bluuebunny/crossref_metadata_2025_split huggingface dataset:
The mdoA locus of Escherichia coli consists of an operon under osmotic control
Authors: Lacroix J.‐M. , Loubens I. , Tempête M. , Menichi B. , Bohin J.‐P. 
Published: October 2006

SummaryIn Escherichia coli, the 5 kb mdoA locus is involved in the osmotically controlled biosynthesis of periplasmic membrane‐derived oligosaccharides (MDOs). The structure of this locus was analysed by in vitro cassette insertion, transposon mutagenesis, and gene‐fusion analysis.A ‘neo’ cassette, derived from the neomycin phosphotransferase II region of transposon Tn5, was inserted into mdoA, borne by a multicopy plasmid. This plasmid was shown to complement two previously described mdoA mutations, depending on the orientation of the exogenous gene. Thus, the gene altered by these mutations could be expressed under the control of the exogenous promoter. Moreover, the ‘neo’ cassette inactivated another, uncharacterlzed, mdo gene, because when this insertion was transferred into the chromosome MDO synthesis was abolished.The existence of a second gene was confirmed by complementation analysis with a collection of Tn1000 insertions into mdoA. Two groups were defined, and the two genes are organized into an operon (mdoGH). This conclusion was reached because Tn1000 insertions in the first gene displayed a polar effect on the expression of the second gene.An active gene fusion was obtained on a multicopy plasmid between the beginning of mdoH and lacZ. The hybrid β‐galactosidase activity followed the same osmotically controlled response as that described for of MDO synthesis. This regulation was unaffected by the presence, or absence, of MDOs In the periplasm. Finally, the amount of mdoA‐specific mRNAs, determined by dot blot hybridization, decreased when the osmolarity of the growth medium increased.
Location of the RNA‐processing enzymes RNase III, RNase E and RNase P in the Eschenchia coli cell
Authors: Miczak A. , Srivastava R. A. K. , Apirion O. 
Published: October 2006

SummaryCells overexpressing the RNA‐processing enzymes RNase III, RNase E and RNase P were fractionated into membrane and cytoplasm. The RNA‐processing enzymes were associated with the membrane fraction. The membrane was further separated to inner and outer membrane and the three RNA‐processing enzymes were found in the inner membrane fraction. By assaying for these enzymatic activities we showed that even in a normal wild‐type strain of Escherichia coli these enzymes fractionate primarily with the membrane. The RNA part of RNase P is found in the cytosolic fraction of cells overexpressing this RNA, while the overexpressed RNase P protein sediments with the membrane fraction; this suggests that the RNase P protein anchors the RNA catalytic moiety of the enzyme to a larger entity. The implications of these findings for the cellular organization of the RNA‐processing enzymes in the cell are discussed.
Regulation of sod genes in Escherichia coli: relevance to superoxide dismutase function
Authors: Fee J. A. 
Published: October 2006

SummaryThis review is concerned with the effects of environmental perturbations on the expression of the two superoxide dismutase (SOD) genes in Escherichia coli (sodA, MnSOD; sodB, FeSOD). Early studies using SOD activity, showed that MnSOD levels respond to changes in oxygen tension, type of substrate, redox active compounds, iron concentration, the nature of the terminal oxidant, and the redox potential of the medium. FeSOD levels appeared nominally insensitive to these perturbations. More recent molecular genetic studies revealed that sodA expression is subject to regulation by three major regulatory systems: fur (ferric uptake regulation) and arcA arcB (aerobic respiratory control) mediate repression of sodA, while a relatively new system, soxR soxS (superoxide response), mediates activation of sodA expression. By contrast, sodB expression, which is much less studied at this time, appears to be positively activated in trans by fur. A rudimentary gene regulation model is presented which rationalizes past observations, is experimentally testable, and should serve as a guide to future research in this area.
Activation of cholera toxin and Escherichia coli heat‐labile enterotoxins by ADP‐ribosylation factors, a family of 20 kDa guanine nucleotide‐binding proteins
Authors: Moss J. , Vaughan M. 
Published: October 2006

