bluuebunny/crossref_metadata_2025_split huggingface dataset:
The stable BRP signal peptide causes lethality but is unable to provoke the translocation of cloacin DF13 across the cytoplasmic membrane of Escherichia coli
Authors: van der Wal F. J. , Oudega B. , Kater M. M. , ten Hagen‐Jongman C. M. , de Graaf F. K. , Luirink J. 
Published: October 2006

SummaryThe bacteriocin release protein (BRP) mediates the secretion of cloacin DF13. The BRP precursor is slowly processed to yield the mature BRP and its stable signal peptide which is also involved in cloacin DF13 secretion. The function of the stable BRP signal peptide was analysed by constructing two plasmids. First, the stable BRP signal peptide was fused to the murein lipoprotein and, second, a stop codon was introduced after the BRP signal sequence. Exchange of the unstable murein lipoprotein signal peptide for the stable BRP signal peptide resulted in an accumulation of precursors of the hybrid murein lipoprotein. This indicated that the BRP signal peptide, as part of this hybrid precursor, is responsible for the slow processing. The stable BRP signal peptide itself was not able to direct the transfer of cloacin DF13 into the periplasmic space or into the culture medium. Over‐expression of the BRP signal peptide was lethal and caused ‘lysis’. Subcellular fractionation experiments revealed that the BRP signal peptide is located exclusively in the cytoplasmic membrane whereas the mature BRP, targeted by either the stable BRP signal peptide or the unstable Lpp signal peptide, is located in both the cytoplasmic and outer membrane.These results are in agreement with the hypothesis that the stable signal peptide and the mature BRP together are required for the passage of cloacin DF13 across the cell envelope.
Expression and mutational analysis of the nucleoid‐associated protein H‐NS of Salmonella typhimurium
Authors: Hinton Jay C. D. , Santos Diogenes S. , Seirafi Alexander , Hulton Christopher S. J. , Pavitt Graham D. , Higgins Christopher F. 
Published: October 2006

SummaryThe H‐NS (H1) protein is a major component of bacterial chromatin. Mutations in the hns (osmZ) gene encoding H‐NS are highly pleiotropic, affecting the expression of many unrelated genes in an allele‐specific manner. H‐NS expression was found not to vary with growth phase or growth medium osmolarity. Additionally, 10 independent hns mutations were isolated and characterized. Five of these mutations were the result of an IS10 insertion, each generating a truncated polypeptide. The other five mutations were the same specific deletion of one amino acid, δla46. The various hns mutations exhibited different phenotypes and influenced ONA topology to variable extents. Implications for the mechanism by which H‐NS influences gene expression are discussed.
The plasmid‐encoded chloramphenicol‐resistance protein of Rhodococcus fascians is homologous to the transmembrane tetracycline efflux proteins
Authors: Desomer J. , Vereecke D. , Crespi M. , Van Montagu M. 
Published: October 2006

SummaryThe nucleotide sequence of the chloramphenicol‐resistance gene (cmr) of Rhodococcus fascians NCPPB 1675 (located on the conjugative plasmid pRF2) allowed the identification of two possible open reading frames (ORFs), of which ORF1 was consistent with the mutational analysis. Biochemical analysis of cmr revealed that it does not encode an antibiotic‐modifying enzyme. The amino acid sequence of 0RF1 predicted a hydrophobic protein, with 12 putative membrane‐spanning domains, homologous to proteins involved in the efflux of tetracycline across the plasma membrane. Expression of the cmr gene was induced by addition of chloramphenicol to the growth media. The promoter of this gene was restricted to 50 bp upstream from a 200 bp 5′‐untrans‐lated mRNA region, the latter containing two inverted repeats. At the amino acid level, the cmr gene is 52% identical to a previously identified chloramphenicol‐resistance determinant in Streptomyces lividans, indicating a wider dispersion of this type of cmr gene among the actinomycetes.
Identification and characterization of virK, a virulence‐associated large plasmid gene essential for intercellular spreading of Shigella flexneri
Authors: Nakata N. , Sasakawa C. , Okada N. , Tobe T. , Fukuda I. , Suzuki T. , Komatsu K. , Yoshikawa M. 
Published: October 2006

