bluuebunny/crossref_metadata_2025_split huggingface dataset:
Functional domains of colicin A
Authors: Baty D. , Frenette M. , Lioubès R. , Geli V. , Howard S. P. , Pattus F. , Lazdunski C. 
Published: October 2006

SummaryA large number of mutations which introduce deletions in colicin A have been constructed. The partially deleted colicin A proteins were purified and their activity in vivo (on sensitive cells) and in vitro (in planar lipid bilayers) was assayed. The receptor‐binding properties of each protein were also analysed. From these results, we suggest that the NH2‐terminal region of colicin A (residues 1 to 172) is involved in the translocation step through the outer membrane. The central region of colicin A (residues 173 to 336) contains the receptor‐binding domain. The COOH‐terminal domain (residues 389 to 592) carries the pore‐forming activity.
Specificity of the lactoferrin and transferrin receptors in Neisseria gonorrhoeae
Authors: Lee B. C. , Schryvers A. B. 
Published: October 2006

SummaryLactoferrin (LF) and transferrin (TF) are postulated to be important physiological sources of iron for Neisseria gonorrhoeae. A dot binding assay involving the use of gonococcal total membranes derived from cells grown in iron‐limited conditions demonstrated the presence of separate receptors for LF and TF. The ligand and functional specificities of these receptors were examined in competition‐binding and growth experiments. The results indicate that the LF and TF receptors are highly specific for the human protein, suggesting that this property may be partially responsible for conferring the human host specificity of N. gonorrhoeae.
Identification of the membrane component of the anion pump encoded by the arsenical resistance operon of R‐factor R773
Authors: San Francisco M. J. D. , Tisa L. S. , Rosen B. P. 
Published: October 2006

SummaryThe arsenical resistance (ars) operon of the conjugative R‐factor R773 encodes an ATP‐driven anion extrusion pump, producing bacterial resistance to arsenicals. There are three structural genes, of which the product of the middle gene, arsB, has not previously been identified. From nucleotide sequence data, the ArsB protein is predicted to be a 45577 Dalton hydrophobic protein. A mini‐Mu transposition procedure was used to construct an arsB‐lacZ gene fusion, producing a hybrid ArsB‐β‐galactosidase protein which was localized in the inner membrane. The operon was cloned into a T7 RNA polymerase expression vector. In addition to the previously identified ArsA and ArsC proteins, the cells synthesized an inner membrane protein with an apparent mass of 36 kD identified as the ArsB protein.
Penicillin‐binding protein 2 genes of non‐β‐lactamase‐producing, penicillin‐resistant strains of Neisseria gonorrhoeae
Authors: Dowson C. G. , Jephcott A. E. , Gough K. R. , Spratt B. G. 
Published: October 2006

SummaryOligonucleotides that correspond to regions of the penicillin‐binding protein 2 gene (penA) that differ between penicillin‐sensitive and penicillin‐resistant strains have been used as probes to classify the penA genes in a collection of penicillin‐resistant gonococci isolated in Britain. 44/47 of those gonococcal strains that had minimal inhibitory concentrations of ≥0.25 μg benzylpenicillin per ml contained extensively altered penA genes which appeared to be very similar (or identical) to one or other of the two classes of altered penA genes that have been described previously. Since these two classes of altered penA genes are related, it appears that the great majority of the altered penA genes of non‐β‐lactamase‐producing penicillin‐resistant gonococci have a clonal origin. The other three penicillin‐resistant strains had altered penA genes that were different to those described previously. A crucial step in the development of the altered forms of PBP2 with decreased affinity for penicillin appears to have been the insertion of an extra codon within the transpeptidase domain of the penA gene. This insertion was found in the penA gene of all gonococci with minimal inhibitory concentrations of >0.016μg benzylpenicillin per ml but was not found in any strains with minimal inhibitory concentrations of ≤0.016μg per ml.
Conserved lipoprotein H.8 of pathogenic Neisseria consists entirely of pentapeptide repeats
Authors: Woods J. P. , Spinola S. M. , Strobel S. M. , Cannon J. G. 
Published: October 2006

SummaryThe pathogenic Neisseria, N. gonorrhoeae and N. meningitidis, possess an outer membrane protein (OMP), designated H.8, with a conserved monoclonal antibody (MAb)‐binding epitope. We determined the DNA sequence of a gonococcal H.8 gene, and confirmed the relationship between the cloned gene and the H.8 OMP by constructing a gonococcal mutant lacking H.8. The predicted H.8 OMP is a lipoprotein 71 amino acids in length, composed of 13 repeats of a consensus sequence AAEAP with perfect 5‐residue periodicity. The AAEAP units form a repeating epitope that comprises the entire predicted sequence of the protein.
Extensive re‐modelling of the transpeptidase domain of penicillin‐binding protein 2B of a penicillin‐resistant South African isolate of Streptococcus pneumoniae
Authors: Dowson C. G. , Hutchison A. , Spratt B. G. 
Published: October 2006