SummaryCholera toxin and Escherichia coli heat‐labile enterotoxins are responsible, in part, for the symptomatology of cholera and traveller's diarrhoea, respectively. Effects of the toxins result from ADP‐ribosylation of regulatory guanine nucleotide‐binding (G) proteins; the ADP‐ribosylated G protein is stabilized in an activated state, resulting in prolonged effects on its target. Toxin‐catalysed ADP‐ribosylation is stimulated in vitro by a family of guanine nucleotide‐binding proteins, c. 20 kDa, termed ADP‐ribosylation factors or ARFs. In the presence of GTP, but not GDP or adenine analogues, ARFs serve as allosteric activators of the toxin. The effects are amplified by certain phospholipids and detergents which promote guanine nucleotide binding. Six different mammalian ARF genes have been identified. They encode highly conserved, ubiquitous proteins of 175 to 181 amino acids, containing consensus domains responsible for guanine nucleotide binding. Differences in amino acid sequences are localized near the amino terminus and in the carboxy half of the protein. Although the physiological functions of ARFs have not been precisely defined, their immunological localization to the Golgi is consistent with a role in the regulated orderly movement of newly synthesized proteins from the endoplasmic reticulum, through the Golgi system to their ultimate destination.
Escherichia coli molecular genetic map (1500 kbp): update II
Authors: Médigue C. , Viari A. , Hénaut A. , Danchin A. 
Published: October 2006

SummaryThe DNA sequence data for Escherichia coli deposited in the EMBL library (release 27), together with miscellaneous data obtained from several laboratories, have been localized on an updated and corrected version of the restriction map of the chromosome generated by Kohara et al. (1987) and modified by others. This second update adds a further 500 kbp, increasing the amount of the E. coli chromosome sequenced to about one third of the total: 1510 kbp of sequenced DNA is included in the present data base. The accuracy of the map is assessed, and allows us to propose a precise genetic map position for every sequenced gene. The location of rare‐cutting sites such as AvII, NotI and SfiI have also been included in the update in order to combine the data obtained from different sources into one single file. The distribution of palindromic sequences (to which most restriction sites belong) has been studied in coding sequences. There appears to be a significant counter‐selection against several such sequences in E. coli coding sequences (but not in other organisms such as Saccharomyces cerevisiae), suggesting the existence of constraints on DNA structure in E. coli, perhaps indicative of a functional role for horizontal gene transfer, preserving coding sequences, in this type of bacteria.
Export of infectious particles by Escherichia coli transfected with the RF DNA of Pf1, a virus of Pseudomonas aeruginosa strain K
Authors: Kostrikis L. G. , Reisberg S. A. , Simon M. N. , Wall J. S. , Day L. A. 
Published: October 2006

SummaryPf1 is a filamentous, single‐stranded DNA virus that has Pseudomas aeruginosa (strain K) as host. It is the longest of the filamentous bacterial viruses, and the DNA within it has the most extended conformation known. Pfl virus cannot infect Escherichia coli (strain MM294) cells, but when these cells are transfected with the double‐stranded replicative form of Pt1 DNA (RF DNA, 7.35 kb), they export low levels of infectious particles that create plaques on lawns of P. aeruginosa. Several different structural species, at least two of which are infectious, are exported. One of them, called Epf1, has virtually the same structure as Pf1, but the amount of Epf1 exported by E. coli is 104 lower than the amount of Pf1 exported by P. aeruginosa. The results imply that host factors affect not only the efficiency of virus assembly and export, but also the actual structures of the species exported.
An amplifiable and deletable chloramphenicol‐resistance determinant of Streptomyces lividans 1326 encodes a putative transmembrane protein
Authors: Dittrich W. , Betzler M. , Schrempf H. 
Published: October 2006

SummaryA genetically unstable chloramphenicol resistance gene from Streptomyces lividans 1326 was cloned and characterized. This gene and adjacent DNA regions can be lost spontaneously or amplify within variants. Biochemical studies proved that chloramphenicol is not modified by an acetyltransferase or any other enzyme and that ribosomes of the resistant strain are sensitive to chloramphenicol. Sequence data revealed that the resistance gene encodes a hydrophobic protein predicted to have 12 membrane‐spanning α‐helices and a hydropathic profile similar to the membrane of proteins required for the efflux of tetracydine. Variable proportions of the amino acids (about 16‐24%) within the presumed chloramphenicol‐resistant protein are identical to various aligned tetracycline‐resistant proteins from Gram‐negative and Gram‐positive bacteria and to transporters for citrate in Klebsiella pneumonaie and for ferrichrome in Escherichia coli.
Mechanism of regulation of the formate‐hydrogenlyase pathway by oxygen, nitrate, and pH: definition of the formate regulon
Authors: Rossmann R. , Sawers G. , Böck A. 
Published: October 2006