SummarySeven virulence loci have been identified by Tn5 insertion mutagenesis on the large 230 kb plasmid (pMYSH6000) of Shigella flexneri 2a. In this study, we used Tn10 insertion mutagenesis and identified a novel virulence locus on pMYSH6000 responsible for bacterial spread. Characterization of the invading bacteria of the Tn10 insertion mutants in the epithelial cells revealed that the bacteria were capable of at least some intracellular spreading but not intercellular spreading. Immunoblot analysis of lysates of the Tn10 insertion mutants with a VirG‐specific antipeptide antibody revealed diminished levels of the 116 kDa VirG protein. The virG mRNA in the mutants, however, was expressed at the same level as that in the wild type. The DNA region required for the virulence phenotype was localized to a 1.6 kb DNA sequence in the Sal I‐K fragment on the plasmid, and thus the locus was designated virK. Expression of virK in Escherichia coli using a T7 RNA polymerase‐dependent promoter system yielded a 36 kDa protein. The nucleotide sequence of 1642 bp encoding VirK function was determined, and an open reading frame encoding 316 amino acid residues was shown to encode the VirK protein. The virK region was highly conserved among the large virulence plasmids of shigellae and enteroinvasive Escherichia coli. These results suggest that VirK function is an essential virulence determinant for shigellae Involved in the expression of virG gene product at post‐transcriptional level.
Analysis of the transcriptional activity of the hut promoter in Bacillus subtilis and identification of a cis‐acting regulatory region associated with catabolite repression downstream from the site of transcription
Authors: Oda M. , Katagai T. , Tomura D. , Shoun H. , Hoshino T. , Furukawa K. 
Published: October 2006

SummaryLevels of transcripts initiated at a hut promoter in Bacillus subtilis were analysed. The addition of histidine to the culture medium increased the level of the transcript sixfold. In the presence of histidine and glucose together, the level of the transcript was reduced to the level in the absence of Induction. Furthermore, addition of a mixture of 16 amino acids to cultures of induced cells and of catabolite‐repressed cells decreased levels of the transcript 16‐fold and 2.6‐fold, respectively. Thus, it appears that at least three regulatory mechanisms associated with induction, catabolite repression, and amino acid repression, control the transcriptional activity of the hut promoter. Expression of the hut promoter–lacZ fusions that contained various regions of the hutP gene and deletion analysis of the hutP region revealed a cis‐acting sequence associated with catabolite repression that was located between positions +204 and +231 or around position +203.
Antibodies to N‐terminal peptides of gonococcal porin are bactericidal when gonococcal lipopolysaccharide is not sialylated
Authors: Elkins Christopher , Carbonetti Nicholas H. , Varela Victor A. , Stirewalt Derek , Klapper David G. , Sparling P. Frederick 
Published: October 2006

SummarySix synthetic 25‐mer peptides corresponding to certain presumed surfaced‐exposed regions of gonococcal porin protein I (PI) were made from strains FA19 (PIA) and MS11 (PIB). Four peptides were immunogenic in rabbits. Affinity‐purified antisera against both PIA and PIB N‐terminal peptides were bactericidal for homologous gonococci and many heterologous PI serovars. However, sialylation of gonococcal lipopolysaccharide (LPS) by growth of gonococci in the presence of cytidine monophosphate–neuraminic acid (CMP–NANA) abrogated the bactericidal activity of these antisera. Binding of anti‐PI monoclonal antibodies to whole gonococci was reduced two‐ to fourfold by sialylation of LPS, suggesting that sialylation may inhibit bactericidal activity by masking porin epitopes. However, binding of anti‐PII (Opa) monoclonal antibodies was not inhibited, yet complement‐mediated killing was inhibited by sialylated LPS. Binding of complement components C3 and C9 was inhibited in the presence of either anti‐PI or anti‐PII monoclonals when gonococci were grown in the presence of CMP–NANA. Thus sialylation inhibited both anti‐PI antibody binding and complement deposition, with a resultant decrease in bactericidal activity.
Control of gene expression in the temperate coliphage 186. X. The cl repressor directly represses transcription of the late control gene B
Authors: Dibbens Justin A. , Gregory Stephen L. , Egan J. Barry 
Published: October 2006

SummaryWe have found that the repressor of 186 lytic transcription, CI, represses transcription of the late control gene B, with no involvement of the B protein Itself. In clone studies we showed that CI repressed transcription from the B promoter and that temperature inactivation of CIts led to B derepression. We conclude that CI repressor directly represses transcription of the Bgene and, with prophage induction, it is probable that the inactivation of the CI repressor not only derepresses early lytic transcription, but also derepresses B gene transcription, leading to the activation of transcription from the late promoters.
Characterization of a Bordetella pertussis fimbrial gene cluster which is located directly downstream of the filamentous haemagglutinin gene
Authors: Willems Rob J. L. , van der Heide Han G. J. , Mooi Frits R. 
Published: October 2006

