protein (the HasA haemophore) that acquires free or haemoprotein-bound haem and presents it to a specific outer membrane haemophore receptor (HasR). TonBSM and HasB are significantly similar and can replace each other for haem acquisition. However, TonBSM, but not HasB, mediates iron acquisition from iron sources other than haem and haemoproteins, showing that HasB and TonBSM only display partial redundancy. The reconstitution in Escherichia coli of the S. marcescens Has system demonstrated that haem uptake is dependent on the E. coli ExbB, ExbD and TonB proteins and that HasB is non-functional in E. coli. Nevertheless, a mutation in the HasB transmembrane anchor domain allows it to replace TonBEC for haem acquisition. As the change affects a domain involved in specific TonBEC,ExbBEC interactions, HasB may be unable to interact with ExbBEC, and the HasB mutation may allow this interaction. In E. coli, the HasB mutant protein was functional for haem uptake but could not complement the other TonBEC -dependent functions, such as iron siderophore acquisition, and phage DNA and colicin uptake. Our findings support the emerging hypothesis that TonB homologues are widespread in bacteria, where they may have specific functions in receptor,ligand uptake systems. [source] Fishing new proteins in the twilight zone of genomes: The test case of outer membrane proteins in Escherichia coli K12, Escherichia coli O157:H7, and other Gram-negative bacteriaPROTEIN SCIENCE, Issue 6 2003Rita Casadio Abstract We address the problem of clustering the whole protein content of genomes into three different categories,globular, all-,, and all-, membrane proteins,with the aim of fishing new membrane proteins in the pool of nonannotated proteins (twilight zone). The focus is then mainly on outer membrane proteins. This is performed by using an integrated suite of programs (Hunter) specifically developed for predicting the occurrence of signal peptides in proteins of Gram-negative bacteria and the topography of all-, and all-, membrane proteins. Hunter is tested on the well and partially annotated proteins (2160 and 760, respectively) of Escherichia coli K 12 scoring as high as 95.6% in the correct assignment of each chain to the category. Of the remaining 1253 nonannotated sequences, 1099 are predicted globular, 136 are all-,, and 18 are all-, membrane proteins. In Escherichia coli 0157:H7 we filtered 1901 nonannotated proteins. Our analysis classifies 1564 globular chains, 327 inner membrane proteins, and 10 outer membrane proteins. With Hunter, new membrane proteins are added to the list of putative membrane proteins of Gram-negative bacteria. The content of outer membrane proteins per genome (nine are analyzed) ranges from 1.5% to 2.4%, and it is one order of magnitude lower than that of inner membrane proteins. The finding is particularly relevant when it is considered that this is the first large-scale analysis based on validated tools that can predict the content of outer membrane proteins in a genome and can allow cross-comparison of the same protein type between different species. [source] The Structure of a Novel Neutral Lipid,A from the Lipopolysaccharide of Bradyrhizobium elkanii Containing Three Mannose Units in the BackboneCHEMISTRY - A EUROPEAN JOURNAL, Issue 9 2010Iwona Komaniecka Dr. Abstract The chemical structure of the lipid,A of the lipopolysaccharide (LPS) from Bradyrhizobium elkanii USDA 76 (a member of the group of slow-growing rhizobia) has been established. It differed considerably from lipids,A of other Gram-negative bacteria, in that it completely lacks negatively charged groups (phosphate or uronic acid residues); the glucosamine (GlcpN) disaccharide backbone is replaced by one consisting of 2,3-dideoxy-2,3-diamino- D -glucopyranose (GlcpN3N) and it contains two long-chain fatty acids, which is unusual among rhizobia. The GlcpN3N disaccharide was further substituted by three D -mannopyranose (D -Manp) residues, together forming a pentasaccharide. To establish the structural details of this molecule, 1D and 2D,NMR spectroscopy, chemical composition analyses and high-resolution mass spectrometry methods (electrospray ionisation Fourier-transform ion cyclotron resonance mass spectrometry (ESI FT-ICR MS) and tandem mass spectrometry (MS/MS)) were applied. By using 1D and 2D,NMR spectroscopy experiments, it was confirmed that one D -Manp was linked to C-1 of the reducing GlcpN3N and an ,-(1,6)-linked D -Manp disaccharide was located at C-4, of the non-reducing GlcpN3N (,-linkage). Fatty acid analysis identified 12:0(3-OH) and 14:0(3-OH), which were amide-linked to GlcpN3N. Other lipid,A constituents were long (,-1)-hydroxylated fatty acids with 26,33 carbon atoms, as well as their oxo forms (28:0(27-oxo) and 30:0(29-oxo)). The 28:0(27-OH) was the most abundant acyl residue. As confirmed by high-resolution mass spectrometry techniques, these long-chain fatty acids created two acyloxyacyl residues with the 3-hydroxy fatty acids. Thus, lipid,A from B. elkanii comprised six acyl residues. It was also shown that one of the acyloxyacyl residues could be further acylated by 3-hydroxybutyric acid (linked to the (,-1)-hydroxy group). [source] Burkholderia pseudomallei stimulates low interleukin-8 production in the human lung epithelial cell line A549CLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 1 2004P. UTAISINCHAROEN SUMMARY Melioidosis is a life-threatening disease caused by Burkholderia pseudomallei. The lung is the most commonly affected organ, resulting in abscess formation in patients with chronic melioidosis. Previous study has shown that B. pseudomallei was able to invade and multiply in epithelial cells. In the present study, we have demonstrated that B. pseudomallei is able to stimulate interleukin 8 (IL-8) production from the human alveolar lung epithelium cell line A549. However, the level of IL-8 production was significantly lower than when the cells were infected with other Gram-negative bacteria such as Salmonella enterica serovar Typhi (S. typhi) which were used for comparison. The degree of I,B, degradation in the B. pseudomallei -infected cells was lower than that of the S. typhi -infected cells, suggesting that B. pseudomallei is also a poorer cell activator. Inhibition of B. pseudomallei invasion by cytochalasin D did not interfere with either IL-8 production or I,B, degradation, indicating that bacterial uptake is not required for the production of this chemokine. Thus, it appears that the signalling initiated by the interaction of B. pseudomallei with the epithelial cell surface is sufficient for epithelial cell activation. [source] | |||||||||||||