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Bacterium Escherichia Coli (bacterium + escherichia_coli)
Selected AbstractsTheoretical study of lipid biosynthesis in wild-type Escherichia coli and in a protoplast-type L-form using elementary flux mode analysisFEBS JOURNAL, Issue 4 2010Dimitar Kenanov In the present study, we investigated lipid biosynthesis in the bacterium Escherichia coli by mathematical modeling. In particular, we studied the interaction between the subsystems producing unsaturated and saturated fatty acids, phospholipids, lipid A, and cardiolipin. The present analysis was carried out both for the wild-type and for several in silico knockout mutants, using the concept of elementary flux modes. Our results confirm that, in the wild type, there are four main products: L1-phosphatidylethanolamine, lipid A, lipid A (cold-adapted), and cardiolipin. We found that each of these compounds is produced on several different routes, indicating a high redundancy of the system under study. By analysis of the elementary flux modes remaining after the knockout of genes of lipid biosynthesis, and comparison with publicly available data on single-gene knockouts in vivo, we were able to determine the metabolites essential for the survival of the cell. Furthermore, we analyzed a set of mutations that occur in a cell wall-free mutant of Escherichia coli W1655F+. We postulate that the mutant is not capable of producing both forms of lipid A, when the combination of mutations is considered to make a nonfunctional pathway. This is in contrast to gene essentiality data showing that lipid A synthesis is indispensable for the survival of the cell. The loss of the outer membrane in the cell wall-free mutant, however, shows that lipid A is dispensable as the main component of the outer surface structure in this particular E. coli strain. [source] 2-Methylisocitrate lyases from the bacterium Escherichia coli and the filamentous fungus Aspergillus nidulansFEBS JOURNAL, Issue 12 2001Characterization, comparison of both enzymes In Escherichia coli and Aspergillus nidulans, propionate is oxidized to pyruvate via the methylcitrate cycle. The last step of this cycle, the cleavage of 2-methylisocitrate to succinate and pyruvate is catalysed by 2-methylisocitrate lyase. The enzymes from both organisms were assayed with chemically synthesized threo -2-methylisocitrate; the erythro -diastereomer was not active. 2-Methylisocitrate lyase from E. coli corresponds to the PrpB protein of the prp operon involved in propionate oxidation. The purified enzyme has a molecular mass of approximately 32 kDa per subunit, which is lower than those of isocitrate lyases from bacterial sources (, 48 kDa). 2-Methylisocitrate lyase from A. nidulans shows an apparent molecular mass of 66 kDa per subunit, almost equal to that of isocitrate lyase of the same organism. Both 2-methylisocitrate lyases have a native homotetrameric structure as identified by size-exclusion chromatography. The enzymes show no measurable activity with isocitrate. Starting from 250 mm pyruvate, 150 mm succinate and 10 µm PrpB, the enzymatically active stereoisomer could be synthesized in 1% yield. As revealed by chiral HPLC, the product consisted of a single enantiomer. This isomer is cleaved by 2-methylisocitrate lyases from A. nidulans and E. coli. The PrpB protein reacted with stoichiometric amounts of 3-bromopyruvate whereby the activity was lost and one amino-acid residue per subunit became modified, most likely a cysteine as shown for isocitrate lyase of E. coli. PrpB exhibits 34% sequence identity with carboxyphosphoenolpyruvate phosphonomutase from Streptomyces hygroscopicus, in which the essential cysteine residue is conserved. [source] Growth rate dependent numbers of SeqA structures organize the multiple replication forks in rapidly growing Escherichia coliGENES TO CELLS, Issue 5 2009Morigen When the bacterium Escherichia coli is grown in rich medium, the replication and segregation periods may span two, three or four generations and cells may contain up to 24 replication forks. The newly synthesized, hemimethylated DNA at each fork is bound by SeqA protein. The SeqA,DNA structures form distinct foci that can be observed by immunofluorescence microscopy. The numbers of foci were lower than the numbers of replication forks indicating fork co-localization. The extent of co-localization correlated with the extent of replication cycle overlap in wild-type cells. No abrupt increase in the numbers of foci occurred at the time of initiation of replication, suggesting that new replication forks bind to existing SeqA structures. Manipulations with replication control mechanisms that led to extension or reduction of the replication period and number of forks, did not lead to changes in the numbers of SeqA foci per cell. The results indicate that the number of SeqA foci is not directly governed by the number of replication forks, and supports the idea that new DNA may be ,captured' by existing SeqA structures. [source] Antibacterial effect of silver nanoparticles deposited on corona-treated polyester and polyamide fabricsPOLYMERS FOR ADVANCED TECHNOLOGIES, Issue 12 2008Maja Radeti Abstract The possibility of using a corona treatment (electrical discharge at atmospheric pressure) for fiber surface activation, which can facilitate the loading of silver nanoparticles (NPs) from colloids onto the polyester (PES) and polyamide (PA) fabrics and thus improve