bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular analysis of the virulence locus of the Salmonella dublin plasmid pSDL2
Authors: Krause M. , Roudier C. , Fierer J. , Harwood J. , Guiney D. 
Published: October 2006

SummaryThe virulence properties of various non‐typhoid Salmonella serotypes depend on the presence of large plasmids 60–100 kb in size. We have shown previously that the virulence region on the 80 kb plasmid pSDL2 of Salmonella dublinLane maps within a 14kb SalI fragment. In this report we show that an 8.2kb region within this fragment is sufficient to express lethal disease in BALB/c mice. Sequence analysis of this segment revealed six sequential open reading frames designated vsdA–F, which encode putative proteins of 13–65kDa. Deletion analysis and location of Tn5‐oriT inserts which abolish virulence suggest that vsdA, vsdC, vsdD and vsdE are essential for virulence expression. Downstream of vsdF we discovered a locus involved in stable plasmid maintenance. Deletion of that region resulted in plasmid multimerization and instability.
The TraM protein of plasmid R1 is a DNA‐binding protein
Authors: Schwab M. , Gruber H. , Högenauer G. 
Published: October 2006

SummaryThe TraM protein of the resistance plasmid R1 was purified to homogeneity and used for DNA‐binding studies. Both gel retardation‐and footprint experiments showed that TraM specifically binds to DNA of plasmid R1 comprising the region between the origin of transfer and the traM gene. Several TraM molecules bind and, according to the footprint experiments, two distinct sites of specific binding exist The two sites are separated from each other by 12 nucleotides and each contains an inverted repeat. DNase I protection assays showed that the initial TraM binding occurs at these palindromic sequences. At higher protein concentrations the lengths of the DNA segments protected by TraM were increased towards the traM gene. In one region this extension leads to binding of TraM protein at its own promoters.
The secretion genes of Pseudomonas aeruginosa alkaline protease are functionally related to those of Erwinia chrysanthemi proteases and Escherichia coliα‐haemolysin
Authors: Guzzo J. , Duong F. , Wandersman C. , Murgier M. , Lazdunski A. 
Published: October 2006

SummaryThe extracellular alkaline protease produced by Pseudomonas aeruginosa is secreted by a specific pathway, independent of the pathway used by most of the other extracellular proteins of this organism. Secretion of this protease is dependent on the presence of several genes located adjacent to the apr gene. Complementation studies have shown that PrtD, E, and F, the three secretion functions for Erwinia chrysanthemi proteases B and C (Létofféet al., 1990), can mediate the secretion of the alkaline protease by Escherichia coli. he secretion functions involved in α‐haemloysin secretion in E. coli(hlyB, hlyD, tolC) can also be used to complement alkaline protease secretion by E. coli, although less efficiently. These data indicate that protease secretion mechanisms in Pseudomonas and Erwinia are very similar and are homologous to that of E. coliα‐haemolysin.
The trpC gene of Phanerochaete chrysosporium is unique in containing an intron but nevertheless maintains the order of functional domains seen in other fungi
Authors: Schrank A. , Tempelaars C. , Sims P. F. G. , Oliver S. G. , Broda P. 
Published: October 2006

SummaryThe Phanerochaete chryososporium trpC gene has been isolated by complementation of an Escherichia coli trpC mutant. The full extent of the fungal gene, determined by sequence analysis, was found to be 2414bp. This includes a single intron of 50bp, the presence of which was confirmed by RNA‐primed polymerase chain reaction analysis. This feature makes the P. chrysosporium gene unique when compared to equivalent genes from other filamentous fungi. The P. chrysosporium trpC gene encodes a single protein containing three enzyme activities involved in tryptophan biosynthesis arranged in the order: NH2–GAT–IGPS–PRAI–COOH. This order is conserved in all filamentous fungi so far examined and, indeed, is the gene order within the E. coli trp operon.
The role of the ‘gearbox’ in the transcription of essential genes
Authors: Vicente M. , Kushner S. R. , Garrido T. , Aldea M. 
Published: October 2006