SummaryClinical isolates of Streptococcus pneumoniae that have greatly increased levels of resistance to penicillin (>1000‐fold) have been reported from South Africa during the last ten years. Penicillin resistance in these strains is entirely due to the development of penicillin‐binding proteins (PBPs) with decreased affinity for penicillin. We have cloned and sequenced the coding region for the transpeptidase domain of penicillin‐binding protein 2B from three penicillin‐sensitive strains of S. pneumoniae and from a penicillin‐resistant South African strain. The amino acid sequences of the transpeptidase domains of PBP2B of the three penicillin‐sensitive strains were identical and there were only between one and four differences in the nucleotide sequences of their coding regions. The corresponding region of the PBP2B gene from the penicillin‐resistant strain differed by 74 nucleotide substitutions which resulted in 17 alterations in the amino acid sequence of PBP2B. The most remarkable alteration that has occurred during the development of the‘penicillin‐resistant’form of PBP2B is the substitution of seven consecutive residues in a region that is predicted to form a loop at the bottom of the penicillin‐binding site.
Phosphoenolpyruvate:sugar phosphotransferase system of Bacillus subtilis: nucleotide sequence of ptsX, ptsH and the 5’‐end of ptsl and evidence for a ptsHI operon
Authors: Gonzy‐Tréboul G. , Zagorec M. , Rain‐Guion M.‐C. , Steinmetz M. 
Published: October 2006

SummaryThe nucleotide sequence of a 1689bp fragment of the Bacillus subtiiis locus containing ptsX (a crr‐like gene), ptsH (coding for HPr), and the 5’‐end of ptsl (coding for Enzyme I) was determined. The deduced amino acid sequences of ptsH and the N‐terminal part of ptsl were compared to those of Streptococcus faecalis and Escherichia coli. Transcription fusion demonstrated that ptsHl constitutes an operon. An open reading frame overlapping the main part of ptsH in the opposite sense was shown to be expressed in vivo, using protein fusions with β‐galactosidase. The deduced amino acid sequence of ptsX showed significant homology with that of Salmonella typhimurium glucose‐specific Enzyme III. ptsX was preceded by an open reading frame whose amino acid sequence showed strong homology with the C‐terminal part of E.coli Enzyme IIGlc.
Nucleotide sequences of the pbpX genes encoding the penicillin‐binding proteins 2x from Streptococcus pneumoniae R6 and a cefotaxime‐resistant mutant, C506
Authors: Laible G. , Hakenbeck R. , Sicard M. A. , Joris B. , Ghuysen J.‐M. 
Published: October 2006

SummaryDevelopment of penicillin resistance in Streptococcus pneumoniae is due to successive mutations in penicillin‐binding proteins (PBPs) which reduce their affinity for β‐lactam antibiotics. PBP2x is one of the high‐Mr PBPs which appears to be altered both in resistant clinical isolates, and in cefotaxime‐resistant laboratory mutants. In this study, we have sequenced a 2564 base‐pair chromosomal fragment from the penicillin‐sensitive S. pneumoniae strain R6, which contains the PBP2x gene. Within this fragment, a 2250 base‐pair open reading frame was found which coded for a protein having an Mr of 82.35 kD, a value which is in good agreement with the Mr of 80–85kD measured by SDS‐gel electrophoresis of the PBP2x protein itself. The N‐terminal region resembled an unprocessed signal peptide and was followed by a hydrophobic sequence that may be responsible for membrane attachment of PBP2x. The corresponding nucleotide sequence of the PBP2x gene from C504, a cefotaxime‐resistant laboratory mutant obtained after five selection steps, contained three nucleotide substitutions, causing three amino acid alterations within the β‐lactam binding domain of the PBP2x protein. Alterations affecting similar regions of Escherichia coli PBP3 and Neisseria gonorrhoeae PBP2 from β‐lactam‐resistant strains are known. The penicillin‐binding domain of PBP2x shows highest homology with these two PBPs and S. pneumoniae PBP2b. In contrast, the N‐terminal extension of PBP2x has the highest homology with E. coli PBP2 and methicillin‐resistant Staphylococcus aureus PBP2′. No significant homology was detected with PBP1a or PBP1b of Escherichia coli, or with the low‐Mr PBPs.
Positive control of colanic acid synthesis in Escherichia coli by rmpA and rmpB, two virulence‐plasmid genes of Kiebsiella pneumoniae
Authors: Nassrf X. , Honoré N. , Vasselon T. , Cole S. T. , Sansonetti P. J. 
Published: October 2006