their antibacterial properties, was studied. Bactericidal efficiency and its laundering durability on silver-loaded fabrics for Gram-positive bacterium Staphylococcus aureus and Gram-negative bacterium Escherichia coli were evaluated. The fiber morphology after corona treatment and subsequent loading of silver NPs was followed by SEM. Corona-treated fabrics loaded with silver NPs exhibited better antibacterial properties in comparison with untreated fabrics. In order to obtain acceptable laundering durability, it is necessary to use highly concentrated silver colloids. Copyright © 2008 John Wiley & Sons, Ltd. [source] Microplitis demolitor bracovirus inhibits phagocytosis by hemocytes from Pseudoplusia includens,ARCHIVES OF INSECT BIOCHEMISTRY AND PHYSIOLOGY (ELECTRONIC), Issue 3 2006Michael R. Strand Abstract The braconid wasp Microplitis demolitor carries Microplitis demolitor bracovirus (MdBV) and parasitizes the larval stage of several noctuid moths. A key function of MdBV in parasitism is suppression of the host's cellular immune response. Prior studies in the host Pseudoplusia includens indicated that MdBV blocks encapsulation by preventing two types of hemocytes, plasmatocytes and granulocytes, from adhering to foreign targets. The other main immune response mediated by insect hemocytes is phagocytosis. The goal of this study was to determine which hemocyte types were phagocytic in P. includens and to assess whether MdBV infection affects this defense response. Using the bacterium Escherichia coli and inert polystyrene beads as targets, our results indicated that the professional phagocyte in P. includens is granulocytes. The phagocytic responses of granulocytes were very similar to those of High Five cells that prior studies have suggested are a granulocyte-like cell line. MdBV infection dose-dependently disrupted phagocytosis in both cell types by inhibiting adhesion of targets to the cell surface. The MdBV glc1.8 gene encodes a cell surface glycoprotein that had previously been implicated in disruption of adhesion and encapsulation responses by immune cells. Knockdown of glc1.8 expression by RNA interference (RNAi) during the current study rescued the ability of MdBV-infected High Five cells to phagocytize targets. Collectively, these results indicate that glc1.8 is a key virulence determinant in disruption of both adhesion and phagocytosis by insect immune cells. Arch. Insect Biochem. Physiol. 61:134,145, 2006. © 2006 Wiley-Liss, Inc. [source] Genetic analysis of G protein-coupled receptor expression in Escherichia coli: Inhibitory role of DnaJ on the membrane integration of the human central cannabinoid receptorBIOTECHNOLOGY & BIOENGINEERING, Issue 2 2009Georgios Skretas Abstract The overexpression of G protein-coupled receptors (GPCRs) and of many other heterologous membrane proteins in simple microbial hosts, such as the bacterium Escherichia coli, often results in protein mistargeting, aggregation into inclusion bodies or cytoplasmic degradation. Furthermore, membrane protein production is very frequently accompanied by severe cell toxicity. In this work, we have employed a genetic strategy to isolate E. coli mutants that produce markedly increased amounts of the human central cannabinoid receptor (CB1), a pharmacologically significant GPCR that expresses very poorly in wild-type E. coli. By utilizing a CB1 fusion with the green fluorescent protein (GFP) and fluorescence-activated cell sorting (FACS), we screened an E. coli transposon library and identified an insertion in dnaJ that resulted in a large increase in CB1-GFP fluorescence and a dramatic enhancement in bacterial production of membrane-integrated CB1. Furthermore, the dnaJ::Tn5 inactivation suppressed the severe cytotoxicity associated with CB1 production. This revealed an unexpected inhibitory role of the chaperone/ co-chaperone DnaJ in the protein folding or membrane insertion of bacterially produced CB1. Our strategy can be easily adapted to identify expression bottlenecks for different GPCRs or any other integral membrane protein, provide useful and unanticipated mechanistic insights, and assist in the construction of genetically engineered E. coli strains for efficient heterologous membrane protein production. Biotechnol. Bioeng. 2009;102: 357,367. © 2008 Wiley Periodicals, Inc. [source] Quantum-Dot-Labeled DNA Probes for Fluorescence In Situ Hybridization (FISH) in the Microorganism Escherichia coliCHEMPHYSCHEM, Issue 5 2006Sheng-Mei Wu Abstract Semiconductor quantum dots (QDs) as a kind of nonisotopic biological labeling material have many unique fluorescent properties relative to conventional organic dyes and fluorescent proteins, such as composition- and size-dependent absorption and emission, a broad absorption spectrum, photostability, and single-dot sensitivity. These properties make them a promising stable and sensitive label, which can be used for long-term fluorescent tracking and subcellular location of genes and proteins. Here, a simple approach for the construction of QD-labeled DNA probes was developed by attaching thiol-ssDNA to QDs via a metal,thiol bond. The as-prepared QD-labeled DNA probes had high dispersivity, bioactivity, and specificity for hybridization. Based on such a kind of probe with a sequence complementary to multiple clone sites in plasmid pUC18, fluorescence in situ hybridization of the tiny bacterium Escherichia coli has been realized for the first time. [source] | |||||||||||||