scholarly journals The reconstitution of oxidase activity in membranes derived from a 5-aminolaevulinic acid-requiring mutant of Escherichia coli

1973 ◽  
Vol 136 (4) ◽  
pp. 877-884 ◽  
Author(s):  
Bruce A. Haddock
Keyword(s):  
Escherichia Coli ◽  
Electron Transport ◽  
Cytochrome B ◽  
Oxidase Activity ◽  
Nadh Dehydrogenase ◽  
Carbon Sources ◽  
Protoporphyrin Ix ◽  
Side Chain ◽  
E Coli ◽  
Aminolaevulinic Acid

1. The reconstitution of oxidase activity in cell-free extracts of a mutant of Escherichia coli K12Ymel, that require 5-aminolaevulinic acid for growth on non-fermentable carbon sources, is described. 2. The reconstitution is dependent on haematin or a haem extract from a prototrophic strain of E. coli, and the product of the reaction has been identified as NADH-reducible cytochrome b. 3. The requirement for haematin cannot be replaced by four other porphyrins. Coproporphyrin III does not inhibit the haematin-dependent reconstitution, mesoporphyrin IX and protoporphyrin IX apparently compete with haematin for a binding site on the cytochrome apoprotein(s) and deuteroporphyrin IX binds to cytochrome apoprotein(s) and cannot be subsequently replaced by haematin. 4. The properties of electron-transport particles from cell-free extracts of the mutant strain, grown aerobically in the presence or absence of 5-aminolaevulinic acid, are described. In the absence of 5-aminolaevulinic acid no detectable cytochromes are produced, and oxidase activities are lowered but there is no apparent effect on the activities of the NADH dehydrogenase and d-lactate dehydrogenase. 5. The reconstitution of oxidase activity by electron-transport particles from cells grown in the absence of 5-aminolaevulinic acid requires ATP and haematin, and the product of the reaction was identified as NADH-reducible cytochrome b. 6. It is concluded that the cytochrome apoproteins are synthesized and incorporated into the cytoplasmic membrane of E. coli in the absence of haem synthesis. The subsequent reconstitution of functional cytochrome(s) requires protohaem, but the nature of the side chain on the 2 and 4 positions of the porphyrin appears to be important.

scholarly journals Characterization and phenotypic control of the cytochrome content of Escherichia coli

1979 ◽  
Vol 182 (2) ◽  
pp. 465-472 ◽  
Author(s):  
Graeme A. Reid ◽  
W. John Ingledew
Keyword(s):  
Escherichia Coli ◽  
Electron Transport ◽  
Cytochrome B ◽  
Terminal Electron Acceptor ◽  
E Coli ◽  
Reduction Potentials ◽  
Electrode Potentials ◽  
Point Potential ◽  
Oxidation Reduction ◽  
Cytochrome D

1. Electron-transport particles derived from Escherichia coli grown aerobically contain three b-type cytochromes with mid-point oxidation–reduction potentials at pH7 of +260mV, +80mV and −50mV, with n=1 for each. The variation of these values with pH was determined. 2. E. coli develops a different set of b-type cytochromes when grown anaerobically on glycerol with fumarate or nitrate as terminal electron acceptor. Electron-transport particles of fumarate-grown cells contain b-type cytochromes with mid-point potentials at pH7 of +140mV and +250mV (n=1). These two cytochromes are also present in cells grown with nitrate as terminal acceptor, where an additional cytochrome b with a mid-point potential of +10mV (n=1) is developed. 3. The wavelengths of the α-absorption-band maxima of the b-type cytochromes at 77K were: (a) for aerobically grown cells, cytochrome b (Em7 +260mV), 556nm and 563nm, cytochrome b (Em7 +80mV), 556nm and cytochrome b (Em7−50mV), 558nm; (b) for anaerobically grown cells, cytochrome b (Em7 +250mV), 558nm, cytochrome b (Em7 +40mV), 555nm and cytochrome b (Em7 +10mV), 556nm. 4. Cytochrome d was found to have a mid-point potential at pH7 of +280mV (n=1). 5. Cytochrome a1 was resolved as two components of equal magnitude with mid-point potentials of +260mV and +160mV (n=1). 6. Redox titrations performed in the presence of CO showed that one of the b-type cytochromes in the aerobically grown cultures was reduced, even at the upper limits of our range of electrode potentials (above +400mV). Cytochrome d was also not oxidizable in the presence of CO. Neither of the cytochromes a1 was affected by the presence of CO.