SummaryRegulation of transcription occurs at different levels, one being in the presence of sequences specifically recognized by different forms of RNA polymerase, i.e. the promoters. Three different kinds of promoter are defined according, among other things, to their dependence on the growth rate of the cell: the house‐keeper' promoter of many metabolic genes, the stringent promoter found at several rRNA and ribosomal protein genes, and the ‘gearbox’ at genes whose products are required at higher relative amounts at lower growth rates. The identified gearbox promoters of Escherichia coli share specific homologies in the ‐10, ‐35 and upstream regions. Although there may be different types of gearbox promoters, the ‐10 sequence of one of these promoters has been found to be essential for functioning as a gearbox. This suggests the existence of specific sigma factors for its transcription. RpoS (KatF) is a likely candidate for being one of these sigma factors. Computer simulation allows us to predict that such sigma factors should, in turn, be expressed following a gearbox mode, which would then imply the existence of self‐regulated loops contributing to the expression of some genes of bacterial division. Some bacterial gene products need to be synthesized al fixed amounts per cell
Occurrence and biological function of cytochrome P450 monooxygenases in the actinomycetes
Authors: O'Keefe D. P. , Harder P. A. 
Published: October 2006

SummaryMany species within the order Actinomycetales contain one or more soluble cytochrome P450 monooxygenases, often substrate‐inducible and responsible for a variety of xenobiotic transformations. The individual cytochromes exhibit a relatively broad substrate specificity, and some strains have the capacity to synthesize large amounts of the protein(s) to compensate for low catalytic turnover with some substrates. All three of the Streptomyces cytochromes sequenced to date are exclusive members of one P450 family, CYP105. In several instances, monooxygenase activity arises from induction of a P450 and associated ferredoxin, or of a P450 only, suggesting that some essential electron donor proteins (reductase and ferredoxin) are not coordinately regulated with the cytochrome. The overall properties of these systems suggest an adaptive strategy whose twofold purpose is to maintain a competitive advantage via the production of secondary metabolites, and, whenever possible, to utilize unusual growth substrates by introducing metabolites from these reactions into the more substrate‐specific primary metabolic pathways.
IIIegitimate integration of non‐replicative vectors in the genome of Rhodococcus fascians upon electro‐transformation as an insertional mutagenesis system
Authors: Desomer J. , Crespi M. , Van Montagu M. 
Published: October 2006

SummaryElectrotransformation of Rhodococcus fascians by non‐replicating plasmids containing a suitable resistance marker resulted in stable transformants by integration of these constructs at various sites in the genome, thereby generating different mutations. Tagged genes could be isolated in Escherichia coli owing to the presence of a CoIE1 replicon and an ampicillin resistance gene in the inserted sequences.Southern analysis and nucleotide sequencing revealed that recombination can occur at defined locations in the plasmid, while no site preference for target sequences could be detected. Low homology between the recombining sequences indicates illegitimate recombination. The specificity of the plasmid sites could be explained by assuming a linear recombination intermediate, generated by cleavage of the transformed plasmid.
Characterization of two insertion sequences, IS701 and IS702, from the cyanobacterium Calothrix species PCC 7601
Authors: Mazel D. , Bernard C. , Schwarz R. , Castets A. M. , Houmard J. , de Marsac N. Tandeau 
Published: October 2006

SummaryWe describe the characterization of two insertion elements, IS701 and IS702, isolated from Calothrix species PCC 7601. These insertion elements were cloned from spontaneous pigmentation mutants. Both show the characteristics of typical bacterial insertion sequences, i.e. they present long terminal inverted repeats and they duplicate target DNA upon insertion. These elements share no homology with the only other cyanobacterial insertion sequence described so far, IS891. At least 15 copies of IS701 and 9 copies of IS702 were detected by hybridization experiments in the Calothrix 7601 genome. Their occurrence in several cyanobacterial strains is also reported.
In vivo analysis of integration of membrane proteins in Escherichia coli
Authors: Ito K. , Akiyama Y. 
Published: October 2006