SummaryThe products of a minimum of 15 genes are required for the synthesis of an active formate‐hydrogenlyase (FHL) system in Escherichia coli. All are co‐ordinately regulated in response to variations in the oxygen and nitrate concentration and the pH of the culture medium. Formate is obligately required for transcriptional activation of these genes. Analysis of the transcription of one of these genes, hycB linked to the iacZ reporter gene, revealed that oxygen and nitrate repression of transcription could be relieved completely, or partially in the case of nitrate, either by the addition of formate to the medium or by increasing the copy number of the gene encoding the transcriptional activator (fhlA) of this regulon. These studies uncovered a further level of regulation in which the transcription of hycB was reduced in cells grown on glucose. This effect was most clearly seen in aerobically grown cells when formate was added externally. Addition of cAMP overcame this glucose repression, which could be shown to be mediated by the cAMP receptor protein. These results would be consistent with the transport of formate being regulated by catabolite repression. Moreover, the repression of transcription through high pH also could be partially overcome by addition of increasing concentrations of formate to the medium, again being consistent with regulation at the level of formate import and export. Taken together, all these observations indicate that it is the intracellular level of formate that determines the transcription of the genes of the formate regulon by FhlA. This represents a novel positive feedback mechanism in which the activator of a regulon induces its own synthesis in response to increases in the concentration of the catabolic substrate, and this in turn is governed by the relative affinities of FhlA and the three formate dehydrogenase isoenzymes for formate.
Pseudomonas aeruginosa elastase and elastolysis revisited: recent developments
Authors: Galloway D. R. 
Published: October 2006

SummaryWith the determination of the three‐dimensional structure of elastase and the probable identification of the active site and key residues involved in proteolytic activity, our knowledge of the molecular details of this interesting protease is rapidly increasing. Pseudomonas elastase appears to be remarkably similar to the Bacillus metalloproteinase thermolysin. A further significant development has been the discovery of the lasA gene and the fact that Pseudomonas elastase and alkaline proteinase appear to act in concert with the LasA protein to display the notable elastolytic activity exhibited by isolates of this organism. Biochemical and genetic studies indicate that LasA is a second elastase which may be an important virulence factor that has been overlooked in previous studies.
Sensory transduction in the gliding bacterium Myxococcus xanthus
Authors: Zusman D. R. , McBride M. J. 
Published: October 2006

SummarySensory transduction in the gliding bacterium Myxococcus xanthus is mediated by the frz genes. These genes are homologous to the chemotaxis genes of enteric bacteria and control the rate of cell reversal during gliding. Sensory transduction is hypothesized to involve the recognition of substances present in the medium at the cell surface and the subsequent stimulation of a cytoplasmic methyl‐accepting protein, FrzCD. Phosphorylation of FrzE is also involved in the sensory transduction pathway. Despite the similarities between the chemotaxis proteins of enteric bacteria and M. xanthus Frz proteins, fundamental differences exist between these different bacteria in terms of the ability of cells to recognize and respond to substances in their environment. The mechanism of directional switching and the nature of the gliding motor remain obscure. It is hoped that the study of the interaction of the Frz proteins will allow greater understanding of these problems.
K1 killer toxin, a pore‐forming protein from yeast
Authors: Bussey H. 
Published: October 2006

SummaryK1 killer toxin is a secreted, pore‐forming protein that kills sensitive yeast cells. The heterodimeric toxin is processed from a precursor in the Golgi, and has allowed identification of the KEX2‐ and KEX1‐encoded proteases. The toxin binds to a glucan receptor on the cell wall of target yeast, and mutational analysis implicates both the a‐ and |5‐toxin sub‐units in receptor binding. Toxin‐resistant mutants with altered cell‐wall glucans have helped to outline a pathway of assembly of these polysaccharides. Patch‐clamp technology has demonstrated the nature of the lethal channel in toxin‐treated plasma membranes. The hydrophobic α‐subunit‐encoding region is the site of all mutations affecting channel formation. Immunity to the toxin is conferred by the toxin precursor, and immunity mutations map to the region encoding the α subunit. The precursor probably competes with the toxin to prevent channel formation in toxin‐producing cells, but the basis of this remains unknown. This toxin/immunity system has a domain structure that differs from that of other characterized toxins and has no known homologues.
H‐DNA formation by the coding repeat elements of neisserial opa genes
Authors: Belland R. J. 
Published: October 2006