SummaryIn Kiebsiella pneumoniae, the mucoid phenotype, which is a virulence factor, is distinct from capsule production. It is positively controlled by a plasmid gene, designated rmpA. When introduced into certain Escherichia coli strains, rmpA induces expression of a mucoid phenotype, which results from overproduction of colanic acid at 30° C but not at 37°C. In E. coli, production of colanic acid is regulated by three genes: rcsA and rcsB which act as positive regulators, and rcsC which is a negative effector. In this work we present evidence that the rmpA gene complemented an rcsA, Ion double mutant of E. coli, but not an rcsA, ion+ isolate. This leads to the suggestion that rmpA expressed an rcsA‐like activity and like rcsA, was negatively controlled at post‐transcriptional level by the Lon protease. The nucleotide sequence of rmpA is reported. No homology could be found between the 27 kiloDalton RcsA protein and the deduced amino acid sequence of the 15.5 kiloDalton RmpA protein. Another gene, rmpB, which was required in E. coli recA isolates for full expression of rmpA at 30° C, has been identified on the K. pneumoniae virulence plasmid and shown to encode a 37 kiloDalton protein. Although rmpB was closely linked to rmpA, it was not present on the same transcriptional unit. These results suggested that induction of colanic acid synthesis by the K. pneumoniae virulence gene rmpA, was, at least in E. coli, under the control of the RecA network via rmpB, which may act as a positive regulator of rmpA. We conclude that these plasmid genes may function in K. pneumoniae as regulatory genes controlling the mucoid phenotype, which is itself encoded by the chromosome.
CRP/cAMP‐ and CytR‐regulated promoters in Escherichia coli K12: the cdd promoter
Authors: Valentin‐Hansen P. , Holst B. , Josephsen J. , Hammer K. , Albrechtsen B. 
Published: October 2006

SummaryTranscriptional regulation of the deoP2 promoter by the cyclic AMP/cyclic AMP receptor protein complex (cAMP/CRP) and the CytR repressor requires two high‐affinity CRP targets located around ‐41 and ‐93 bp preceding the start site for transcription. Here we report the structure of cddP, another CRP/CytR‐regulated promoter. In common with what was found in deo, the cdd promoter also contains multiple CRP targets. Thus, using the DNasel footprinting procedure, tandem CRP binding sites were identified around ‐41 and ‐93. These findings support a general model for CytR binding and CytR regulation, in which (i) CytR and the CRP/cAMP complex bind to similar or Identical targets, (ii) two or more targets are necessary for proper binding of CytR to a promoter region, and (iii) CytR represses transcription by antagonizing cAMP/CRP activation.
The Yersinia yop regulon
Authors: Cornelis G. R. , Biot T. , de Rouvroit C. Lambert , Michiels T. , Mulder B. , Sluiters C. , Sory M.‐P. , Van Bouchaute M. , Vanooteghem J.‐C. 
Published: October 2006

SummaryGrowth of yersiniae is restricted at 37°C in the absence of calcium ions. This phenomenon correlates with the massive release of a set of proteins called Yops. Growth restriction and Yops production are governed by a 70 kb plasmid called pYV. yop genes are distributed throughout pYV and constitute a thermoactivated regulon controlled by the gene virF. The transcription activator VirF is a member of a new family of regulators including those of the arabinose and rhamnose operons as well as a regulator of enteric colonization pili. The role of calcium ions on the release of Yops remains largely unknown.
A family of genes encoding a cell‐killing function may be conserved in all Gram‐negative bacteria
Authors: Poulsen L. K. , Larsen N. W. , Molin S. , Andersson P. 
Published: October 2006

SummaryThe relF gene in Escherichia coli is related to the hok gene on plasmid R1. Both genes encode small proteins which, when overexpressed In E. coli lead to collapse of the membrane potential and cell death. A third gene, designated gef, which encodes a homologous cell‐toxic protein, has been isolated from E. coli DNA. Both gef and relF are transcribed in E. coli and subject to post‐transcriptional regulation which, in the case of gef, is coupled to translation of a leader sequence. The finding of homologous sequences in such distantly related bacteria as Agrobacterium and Rhizobium species suggests an important physiological role.
The Escherichia coli regulatory protein MetJ binds to a tandemly repeated 8bp palindrome
Authors: Davidson B. E. , Girons I. Saint 
Published: October 2006