SummaryThe in vivo process of membrane protein integration was studied by pulse‐labelling Escherichia coli cells, and assessing integral anchoring of labelled proteins to the lipid bilayer based on their resistance to alkali extraction. To conduct this experiment, conditions for extracting E. coli proteins with alkali were refined, and the immunoprecipitation procedures were improved to allow effective detection of integral membrane proteins. Examination of pulse‐labelled, integral membrane proteins, including lactose permease (LacY), SecY, cytochrome o subunit II and leader peptidase revealed that all were in the alkali‐insoluble fraction, indicating that membrane integration of these proteins takes place rapidly in wild‐type cells. However, when LacY was synthesized in excess from a multicopy plasmid, significant proportions were found in the alkali‐soluble fraction, indicating that the solubility in alkali is not an intrinsic property of the protein, and suggesting that LacY depends on some limited cellular factor for membrane integration. The unintegrated species of LacY sedimented slowly through an alkaline sucrose gradient. The secY24 mutant cells accumulated higher proportions of unintegrated LacY molecules at lower levels of overproduction than the sec+ cells. LacY overproduction in wild‐type cells was found to inhibit processing (export) of β‐lactamase but not of OmpA and OmpF. These results are interpreted to mean that integration of LacY depends on multiple cellular components, one of which is also involved in export of β‐lactamase.
Membrane topology of the integral membrane components, OppB and OppC, of the oligopeptide permease of Salmonella typhimurium
Authors: Pearce S. R. , Mimmack M. L. , Gallagher M. P. , Gileadi U. , Hyde S. C. , Higgins C. F. 
Published: October 2006

SummaryThe oligopeptide permease of Salmonella typhimurium is a periplasmic binding protein‐dependent transport system. Five gene products, OppABCDF, are required for the functioning of this transporter, two of which (OppB and OppC) are highly hydrophobic, integral membrane proteins and are responsible for mediating passage of peptides across the cytoplasmic membrane. OppB and OppC are each predicted, from their sequences, to span the membrane many times. In this paper we describe experimental evidence confirming these predictions using a combination of biochemical, immunological and genetic procedures. Each of these two proteins is shown to span the membrane six times, with the N‐ and C‐termini both being located at the cytoplasmic face of the membrane. Opp is apparently a typical member of the ABC (ATP‐binding cassette) superfamily of transporters. These findings, therefore, have general implications for the organization and function of other ABC transporters, including the human multidrug resistance protein and the product of the cystic fibrosis gene.
The copy number of plasmid pLS1 is regulated by two trans‐acting plasmid products: the antisense RNA II and the repressor protein, RepA
Authors: del Solar G. , Espinosa M. 
Published: October 2006

SummaryThe promiscuous plasmid pLS1 encodes two transacting elements that regulate its copy number: protein RepA and antisense RNA II. In vitro transcription showed that RNAs for both repressors are synthesized from two promoters, PAB and PII From PAB, genes encoding RepA (transcriptional repressor) and RepB (initiator of replication) are cotranscribed, the target of RepA being located within PAB Mutants in repA or in PAB are still sensitive to RepA. However, cloning of the repA gene in a compatible replicon did not result in incompatibility towards pLS1. From PII, the 50‐nucleotide RNA II is synthesized. The main incompatibility determinant towards pLS1 corresponds to the coding sequence for RNA II. The RNA II target could be reduced to 21 nucleotides, including the RepB initiation of translation signals. We propose that plasmids of the pLS1 family (pE194, pADB201, and pLB4) share functional and structural characteristics for the regulation of their copy numbers.
Inducible DNA repair and differentiation in Bacillus subtilis: interactions between global regulons
Authors: Yasbin Ronald E. , Cheo David L. , Bayles Ken W. 
Published: October 2006