SummaryThe coding repeat region of opa genes from Neisseria gonorrhoeae and Neisseria meningitidis determines the expression state of their respective genes through high‐frequency addition or deletion of pentanucleotide coding repeat units (CRs; CTTCT). In vitro analyses of cloned opa gene CR regions using single‐strand specific nucleases, oligonucleotide protection experiments, and modifications of non‐B‐DNA residues indicate that the regions form structures resembling H‐DNA under acidic conditions in the presence of negative supercoiling. The purine/pyrimidine strand bias and H‐palindromic nature of the repeat region are consistent with sequence requirements for H‐DNA formation. Sequences flanking the repeat elements are required to form the H‐DNA structure in vitro as judged by the pattern of exposed non‐B‐DNA residues in CR sequences synthesized as oligonucleotides to form β‐galactosidase::CR translational fusions. The fusions phase vary by addition and deletion of CR elements and the rate of phase variation increases upon induction of the fusion genes. The opa gene CR region is the first reported bacterial H‐DNA structure and is unique in that it ties within the coding sequence for the gene.
Mutational analysis supports a role for multiple structural features in the C‐terminal secretion signal of Escherichia coli haemolysin
Authors: Stanley P. , Koronakis V. , Hughes C. 
Published: October 2006

SummaryWe have carried out an extensive mutational analysis of the C‐terminal signal which targets the export of the 1024‐residue haemolysin protein (HlyA) of Escherichia coli across both bacterial membranes into the surrounding medium. Over 60 variants of the HlyA C‐terminal 53‐amino‐acid sequence were created by oligonucleotide‐directed mutagenesis and fused to the HlyA N‐terminal 830 residues. Transport of the HlyA derivatives by the HlyB/HlyO system was compared with the wild‐type level and the data indicate that the HlyA C‐terminal export signal lies within the last 48 amino acids and comprises three functional domains: an amphipathic, charged helix between residues 1,977 and R,996; a 13‐amino‐acid uncharged region from residue T,997 to S,1009; and an 8‐aminoacid hydroxylated tail at the extreme C‐terminus. Analogous features were found in the C‐terminal sequences of an extended family of haemolysins, leukotoxins and proteases which are secreted by HlyB/HlyD‐type translocators. In particular, all nine proteins which are secreted into the extracellular medium possess potential extended amphipathic helices. These results suggest a possible role for multiple regions of the HlyA C‐terminal export signal in which the first two domains span the membranes and the third domain remains in the cytoplasm.
Identification of gene products of the P1 operon of Mycoplasma pneumoniae
Authors: Sperker B. , Hu P.‐c. , Herrmann R. 
Published: October 2006

SummaryGene P1 of Mycoplasma pneumoniae, which codes for a major adhesin, is flanked by two sequences with open reading frames designated QRF4 and QRF6 (Inamine et al., 1988b). In order to identify proteins translated from these ORFs, gene fusions between the N‐terminus of the RNA replicase of the Escherichia coli bacteriophage MS2 and selected regions of ORF4 and ORF6 were constructed. The corresponding fusion proteins synthesized in Escherichia coli were used to immunize mice. Antisera directed against ORF4‐related sequences did not recognize M. pneumoniae antigens in Western blot analysis, but antisera directed against ORF‐6‐derived fusion proteins reacted with two M. pneumoniae proteins of 40kDa and 90kDa. In addition, some of the antisera also recognized proteins that formed in a sodium dodecyl sulphate/polyacrylamide gel a protein ladder between 115 and 145 kDa.
Identification of phosphatidylinositol‐specific phospholipase C activity in Listeria monocytogenes: a novel type of virulence factor?
Authors: Mengaud J. , Braun‐Breton C. , Cossart P. 
Published: October 2006

SummaryA phospholipase C which cleaves phosphatidylinositol and glycosylphosphatidylinositol (GPI) anchors was identified in Listeria monocytogenes. This 36kDa protein is encoded by the gene plcA, and is homologous to the Bacillus cereus, Bacillus thuringiensis and eukaryotic phosphatidylinositol‐specific phospholipases C (PI‐PLC). Expression of the plcA gene in Escherichia coli correlates with the appearance of PI‐PLC activity in the cells. In Listeria monocytogenes, the activity is secreted to the culture medium. PI‐PLC activity was only found in the two pathogenic species of the genus Listeria, namely L. monocytogenes and L. ivanovii. PI‐PLC activity was lost and virulence decreased when the plcA gene was disrupted in the chromosome. This suggests that the PI‐PLC of L. monocytogenes might be involved in virulence.
P.70 pertactin, an outer‐membrane protein from Bordetella parapertussis: cloning, nucleotide sequence and surface expression in Escherichia coli
Authors: Li L. J. , Dougan G. , Novotny P. , Charles I. G. 
Published: October 2006