SummaryThe biosynthesis of fimbriae is a complex process requiring multiple genes which are generally found clustered on the chromosome. In Bordetella pertussis, only major fimbrial subunit genes have been identified, and no evidence has yet been found that they are located in a fimbrial gene cluster. To locate additional genes involved in the biosynthesis of B. pertussis fimbriae, we used TnphoA mutagenesis. A PhoA+ mutant (designated B176) was isolated which was affected in the production of both serotype 2 and 3 fimbriae. Cloning and sequencing of the DNA region harbouring the transposon insertion revealed the presence of at least three additional fimbrial genes, designated fimB, fimC and fimD. The transposon was found to be located In fimD. Analysis of PhoA activity indicated that the fimbrial gene cluster was positively regulated by the bvg locus. A potential binding site for BvgA was observed upstream of fimB. FimB showed homology with the so‐called chaperone‐like fimbrial proteins, while FimC was homologous with a class of fimbrial proteins located in the outer membrane and presumed to be involved in transport and anchorage of fimbrial subunits. An insertion mutation in fimB abolished the expression of fimbrial subunits, implicating this gene in the biosynthesis of both serotype 2 and 3 fimbriae. Upstream of fimB a pseudogene (fimA) was observed which showed homology with the three major fimbrial subunit genes, fim2, fim3 and fimX. The construction of a phylogenetic tree suggested that fimA may be the primordial major fimbrial subunit gene from which the other three were derived by gene duplication. Interestingly, the fimbrial gene cluster was found to be located directly downstream from the gene coding for the filamentous haemagglutinin, an important B. pertussis adhesin, possibly suggesting co‐operation between the two loci in the pathogenesis of pertussis.
Identification and sequence of a Na+‐linked gene from the marine bacterium Alteromonas haloplanktis which functionally complements the dagA gene of Escherichia coli
Authors: MacLeod Patricia R. , MacLeod Robert A. 
Published: October 2006

SummaryA 4.0 kb fragment from a plasmid genomic DNA library of the marine bacterium Alteromonas haloplanktis ATCC 19855 was found in the presence of Na+ to complement the dagA gene of Escherichia coli. We have completely sequenced this fragment and the position of the Na+‐linked D‐alanine glycine permease gene (dagA) on the fragment has been determined by complementation. The predicted carrier protein consists of 542 amino acid residues (Mr 58955). Its hydropathy profile suggests it is composed of eight transmembrane segments with a long hydrophilic region between segments six and seven. Significant similarity has been found between this Na+‐linked permease and the Na*/proline permeases of E. coli and Salmonella typhimurium and the human and rabbit intestinal Na+/glucose cotransporters.
Identification and characterization of FliY, a novel component of the Bacillus subtilis flagellar switch complex
Authors: Bischoff David S. , Ordal George W. 
Published: October 2006

SummaryThe Bacillus subtilis gene encoding FMY has been cloned and sequenced. The gene encodes a 379‐amino‐acid protein with a predicted molecular mass of 41 054 daltons. FliY is partly homologous to the Escherichia coli and Salmonella typhimurium switch proteins FliM and FliN. The N‐terminus of FliY has 33% identity with the first 122 amino acids of FliM, whereas the C‐terminus of FliY has 52% identity with the last 30 amino acids of FMN. The middle 60% of FliY is not significantly homologous to either of the proteins. A fliY::cat null mutant has no flagella. Motility can be restored to the mutant by expression of fliY from a plasmid, although chemotaxis is still defective since the strain exhibits smooth swimming behaviour. fliY::cat is in the cheD complementation group. One of the cheD point mutants does not switch although the population grown from a single cell has both smooth swimming and tumbling bacteria, implying that the switch is locked. Expression of fliY in wild‐type B. subtilis makes the cells more smooth‐swimming but does not appear to affect chemotaxis. Expression of fliY in wild‐type S. typhimurium severely inhibits chemotaxis and also makes the cells smooth swimming. Expression in a non‐motile S. typhimurium fliN mutant restores motility but not chemotaxis, although expression in a non‐motile E. coli fliM mutant does not restore motility. The homology, multiple phenotypes, and inter‐species complementation suggest that FliY forms part of the B. subtilis switch complex. The function of the unique part of FliY and the evolutionary relationship between FliY and FliN are discussed in terms of the novel properties of B. subtilis chemotaxis.
The molecular basis for positive regulation of cys promoters in Salmonella typhimurium and Escherichia coli
Authors: Kredich Nicholas M. 
Published: October 2006