scholarly journals Binding Site of the Bridged Macrolides in the Escherichia coli Ribosome

2005 ◽  
Vol 49 (1) ◽  
pp. 281-288 ◽  
Author(s):  
Liqun Xiong ◽  
Yakov Korkhin ◽  
Alexander S. Mankin
Keyword(s):  
Escherichia Coli ◽  
Tight Binding ◽  
Dimethyl Sulfate ◽  
23S Rrna ◽  
Side Chain ◽  
Macrolide Antibiotics ◽  
Side Chains ◽  
Alkyl Aryl ◽  
E Coli ◽  
Exit Tunnel

ABSTRACT Ketolides represent the latest group of macrolide antibiotics. Tight binding of ketolides to the ribosome appears to correlate with the presence of an extended alkyl-aryl side chain. Recently developed 6,11-bridged bicyclic ketolides extend the spectrum of platforms used to generate new potent macrolides with extended alkyl-aryl side chains. The purpose of the present study was to characterize the site of binding and the action of bridged macrolides in the ribosomes of Escherichia coli. All the bridged macrolides investigated efficiently protected A2058 and A2059 in domain V of 23S rRNA from modification by dimethyl sulfate and U2609 from modification by carbodiimide. In addition, bridged macrolides that carry extended alkyl-aryl side chains protruding from the 6,11 bridge protected A752 in helix 35 of domain II of 23S rRNA from modification by dimethyl sulfate. Bridged macrolides efficiently displaced erythromycin from the ribosome in a competition binding assay. The A2058G mutation in 23S rRNA conferred resistance to the bridged macrolides. The U2609C mutation, which renders E. coli resistant to the previously studied ketolides telithromycin and cethromycin, barely affected cell susceptibility to the bridged macrolides used in this study. The results of the biochemical and genetic studies indicate that in the E. coli ribosome, bridged macrolides bind in the nascent peptide exit tunnel at the site previously described for other macrolide antibiotics. The presence of the side chain promotes the formation of specific interactions with the helix 35 of 23S rRNA.

scholarly journals Variations in O-Antigen Biosynthesis and O-Acetylation Associated with Altered Phage Sensitivity in Escherichia coli 4s

2014 ◽  
Vol 197 (5) ◽  
pp. 905-912 ◽  
Author(s):  
Yuriy A. Knirel ◽  
Nikolai S. Prokhorov ◽  
Alexander S. Shashkov ◽  
Olga G. Ovchinnikova ◽  
Evelina L. Zdorovenko ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Host Cells ◽  
Side Chain ◽  
Gram Negative Bacteria ◽  
Gene Encoding ◽  
Content Type ◽  
Surface Receptors ◽  
E Coli ◽  
O Antigen ◽  
Phage Sensitivity

The O polysaccharide of the lipopolysaccharide (O antigen) of Gram-negative bacteria often serves as a receptor for bacteriophages that can make the phage dependent on a given O-antigen type, thus supporting the concept of the adaptive significance of the O-antigen variability in bacteria. The O-antigen layer also modulates interactions of many bacteriophages with their hosts, limiting the access of the viruses to other cell surface receptors. Here we report variations of O-antigen synthesis and structure in an environmentalEscherichia coliisolate, 4s, obtained from horse feces, and its mutants selected for resistance to bacteriophage G7C, isolated from the same fecal sample. The 4s O antigen was found to be serologically, structurally, and genetically related to the O antigen ofE. coliO22, differing only in side-chain α-d-glucosylation in the former, mediated by agtrlocus on the chromosome. Spontaneous mutations ofE. coli4s occurring with an unusually high frequency affected either O-antigen synthesis or O-acetylation due to the inactivation of the gene encoding the putative glycosyltransferase WclH or the putative acetyltransferase WclK, respectively, by the insertion of IS1-like elements. These mutations induced resistance to bacteriophage G7C and also modified interactions ofE. coli4s with several other bacteriophages conferring either resistance or sensitivity to the host. These findings suggest that O-antigen synthesis and O-acetylation can both ensure the specific recognition of the O-antigen receptor following infection by some phages and provide protection of the host cells against attack by other phages.

