scholarly journals Ubiquinone biosynthesis in Escherichia coli K-12. Accumulation of an octaprenol, farnesylfarnesylgeraniol, by a multiple aromatic auxotroph

1971 ◽  
Vol 123 (3) ◽  
pp. 435-443 ◽  
Author(s):  
J. A. Hamilton ◽  
G. B. Cox
Keyword(s):  
Escherichia Coli ◽  
Mass Spectroscopy ◽  
Active Form ◽  
Side Chain ◽  
Whole Cells ◽  
Cell Extracts ◽  
K 12

Cell extracts of a multiple aromatic auxotroph of Escherichia coli K-12, strain AB2830, grown in the absence of precursors of the quinone rings of the ubiquinone and menaquinone molecules, converted 4-hydroxy[U-14C]benzoate into a mixture of 3-octaprenyl-4-hydroxybenzoate and 2-octaprenylphenol. An octaprenol, farnesylfarnesylgeraniol, was isolated from such cell extracts and characterized by n.m.r. and mass spectroscopy. Neither the octaprenol, nor polyprenylation of 4-hydroxy[U-14C]benzoate, could be detected in cell extracts of strain AB2830 grown in the presence of 0.1mm-4-hydroxybenzoate. It was concluded that, in the biosynthesis of ubiquinone, the polyprenyl side chain is added to 4-hydroxybenzoate as a C40 unit, the active form of which is converted by cell extracts into farnesylfarnesylgeraniol. The multiple aromatic auxotroph, when grown in the absence of 4-hydroxybenzoate but in the presence of 4-aminobenzoate, converted the latter compound into 3-octaprenyl-4-aminobenzoate. This compound was isolated from whole cells and characterized by n.m.r. and mass spectroscopy.

scholarly journals Purification and Characterization of the Recombinant Thermus sp. Strain T2 α-Galactosidase Expressed in Escherichia coli

2001 ◽  
Vol 67 (4) ◽  
pp. 1601-1606 ◽  
Author(s):  
Mitsunori Ishiguro ◽  
Satoshi Kaneko ◽  
Atsushi Kuno ◽  
Yoshinori Koyama ◽  
Shigeki Yoshida ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Active Form ◽  
Gel Filtration ◽  
Amino Acid Sequences ◽  
Side Chain ◽  
Catalytic Function ◽  
Native Polyacrylamide Gel Electrophoresis

ABSTRACT The nucleotide sequence of the Thermus sp. strain T2 DNA coding for a thermostable α-galactosidase was determined. The deduced amino acid sequence of the enzyme predicts a polypeptide of 474 amino acids (M r, 53,514). The observed homology between the deduced amino acid sequences of the enzyme and α-galactosidase from Thermus brockianus was over 70%.Thermus sp. strain T2 α-galactosidase was expressed in its active form in Escherichia coli and purified. Native polyacrylamide gel electrophoresis and gel filtration chromatography data suggest that the enzyme is octameric. The enzyme was most active at 75°C forp-nitrophenyl-α-d-galactopyranoside hydrolysis, and it retained 50% of its initial activity after 1 h of incubation at 70°C. The enzyme was extremely stable over a broad range of pH (pH 6 to 13) after treatment at 40°C for 1 h. The enzyme acted on the terminal α-galactosyl residue, not on the side chain residue, of the galactomanno-oligosaccharides as well as those of yeasts and Mortierella vinacea α-galactosidase I. The enzyme has only one Cys residue in the molecule.para-Chloromercuribenzoic acid completely inhibited the enzyme but did not affect the mutant enzyme which contained Ala instead of Cys, indicating that this Cys residue is not responsible for its catalytic function.

scholarly journals Aerobic Growth of Escherichia coli with 2,4,6-Trinitrotoluene (TNT) as the Sole Nitrogen Source and Evidence of TNT Denitration by Whole Cells and Cell-Free Extracts