SummaryMost genes required for cysteine biosynthesis in Salmonella typhimurium and Escherichia coli are positively regulated by cysB, which encodes a transcriptional activator belonging to the LysR family of regulatory proteins. CysB protein binds just upstream of the ‐35 region of positively regulated promoters, where in the presence of inducer it facilitates formation of a transcription initiation complex. CysB protein also autoregulates its own synthesis by binding to the cysB promoter as a repressor. Cysteine down‐regulates the pathway by inhibiting synthesis of O‐acetylserine, a direct cysteine precursor and possibly an inducer of gene expression. O‐Acetylserine spontaneously isomerizes to N‐acetyl‐serine, which is clearly an inducer. Sulphide and thio‐sulphate provide additional regulation by acting as anti‐inducers. Inducer stimulates CysB protein binding to sites involved in positive regulation, and inhibits binding to the negatively autoregulated cysB promoter. For three sites with unknown function, binding is stimulated at one and inhibited at the other two.
Biological roles of the Escherichia coli RuvA, RuvB and RuvC proteins revealed
Authors: West Stephen C. , Connolly Bernadette 
Published: October 2006

SummaryIn Escherichia coli, the ruvA, ruvB and ruvC gene products are required for genetic recombination and the recombinational repair of DNA damage. New studies suggest that these three proteins function late in recombination and process Holliday junctions made by RecA protein‐mediated strand exchange. In vitro, RuvA protein binds a Holliday junction with high affinity and, together with RuvB (an ATPase), promotes ATP‐dependent branch migration of the junction leading to the formation of heteroduplex DNA. The third protein, RuvC, which acts independently of RuvA and RuvB, resolves recombination intermediates by specific endonucleolytic cleavage of the Holliday junction.
Expression of the Opc protein correlates with invasion of epithelial and endothelial cells by Neisseria meningitidis
Authors: Virji Mumtaz , Makepeace Katherine , Ferguson David J. P. , Achtman Mark , Sarkari Jasmine , Moxon E. Richard 
Published: October 2006

SummaryWhereas capsulate strains of Neisseria meningitidis are dependent on pili for adhesion to human endothelial and epithelial cells, strains which lacked assembled pili and were partially capsule‐deficient adhered to and invaded human endothelial and epithelial cells if they expressed the Opc protein. Bacteria expressing low or undetectable levels of Opc protein failed to adhere to or invade eukaryotic cells. In addition, the presence of OpaAc751 protein on the surface of bacteria did not increase bacterial interactions with host cells. Association of Opc‐expressing bacteria was inhibited by antibodies against Opc. Invasion was dependent on the host‐cell cytoskeletal activity and was inhibited by cytochalasin D. In some cells, infected at the apical surface, bacteria emerging from basal surface were detected by electron microscopy. Opc is found in diverse meningococci and may represent a common virulence factor, which facilitates adherence and invasion by these bacteria.
Transcriptional regulation of the redD transcriptional activator gene accounts for growth‐phase‐dependent production of the antibiotic undecylprodigiosin in Streptomyces coelicolor A3(2)
Authors: Takano E. , Gramajo H. C. , Strauch E. , Andres N. , White J. , Bibb M. J. 
Published: October 2006

SummaryTranscription of redD, the activator gene required for production of the red‐pigmented antibiotic undecylprodigiosin by Streptomyces coelicolor A3(2), showed a dramatic increase during the transition from exponential to stationary phase. The increase in redD expression was followed by transcription of redX, a biosynthetic structural gene, and the appearance of the antibiotic in the mycelium, and coincided with the intracellular appearance of ppGpp. However, ppGpp production elicited either by nutritional shift‐down of, or addition of serine hydroxamate to, exponentially growing cultures had no stimulatory effect on redD transcription. The presence of redD on a multicopy plasmid resulted in elevated levels of the redD transcript and production of redX and undecylprodigiosin during exponential growth; the normal growth‐phase‐dependent production of undecylprodigiosin appeared to be mediated entirely through the redD promoter, which shows limited similarity to the consensus sequence for the major class of eubacterial promoters.
Characterization and functional expression in Escherichia coli of the sodium/proton/glutamate symport proteins of Bacillus stearothermophilus and Bacillus caldotenax
Authors: Tolner Berend , Poolman Bert , Konings Wil N. 
Published: October 2006