scholarly journals DNA Microarray Analyses of the Long-Term Adaptive Response of Escherichia coli to Acetate and Propionate

2003 ◽  
Vol 69 (3) ◽  
pp. 1759-1774 ◽  
Author(s):  
T. Polen ◽  
D. Rittmann ◽  
V. F. Wendisch ◽  
H. Sahm
Keyword(s):  
Escherichia Coli ◽  
Dna Microarray ◽  
Adaptive Response ◽  
Carbon Sources ◽  
Short Chain Fatty Acids ◽  
Adaptive Responses ◽  
General Stress Response ◽  
E Coli ◽  
Short Term Exposure

ABSTRACT In its natural environment, Escherichia coli is exposed to short-chain fatty acids, such as acetic acid or propionic acid, which can be utilized as carbon sources but which inhibit growth at higher concentrations. DNA microarray experiments revealed expression changes during exponential growth on complex medium due to the presence of sodium acetate or sodium propionate at a neutral external pH. The adaptive responses to acetate and propionate were similar and involved genes in three categories. First, the RNA levels for chemotaxis and flagellum genes increased. Accordingly, the expression of chromosomal fliC′-′lacZ and flhDC′-′lacZ fusions and swimming motility increased after adaptation to acetate or propionate. Second, the expression of many genes that are involved in the uptake and utilization of carbon sources decreased, indicating some kind of catabolite repression by acetate and propionate. Third, the expression of some genes of the general stress response increased, but the increases were more pronounced after short-term exposure for this response than for the adaptive response. Adaptation to propionate but not to acetate involved increased expression of threonine and isoleucine biosynthetic genes. The gene expression changes after adaptation to acetate or propionate were not caused solely by uncoupling or osmotic effects but represented specific characteristics of the long-term response of E. coli to either compound.

scholarly journals Cyclic AMP (cAMP) and cAMP Receptor Protein Influence both Synthesis and Uptake of Extracellular Autoinducer 2 in Escherichia coli

2005 ◽  
Vol 187 (6) ◽  
pp. 2066-2076 ◽  
Author(s):  
Liang Wang ◽  
Yoshifumi Hashimoto ◽  
Chen-Yu Tsao ◽  
James J. Valdes ◽  
William E. Bentley
Keyword(s):  
Escherichia Coli ◽  
Cyclic Amp ◽  
Carbon Sources ◽  
Cell Communication ◽  
Receptor Protein ◽  
Upstream Region ◽  
E Coli ◽  
Autoinducer 2 ◽  
Serovar Typhimurium ◽  
Camp Receptor

ABSTRACT Bacterial autoinducer 2 (AI-2) is proposed to be an interspecies mediator of cell-cell communication that enables cells to operate at the multicellular level. Many environmental stimuli have been shown to affect the extracellular AI-2 levels, carbon sources being among the most important. In this report, we show that both AI-2 synthesis and uptake in Escherichia coli are subject to catabolite repression through the cyclic AMP (cAMP)-CRP complex, which directly stimulates transcription of the lsr (for “luxS regulated”) operon and indirectly represses luxS expression. Specifically, cAMP-CRP is shown to bind to a CRP binding site located in the upstream region of the lsr promoter and works with the LsrR repressor to regulate AI-2 uptake. The functions of the lsr operon and its regulators, LsrR and LsrK, previously reported in Salmonella enterica serovar Typhimurium, are confirmed here for E. coli. The elucidation of cAMP-CRP involvement in E. coli autoinduction impacts many areas, including the growth of E. coli in fermentation processes.