SummaryThe gene prn encoding the outer‐membrane protein P.70 (pertactin) from Bordetella parapertussis has been cloned in Escherichia coli and its DNA sequence determined. Analysis of the DNA sequence reveals that the gene has an open reading frame comprising 922 amino acids capable of encoding a protein with a molecular weight of 95177 (P.95). In vivo processing of this precursor yields a protein with an estimated Mr of 70kDa (P.70) which is located on the surface of B. parapertussis. Homology between the prn gene from B. parapertussis and that from Bordetella pertussis is 91.3%. The homology is 93% when the protein sequence of P.95 is aligned with that of P.93 from B. pertussis. The major differences between the P.70 pertactin from B. parapertussis and the P.69 pertactin from B. pertussis occur in the number of reiterated units within the repeat motifs found in both proteins; the sequence Gly–Gly–Xaa–Xaa–Pro is repeated four times in the P.70 pertactin, and five times in the P.69 pertactin, while the sequence Pro–Gln–Pro occurs nine times in P.70 pertactin and five times in P.69 pertactin. Cloning of the gene for P.95 in an E. coli expression vector results in the synthesis of a protein that mimics native gene expression in B. parapertussis, i.e. the P.95 protein is synthesized and subsequently processed to yield the P.70 form of the protein on the surface of the cell.
Segment IV of a Salmonella flagellin gene specifies flagellar antigen epitopes
Authors: Newton S. M. C. , Wasley R. D. , Wilson A. , Rosenberg L. T. , Miller J. F. , Stocker B. A. 
Published: October 2006

SummaryEach of the two mutants isolated from a fliC (=hag, flagellin‐deficient) Escherichia coli strain made motile by a plasmid carrying the fliC gene of Salmonella muenchen by selection for motility in the presence of anti‐d (Salmonella flagellar antigen) serum had both lost and gained one or more subfactors of the wild‐type antigen. In one more mutant codon 246 was GAC (alanine) instead of GCC (asparagine); the other had a deletion of 105 base pairs, explicable by a 10bp direct repeat, starting at bases 782 and 887. The in vitro removal of a 48bp EcoRV(631)/EcoRV(679) fragment produced plasmid pLS408, which was found to lack a subfactor of wild‐type antigen d but able to confer motility on flagellin‐negative Salmonella sp.(and used for insertion of epitope‐specifying oligonucleotides at its EcoRV site). Immunoblotting with absorbed and unabsorbed sera from rabbits immunized with E. coli with wild‐type or mutated antigen d showed that the fusion proteins specified by λ gt11 with the N‐terminal part of gene lacZ joined to a restriction fragment coding for residues 145–391 of flagellin gave the same patter of parent‐specific and mutant‐specific reactions as the flagellate bacteria. Four out of five similarly selected mutants had the same 105bp deletion as the first‐isolated mutant; the fifth had a 72bp deletion made possible by a 7‐base pair direct repeat, starting at positions 649 and 721. All these changes in serological character without loss of function affected segment IV, specifying residues 182 to 308 of the total of 505, where there is little homology between different flagellar antigen alleles.
Design of camp–CRP‐activated promoters in Escherichia coli
Authors: Valentin‐Hansen P. , Hoist B. , Søgaard‐Andersen L. , Martinussen J. , Nesvera J. , Douthwaite S. R. 
Published: October 2006

SummaryWe have studied the doeP2 promoter of Escherichia coli to define features that are required for optimal activation by the complex of adenosine 3′, 5′ monophosphate (cAMP) and the cAMP receptor protein (CRP). Systematic mutagenesis of deoP2 shows that the distance between the CRP site and the ‐10 hexamer is the crucial factor in determining whether the promoter is activated by camp–CRP. Based on these observations, we propose that camp–CRP‐activated promoters can be created by correctly aligning a CRP target and a ‐ 10 hexamer. This idea has been successfully tested by converting both a CRP‐in‐dependent promoter and a sequence resembling the consensus ‐10 hexamer to strongly camp–CRP‐activated promoters.