2006 ◽  
Vol 72 (12) ◽  
pp. 7945-7948 ◽  
Author(s):  
Ben Stenuit ◽  
Laurent Eyers ◽  
Raoul Rozenberg ◽  
Jean-Louis Habib-Jiwan ◽  
Spiros N. Agathos
Keyword(s):  
Escherichia Coli ◽  
Nitrogen Source ◽  
Cell Suspensions ◽  
Sole Nitrogen Source ◽  
Aerobic Growth ◽  
Whole Cells ◽  
Cell Extracts

ABSTRACT Escherichia coli grew aerobically with 2,4,6-trinitrotoluene (TNT) as sole nitrogen source and caused TNT's partial denitration. This reaction was enhanced in nongrowing cell suspensions with 0.516 mol nitrite released per mol TNT. Cell extracts denitrated TNT in the presence of NAD(P)H. Isomers of amino-dimethyl-tetranitrobiphenyl were detected and confirmed with U-15N-labeled TNT.

THE INHIBITION OF KETO ACID OXIDATION BY PYOCYANINE

1957 ◽  
Vol 3 (2) ◽  
pp. 313-318 ◽  
Author(s):  
J. J. R. Campbell ◽  
A. M. MacQuillan ◽  
B. A. Eagles ◽  
R. A. Smith
Keyword(s):  
Escherichia Coli ◽  
Cell Membrane ◽  
Pseudomonas Fluorescens ◽  
Acid Level ◽  
Divalent Cations ◽  
Acid Oxidation ◽  
Keto Acid ◽  
Proteus Vulgaris ◽  
Whole Cells ◽  
Cell Extracts

When tested against Pseudomonas fluorescens, pyocyanine was found to stop the oxidation of a number of substrates at the keto acid level. This inhibition could be reversed by the addition of divalent cations. Of these, magnesium was most effective. The pigment was found to be similarly effective against the oxidations of Proteus vulgaris. Whole cells of Escherichia coli were not affected by the dye, whereas cell extracts were, indicating that the dye did not penetrate the cell membrane.

scholarly journals Identification of an Escherichia coli Operon Required for Formation of the O-Antigen Capsule

2005 ◽  
Vol 187 (15) ◽  
pp. 5259-5266 ◽  
Author(s):  
Adi Peleg ◽  
Yulia Shifrin ◽  
Ophir Ilan ◽  
Chen Nadler-Yona ◽  
Shani Nov ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Secretion System ◽  
Side Chain ◽  
E Coli ◽  
O Antigen ◽  
Group 4 ◽  
K 12 ◽  
Polysaccharide Capsules ◽  
Polysaccharide Secretion ◽  
Group 1

ABSTRACT Escherichia coli produces polysaccharide capsules that, based on their mechanisms of synthesis and assembly, have been classified into four groups. The group 4 capsule (G4C) polysaccharide is frequently identical to that of the cognate lipopolysaccharide O side chain and has, therefore, also been termed the O-antigen capsule. The genes involved in the assembly of the group 1, 2, and 3 capsules have been described, but those required for G4C assembly remained obscure. We found that enteropathogenic E. coli (EPEC) produces G4C, and we identified an operon containing seven genes, ymcD, ymcC, ymcB, ymcA, yccZ, etp, and etk, which are required for formation of the capsule. The encoded proteins appear to constitute a polysaccharide secretion system. The G4C operon is absent from the genomes of enteroaggregative E. coli and uropathogenic E. coli. E. coli K-12 contains the G4C operon but does not express it, because of the presence of IS1 at its promoter region. In contrast, EPEC, enterohemorrhagic E. coli, and Shigella species possess an intact G4C operon.

scholarly journals Effects of dicyclohexylcarbodi-imide on proton translocation coupled to fumarate reduction in anaerobically grown cells of Escherichia coli K-12