scholarly journals Comparison of methods for the identification and sub-typing of O157 and non-O157 Escherichia coli serotypes and their integration into a polyphasic taxonomy approach

2016 ◽  
Vol 55 (2) ◽  
pp. 81-90 ◽  
Author(s):  
M.A. Prieto-Calvo ◽  
M.K. Omer ◽  
O. Alvseike ◽  
M. López ◽  
A. Alvarez-Ordóñez ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Carbon Source ◽  
Carbon Sources ◽  
23S Rrna ◽  
Local Alignment ◽  
Taxonomic Structure ◽  
Sequencing Data ◽  
E Coli ◽  
Additional Information ◽  
Ft Ir

AbstractPhenotypic, chemotaxonomic and genotypic data from 12 strains ofEscherichia coli werecollected, including carbon source utilisation profiles, ribotypes, sequencing data of the 16S–23S rRNA internal transcribed region (ITS) and Fourier transform-infrared (FT-IR) spectroscopic profiles. The objectives were to compare several identification systems forE. coliand to develop and test a polyphasic taxonomic approach using the four methodologies combined for the sub-typing of O157 and non-O157E. coli. The nucleotide sequences of the 16S–23S rRNA ITS regions were amplified by polymerase chain reaction (PCR), sequenced and compared with reference data available at the GenBank database using the Basic Local Alignment Search Tool (BLAST) . Additional information comprising the utilisation of carbon sources, riboprint profiles and FT-IR spectra was also collected. The capacity of the methods for the identification and typing ofE. colito species and subspecies levels was evaluated. Data were transformed and integrated to present polyphasic hierarchical clusters and relationships. The study reports the use of an integrated scheme comprising phenotypic, chemotaxonomic and genotypic information (carbon source profile, sequencing of the 16S–23S rRNA ITS, ribotyping and FT-IR spectroscopy) for a more precise characterisation and identification ofE. coli. The results showed that identification ofE. colistrains by each individual method was limited mainly by the extension and quality of reference databases. On the contrary, the polyphasic approach, whereby heterogeneous taxonomic data were combined and weighted, improved the identification results, gave more consistency to the final clustering and provided additional information on the taxonomic structure and phenotypic behaviour of strains, as shown by the close clustering of strains with similar stress resistance patterns.

Synthesis, Characterization and Evaluation of Antimicrobial Properties of (R)-(-)-4-Phenyl-2 Oxazolidinone Based Azetidinones

2018 ◽  
Vol 16 (2) ◽  
pp. 104-113 ◽  
Author(s):  
Shyamal Baruah ◽  
Amrit Puzari ◽  
Farhana Sultana ◽  
Jayanta Barman
Keyword(s):  
Escherichia Coli ◽  
Antibacterial Activity ◽  
Acetic Acid ◽  
Bacillus Subtilis ◽  
Antibacterial Property ◽  
Antimicrobial Properties ◽  
Side Chain ◽  
Alkyl Side Chain ◽  
E Coli ◽  
Aromatic Imines

Introduction: A series of (R)-(-)-4-Phenyl-2 oxazolidinone based azetidinones (4a-i) were synthesized from the reaction of (2-Oxo-4-phenyl-oxazolidin-3-yl) acetic acid with aromatic imines (3a-i) in the presence of Thionyl chloride and Triethylamine as a base. Methods: The transformation proceeds through the formation of acid chloride to ketene which finally forms the azetidinones through [2+2] cycloaddition with aromatic imines. Products obtained were screened to evaluate their antibacterial activity with respect to known bacteria like Escherichia Coli (E. Coli) and Bacillus subtilis. Results and Conclusion: In most of the cases, azetidinones were found to exhibit superior antimicrobial properties than oxazolidinones. They were found to be a good inhibitor of gram-positive and gramnegative bacteria. Enhancement of antibacterial property can be attributed to the presence of azetidinone ring and hydrophobic alkyl side chain in the scaffolds.