SummaryThe genes encoding the Na+/H+/L‐glutamate symport proteins of the thermophilic organisms Bacillus stearothermophilus (gltTBs) aid Bacillus caldotenax (gltTBc) were cloned by complementation of Escherichia coli JC5412 for growth on glutamate as sole source of carbon, energy and nitrogen. The nucleotide sequences of the gltTBs and gltTBc genes were determined. In both cases the translated sequences corresponded with proteins of 421 amino acid residues (96.7% amino acid identity between GltTeB and GltTBc)‐ Putative promoter, terminator and ribosome‐binding‐site sequences were found in the flanking regions. These expression signals were functional in E. coli. The hydropathy profiles indicate that the proteins are hydrophobic and could form 12 membrane‐spanning regions. The Na+/H+ coupled L‐glutamate symport proteins GltTBs and GltTBc are homologous to the strictly H+ coupled L‐glutamate transport protern of E. coli K‐12 (overall 57.2% identity). Functional expression of glutamate transport activity was demonstrated by uptake of glutamate in whole cells and membrane vesicles. In accordance with previous observations (de Vrij etal., 1989; Heyne et al., 1991), glutamate uptake was driven by the electrochemical gradients of sodium ions and protons.
Molecular mechanisms of attachment of Rhizobium bacteria to plant roots
Authors: Smit Gerrit , Swart Saskia , Lugtenberg Ben J. J. , Kijne Jan Willem 
Published: October 2006

SummaryAttachment of bacteria to plant cells is one of the earliest steps in many plant‐bacterium interactions. This review covers the current knowledge on one of the best‐studied examples of bacterium‐plant attachment, namely the molecular mechanism by which Rhizobium bacteria adhere to plant roots. Despite differences in several studies with regard to growth conditions of bacteria and plants and to methods used for measuring attachment, an overall consensus can be drawn from the available data. Rhizobial attachment to plant root hairs appears to be a two‐step process. A bacterial Ca2+‐binding protein, designated as rhicadhesin, is involved in direct attachment of bacteria to the surface of the root hair cell. Besides this step, there is another step which results mainly in accumulation and anchoring of the bacteria to the surface of the root hair. This leads to so‐called firm attachment. Depending on the growth conditions of the bacteria, the latter step is mediated by plant lectins and/or by bacterial appendages such as cellulose fibrils and fimbriae. The possible role of these adhesins in root nodule formation is discussed.
An inverted repeat preceding the Bacillus subtilis glpD gene is a conditional terminator of transcription
Authors: Holmberg Christina , Rutberg Lars 
Published: October 2006

SummaryThe Bacillus subtilis glpD gene, encoding glycerol‐3‐phosphate (G3P) dehydrogenase, is preceded by a promoter and an inverted repeat which is located between the promoter and the glpD coding region. The Inverted repeat acts as a transcriptional terminator in vitro. Expression of glpD is induced by G3P in the presence of the glpD gene product. Full‐length glpD transcripts can be detected only in glycerol‐induced cells. The major glpD transcript is initiated from the glpD promoter but minor amounts of larger transcripts, possibly initiated at upstream glp promoters, can also be found. In uninduced cells short transcripts are present, corresponding to initiation at the glpD promoter and termination at the inverted repeat. Upon induction, these short transcripts disappear and are replaced by full‐length glpD transcripts. The 3′‐ends of full‐length glpD transcripts were mapped to an Inverted repeat located immediately downstream of glpD. These results show that glpD of B. subtilis is regulated by termination/antitermination of transcription.
Determination of a specific region of the purine–cytosine permease involved in the recognition of its substrates
Authors: Bloch J. C. , Sychrova H. , Souciet J. L. , Jund R. , Chevallier M. R. 
Published: October 2006

SummaryThree u.v.‐induced mutants of the purine–cytosine permease gene of Saccharomyces cerevisiae, with altered apparent Michaelis constant of transport (Kmapp), were cloned and sequenced. One of the mutants had extensive nucleotide replacement, whereas the other two had a single mutation. To evaluate the contribution of the different amino acid replacements to the phenotype of the complex mutant, simpler mutants were created by site‐directed mutagenesis. All the amino acid replacements found in the segment from amino acids 371 to 377 inclusive, contribute to the determination of the phenotype. According to the model postulated this segment lies on the cell surface. In particular, amino acids at position 374 and 377 modulate the affinity of the permease towards its substrates. In the wild‐type, when asparagine is present at both of these positions, the lowest Kmapp values are found.