SummarySite‐directed oligonucleotide mutagenesis has been used to isolate thirty four new mutants in the regulatory region of the Escherichia coliK12 gene, metF. The mutants include single base pair (bp) substitutions and insertions, double bp substitutions and one 7bp deletion. The effects of these and another five previously described mutants on the transcriptional regulation of metF have been analysed by using a metF′‐lac'Z fusion in a low copy‐number plasmid. These data, and those obtained from DNAse protection studies using pure MetJ with wild‐type and mutant metF operator DNA, show that the metF operator is comprised of five tandem 8bp repeat units that overlap the ‐10 region of the metF promoter. In the presence of the co‐repressor S‐adenosylmethionine, the DNAse protection studies yielded dissociation constants of 150nM and 300 nM for the interaction of MetJ with repeat units 1 to 4 and repeat unit 5, respectively. In the absence of co‐repressor, the dissociation constants obtained for these interactions are four to five times greater. It is proposed that regulation at the metF operator requires four molecules of MetJ dimer to bind to the five 8bp repeat units to form a tandem, overlapping array. Interactions between MetJ molecules make an essential contribution to the stability of this protein‐DNA complex.
RNA polymerase heterogeneity in Streptomyces coelicolor A3(2)
Authors: Buttner M. J. 
Published: October 2006

SummaryRecent genetic and biochemical experiments have revealed the existence of at least seven different Sigma factors in Streptomyces coelicolor and demonstrated a role for alternative sigma factors in the control of differentiation and also in the transcription of primary metabolic genes. S. coelicolor has four genes predicted to encode sigma factors that are very closely related to the σ70 polypeptide of Escherichia coli.
Genes for biosynthesis and assembly of CS3 pili of CFA/II enterotoxigenic Escherichia coli: novel regulation of pilus production by bypassing an amber codon
Authors: Jalajakumari M. B. , Thomas C. J. , Halter R. , Manning P. A. 
Published: October 2006

SummaryThe complete nucleotide sequence of the 4746bp HindIII fragment encoding the genes for the biosynthesis and assembly of CS3 pili has been determined. By site‐directed mutagenesis in conjunction with analysis of the plasmid‐encoded proteins in minicells, the actual reading frames for the various products have been determined. This demonstrated that the genes for four of the proteins (63 kD, 48 kD, 33 kD, and 20kD in size) are encoded entirely within the same open reading frame as a fifth protein (104kD). However, for synthesis of this latter protein, suppression or readthrough of an internal amber codon is required. Termination at this codon is also necessary for synthesis of the former proteins. Two further proteins are also encoded within the HindIII fragment: a 27 kD precursor of a periplasmic protein and the 17.5kD precursor of the major CS3 fimbrial subunit.
Regulation of the RAD2 gene of Saccharomyces cervisiae
Authors: Siede W. , Robinson G. W. , Kalainov D. , Malley T. , Friedberg E. C. 
Published: October 2006

SummaryRegulation of the DNA damage‐inducible RAD2 gene was investigated in yeast cells transformed with centromeric plasmids containing RAD2‐lacZ fusion constructs. Deletion analysis defined several regions in the 350 bp region upstream of the translational start codon which are required for induction of β‐galactosidase activity. No deletions resulted in constitutivety enhanced expression. We therefore conclude that induction of RAD2 by DNA‐damaging agents is positively regulated. Two domains required for induction have a similar sequence and are located ∼70 and ∼140bp upstream of the major transcriptional start site. Four other sequence domains required for induction contain uninterrupted poly(dA) poly(dT) stretches 9‐13 bp long. Deletion of some of these AT‐rich domains also affects constitutive expression of RAD2. Expression of RAD2 is not cell‐cycle‐regulated in mitotic cells. However, meiosis is accompanied by increased steady‐state levels of RAD2 mRNA in the absence of DNA damage. This enhanced transcription is not dependent on the presence of upstream sequences required for regulation of induction by DNA damage. Increased steady‐state levels of RAD2 mRNA are induced by cycloheximide in asynchronously dividing populations of cells, but not in non‐replicating cells arrested in G1 phase of the cell cycle. Following exposure to u.v. irradiation induction is also dramatically reduced in non‐replicating cells.
Nickel deficiency gives rise to the defective hydrogenase phenotype of hydc and fnr mutants in Escherichia coli
Authors: Wu L.‐F. , Mandrand‐Berthelot M.‐A. , Waugh R. , Edmonds C. J. , Holt S. E. , Boxer D. H. 
Published: October 2006