1976 ◽  
Vol 160 (3) ◽  
pp. 813-816 ◽  
Author(s):  
S J Gutowski ◽  
H Rosenberg
Keyword(s):  
Escherichia Coli ◽  
Reductase Activity ◽  
Proton Translocation ◽  
Fumarate Reductase ◽  
Escherichia Coli K12 ◽  
Fumarate Reduction ◽  
Cell Extracts ◽  
Endogenous Substrates ◽  
K 12

The addition of dicyclohexylcarbodi-imide to anaerobic cells of Escherichia coli K12 decreases both the observed extent of proton translocation coupled to fumarate reduction by endogenous substrates and the t1/2 of proton re-entry after such translocation, but does not affect fumarate uptake. Dicyclohexylcarbodi-imide also inhibits fumarate reductase activity in cell extracts.

Metal binding by the peptidoglycan sacculus of Escherichia coli K-12

1984 ◽  
Vol 30 (2) ◽  
pp. 204-211 ◽  
Author(s):  
B. D. Hoyle ◽  
T. J. Beveridge
Keyword(s):  
Escherichia Coli ◽  
Aqueous Solution ◽  
Bacillus Subtilis ◽  
Metal Binding ◽  
Sodium Dodecyl ◽  
Dry Weight ◽  
Metallic Ions ◽  
Trypsin Treatment ◽  
Whole Cells ◽  
K 12

The peptidoglycan of Escherichia coli K-12 strain AB264 was isolated by treating whole cells with sodium dodecyl sulfate and was purified by deoxyribonuclease, ribonuclease, and trypsin treatment. Like the peptidoglycan of Bacillus subtilis, this peptidoglycan proved able to bind substantial amounts of metallic ions from aqueous solution. In particular, most metals of the transition I series were bound from solution in amounts ≥ 1 μmol/mg dry weight peptidoglycan.

scholarly journals NEGATIVE DOMINANT MUTATIONS OF THE uidR GENE IN ESCHERICHIA COLI: GENETIC PROOF FOR A COOPERATIVE REGULATION OF uidA EXPRESSION

Genetics ◽  
1986 ◽  
Vol 112 (2) ◽  
pp. 173-182
Author(s):  
Carlos Blanco ◽  
Paul Ritzenthaler ◽  
Mireille Mata-Gilsinger
Keyword(s):  
Escherichia Coli ◽  
Gene Dosage ◽  
Regulatory Gene ◽  
Active Form ◽  
Point Mutations ◽  
Multicopy Plasmid ◽  
Wild Type ◽  
Cooperative Process ◽  
Mutant Phenotypes ◽  
K 12

ABSTRACT The uidA gene is the first gene involved in the hexuronide-hexuronate pathway in Escherichia coli K-12 and is under the dual control of the uidR and uxuR encoded repressors. Point mutations affecting the uidR regulatory gene were sought to investigate the regulation of uidA. When the uidR mutant allele was on a multicopy plasmid and the wild-type allele was on the chromosome, some of the mutant phenotypes were dominant to the wild-type phenotype, indicating that the active form of the UidR repressor is multimeric. We have demonstrated that expression of the mutant phenotype is dependent on gene dosage. The dominance of the uidR allele was also sensitive to the presence of the wild-type uxuR allele in the cell. This behavior probably results from UidR-UxuR repressor interactions. A mechanism is proposed: we suggest that the UidR and UxuR repressors interact after their binding to the operator site of uidA; the binding of one regulatory molecule may facilitate the binding of the other one in a cooperative process.

scholarly journals Sequence Analysis of the GntII (Subsidiary) System for Gluconate Metabolism Reveals a Novel Pathway for l-Idonic Acid Catabolism in Escherichia coli

1998 ◽  
Vol 180 (14) ◽  
pp. 3704-3710 ◽  
Author(s):  
Christoph Bausch ◽  
Norbert Peekhaus ◽  
Cristina Utz ◽  
Tessa Blais ◽  
Elizabeth Murray ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Sequence Analysis ◽  
Genomic Sequence ◽  
Regulatory Protein ◽  
Gene Encoding ◽  
E Coli ◽  
Cell Extracts ◽  
Genomic Sequence Analysis ◽  
Metabolic Sequence ◽  
K 12