scholarly journals Production in Escherichia coli of Poly(3-Hydroxybutyrate-co-3-Hydroxyvalerate) with Differing Monomer Compositions from Unrelated Carbon Sources

2011 ◽  
Vol 77 (14) ◽  
pp. 4886-4893 ◽  
Author(s):  
Quan Chen ◽  
Qian Wang ◽  
Guoqing Wei ◽  
Quanfeng Liang ◽  
Qingsheng Qi
Keyword(s):  
Escherichia Coli ◽  
Control Strategy ◽  
Biosynthetic Pathway ◽  
Feedback Inhibition ◽  
Carbon Sources ◽  
Threonine Deaminase ◽  
Content Type ◽  
E Coli ◽  
Key Factor ◽  
Threonine Biosynthesis

ABSTRACTThe industrial production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) has been hindered by high cost and a complex control strategy caused by the addition of propionate. In this study, based on analysis of the PHBV biosynthesis process, we developed a PHBV biosynthetic pathway from a single unrelated carbon source via threonine biosynthesis inEscherichia coli. To accomplish this, we (i) overexpressed threonine deaminase, which is the key factor for providing propionyl-coenzyme A (propionyl-CoA), from different host bacteria, (ii) removed the feedback inhibition of threonine by mutating and overexpressing thethrABCoperon inE. coli, and (iii) knocked out the competitive pathways of catalytic conversion of propionyl-CoA to 3-hydroxyvaleryl-CoA. Finally, we constructed a series of strains and mutants which were able to produce the PHBV copolymer with differing monomer compositions in a modified M9 medium supplemented with 20 g/liter xylose. The largest 3-hydroxyvalerate fraction obtained in the copolymer was 17.5 mol%.

Structures and reaction mechanisms of riboflavin synthases of eubacterial and archaeal origin

2005 ◽  
Vol 33 (4) ◽  
pp. 780-784 ◽  
Author(s):  
M. Fischer ◽  
W. Römisch ◽  
B. Illarionov ◽  
W. Eisenreich ◽  
A. Bacher
Keyword(s):  
Escherichia Coli ◽  
Reaction Mechanisms ◽  
Nmr Spectra ◽  
Common Ancestor ◽  
Sequence Similarity ◽  
Side Chain ◽  
Cd Spectra ◽  
Riboflavin Biosynthesis ◽  
Significant Sequence Similarity ◽  
E Coli

The biosynthesis of one riboflavin molecule requires one molecule of GTP and two molecules of ribulose 5-phosphate as substrates. GTP is hydrolytically opened, converted into 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione by a sequence of deamination, side chain reduction and dephosphorylation. Condensation with 3,4-dihydroxy-2-butanone 4-phosphate obtained from ribulose 5-phosphate leads to 6,7-dimethyl-8-ribityllumazine. The dismutation of 6,7-dimethyl-8-ribityllumazine catalysed by riboflavin synthase produces riboflavin and 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione. A pentacyclic adduct of two 6,7-dimethyl-8-ribityllumazines has been identified earlier as a catalytically competent reaction intermediate of the Escherichia coli enzyme. Acid quenching of reaction mixtures of riboflavin synthase of Methanococcus jannaschii, devoid of similarity to riboflavin synthases of eubacteria and eukaryotes, afforded a compound whose optical absorption and NMR spectra resemble that of the pentacyclic E. coli riboflavin synthase intermediate, whereas the CD spectra of the two compounds have similar envelopes but opposite signs. Each of the compounds could serve as a catalytically competent intermediate for the enzyme by which it was produced, but not vice versa. All available data indicate that the respective pentacyclic intermediates of the M. jannaschii and E. coli enzymes are diastereomers. Whereas the riboflavin synthase of M. jannaschii is devoid of similarity with those of eubacteria and eukaryotes, it has significant sequence similarity with 6,7-dimethyl-8-ribityllumazine synthases catalysing the penultimate step of riboflavin biosynthesis. 6,7-Dimethyl-8-ribityllumazine synthase and the archaeal riboflavin synthase appear to have diverged early in the evolution of Archaea from a common ancestor.

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