SummaryHydrogenase activity and other hydrogenase‐related functions can be restored to hydC mutants by the specific addition of nickel salts to the growth medium. These mutants are defective in all three hydrogenase isoenzymes and the restoration is dependent upon protein synthesis. The cellular nickel content of the mutant when grown in LB medium is less than 1% of that of the parental strain. Partial suppression of the hydrogenase phenotype of hydC mutants occurs when growth takes place in a different medium. This correlates with an increased cellular nickel content. The phenotype of the mutant is also fully suppressed by growth in media of very low magnesium content. Such media facilitate nickel uptake via the magnesium transport system, which leads to the acquisition of a normal cellular nickel content. Mutations in the fnr gene, which encodes a transcriptional regulator for several anaerobically expressed enzymes, abolishes hydC expression and gives rise to a defective hydrogenase phenotype. The hydrogenase phenotype of fnr is closely similar to that of hydC in all respects examined. The hydrogenase activity of fnr strains can be restored by the presence of a functional hydC gene on a multicopy plasmid. The hydrogenase phenotype of fnr strains therefore arises indirectly via suppression of hydC, which leads to a low cellular nickel content. Nickel has no influence on fumarate reductase or nitrate reductase activities in fnr strains. The hydrogen‐metabolism phenotype of fnr strains is, therefore, dependent upon their ability to acquire nickel from growth media. It is likely that hydC encodes a specific transport system for nickel.
Multi‐gene amplification: simultaneous detection of three virulence genes in diarrhoeal stool
Authors: Frankel G. , Giron J. A. , Valmassoi J. , Schoolnik G. K. 
Published: October 2006

SummaryEnterotoxigenic Escherichia coli (ETEC) and Shigella account for a substantial proportion of acute diarrhoeal illnesses among Third‐World children. Rapid detection of these infectious agents in faeces followed by the prompt implementation of public health measures could help reduce their spread during the early phase of epidemics. Towards this end, three pairs of synthetic oligonucleotide primers were prepared and shown to hybridize specifically to the genes encoding the heat‐stable (ST) and the heat‐labile (LT) enterotoxins of ETEC and to invasion‐associated loci (ial) of the large Shigella virulence plasmid. When the three primer pairs were used together in the polymerase chain reaction (PCR), the three corresponding genetic loci could be simultaneously amplified using DNA extracted directly from stool; the amplified products were readily detected by ST‐, LT‐ and ial‐specific, alkaline phosphatase‐labelled oligonucleotide probes (AP probes). The performance of this system was evaluated in a Mayan community in southeastern Mexico, where diarrhoeal illnesses are a common cause of childhood morbidity and mortality. Using only simple and inexpensive laboratory equipment, multigene amplification with these primers and probes led to the identification of ETEC and/or Shigella in the stools of 20 out of 71 children with diarrhoea; the procedure could be completed in seven hours and was more sensitive than conventional diagnostic tests or DNA probes used without amplification.
Diversity of the outer membrane protein P2 gene from major clones of Haemophilus influenzae type b
Authors: Munson R. , Bailey C. , Grass S. 
Published: October 2006

SummaryIn previous studies, it has been demonstrated that outer membrane protein P2 from Haemophilus influenzae type b has porin activity and that antibody directed against P2 is protective in an infant rat bacteraemic model.Outer membrane protein subtyping has been employed to subclassify type b Haemophilus isolates. Strain MinnA has the outer membrane protein subtype 1H and is representative of the dominant clonal group of disease‐producing isolates in the United States. In the present study, the P2 gene from strain MinnA was employed to probe EcoRI‐ and PvuII‐digested chromosomal DNA from 24 Haemophilus influenzae type b isolates representative of the common outer membrane protein subtype groups observed throughout the world. Restriction fragment length polymorphisms were identified for the members of the outer membrane protein subtype 3L group, but not for the other subtypes examined.The P2 gene from each of four prototype isolates was then cloned, sequenced and compared to the previously reported sequence of the strain MinnA gene. The P2 gene from each of two isolates with the outer membrane protein subtype 3L was identical to the MinnA P2 sequence. The P2 gene from a subtype 2L isolate differed by a single nucleotide and the gene from a subtype 6U isolate differed by 13 nucleotides. Thus, the P2 protein is highly conserved among type b isolates.