ABSTRACT The presence of two systems in Escherichia coli for gluconate transport and phosphorylation is puzzling. The main system, GntI, is well characterized, while the subsidiary system, GntII, is poorly understood. Genomic sequence analysis of the region known to contain genes of the GntII system led to a hypothesis which was tested biochemically and confirmed: the GntII system encodes a pathway for catabolism of l-idonic acid in whichd-gluconate is an intermediate. The genes have been named accordingly: the idnK gene, encoding a thermosensitive gluconate kinase, is monocistronic and transcribed divergently from the idnD-idnO-idnT-idnRoperon, which encodes l-idonate 5-dehydrogenase, 5-keto-d-gluconate 5-reductase, an l-idonate transporter, and an l-idonate regulatory protein, respectively. The metabolic sequence is as follows: IdnT allows uptake of l-idonate; IdnD catalyzes a reversible oxidation ofl-idonate to form 5-ketogluconate; IdnO catalyzes a reversible reduction of 5-ketogluconate to formd-gluconate; IdnK catalyzes an ATP-dependent phosphorylation of d-gluconate to form 6-phosphogluconate, which is metabolized further via the Entner-Doudoroff pathway; and IdnR appears to act as a positive regulator of the IdnR regulon, withl-idonate or 5-ketogluconate serving as the true inducer of the pathway. The l-idonate 5-dehydrogenase and 5-keto-d-gluconate 5-reductase reactions were characterized both chemically and biochemically by using crude cell extracts, and it was firmly established that these two enzymes allow for the redox-coupled interconversion of l-idonate andd-gluconate via the intermediate 5-ketogluconate. E. coli K-12 strains are able to utilize l-idonate as the sole carbon and energy source, and as predicted, the ability ofidnD, idnK, idnR, andedd mutants to grow on l-idonate is altered.

scholarly journals Assembly of Cyclic Enterobacterial Common Antigen in Escherichia coli K-12

2005 ◽  
Vol 187 (20) ◽  
pp. 6917-6927 ◽  
Author(s):  
Junko Kajimura ◽  
Arifur Rahman ◽  
Paul D. Rick
Keyword(s):  
Escherichia Coli ◽  
Subcellular Location ◽  
Dry Weight ◽  
Water Soluble ◽  
Cell Fractionation ◽  
Common Antigen ◽  
Enterobacterial Common Antigen ◽  
Repeat Units ◽  
Cell Extracts ◽  
K 12

ABSTRACT We describe here the purification and quantification of a water-soluble cyclic form of enterobacterial common antigen (ECACYC) from Escherichia coli K-12 as well as information regarding its subcellular location and the genetic loci involved in its assembly. Structural characterization of purified ECACYC molecules obtained from E. coli K-12 revealed that they uniformly contained four trisaccharide repeat units, and they were substituted with from zero to four O-acetyl groups. Cells from overnight cultures contained approximately 2 μg ECACYC per milligram (dry weight), and cell fractionation studies revealed that these molecules were localized exclusively in the periplasm. The synthesis and assembly of ECACYC were found to require the wzxE and wzyE genes of the wec gene cluster. These genes encode proteins involved in the transmembrane translocation of undecaprenylpyrophosphate-linked ECA trisaccharide repeat units and the polymerization of trisaccharide repeat units, respectively. Surprisingly, synthesis of ECACYC was dependent on the wzzE gene, which is required for the modulation of the polysaccharide chain lengths of phosphoglyceride-linked ECA (ECAPG). The presence of ECACYC in extracts of several other gram-negative enteric organisms was also demonstrated; however, it was not detected in cell extracts of Pseudomonas aeruginosa. These data suggest that in addition to ECAPG, ECACYC may be synthesized in many, if not all, members of the Enterobacteriaceae.

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