SummaryThe SOS response of Escherichia coli has become a paradigm for the study of inducible DNA repair and recombination processes in many different organisms. While these studies have demonstrated that the components of the SOS response appear to be highly conserved among bacterial species, as with most models, there are some significant variations. Perhaps the best example of this comes from an analysis of the SOS‐like system of the developmental organism. Bacillus subtilis. Accordingly, the most striking difference is the complex developmental regulation of the SOS system as this organism differentiates into its competent state. In this review we have given an overview of the elements that comprise the SOS system of B. subtilis. Additionally, we have summarized our most recent findings regarding the regulation of this regulon. Using these results along with new findings from other laboratories we have provided provocative molecular models for the regulation of the B. subtilis SOS system in response to DNA damage and during competent cell formation.
Molecular analysis of the rfb gene cluster of Salmonella serovar muenchen (strain M67): the genetic basis of the polymorphism between groups C2 and B
Authors: Brown P. K. , Romana L. K. , Reeves P. R. 
Published: October 2006

SummaryThe rfb (O antigen) gene cluster of group C2 Salmonella differs from that of group B in a central region of 12.4kb: we report the sequencing of this region of strain M67 (group C2) and a subsequent comparison with the central region of strain LT2 (group B). We find a block of seven open reading frames unique to group C2 which encode the O antigen polymerase (rfc) and the transferases responsible for assembly of the group C2 O antigen. The remaining rfb genes are common to strains M67 and LT2, but rfbJ (GDP‐abequose synthase) and rfbM and rfbK (GDP‐mannose synthesis), which are immediately adjacent to the central region, are highly divergent. All these genes have a low G+C content and appear to have been recent additions to Salmonella enterica. We discuss the evolutionary significance of the arrangement and divergence of the genes in the polymorphism of the rfb cluster.
Involvement of the Arc system in redox regulation of the Cob operon in Salmonella typhimurium
Authors: Andersson Dan I. 
Published: October 2006

SummaryTranscription of the cobalamin biosynthetic (cob) genes in Salmonella typhimurium is induced during anaerobic growth. Two mutants defective in induction of the Cob operon were isolated. The mutants showed decreased anaerobic induction, both during conditions of fermentation and anaerobic respiration, relative to the wild type. The mutant genes were localized by Hfr and transduction mapping to 100 minutes and 69 minutes on the chromosome. The mutants were sensitive to The dye toluidine blue. The mapping and the dye sensitivity suggest that these mutants are defective in The arcA and arcB genes, not previously identified in Salmonella typhimurium. Thus The regulatory function of the Arc system is expanded to include anaerobic activation of The Cob operon.
AIDA‐I, the adhesin involved in diffuse adherence of the diarrhoeagenic Escherichia coli strain 2787 (O126:H27), is synthesized via a precursor molecule
Authors: Benz Inga , Schmidt M. Alexander 
Published: October 2006

SummaryThe adherence mechanisms of enteropathogenic Escherichia coli (EPEC) to epithelial cells are still not understood. To study the molecular basis of the diffuse adherence (DA) phenotype exhibited by diarrhoeagenic E. coli expressing classical EPEC serotypes we investigated strain 2787 (O126:H27) isolated from a case of infantile diarrhoea. A 6.0 kb plasmid‐derrved DNA fragment mediates the DA phenotype and encodes the 100 kDa adhesin protein AIDA‐I (adhesin involved in diffuse adherence). Sequencing of the entire fragment revealed two open reading frames which encoded proteins of 45 kDa and 132 kDa, respectively. The 132 kDa protein has been identified as an AIDA‐I precursor protein. After cleavage of the signal sequence further processing at the C‐terminus of the 132 kDa precursor leads to the mature ∼100 kDa AIDA‐I. While the exact function of the cytopiasmic 45 kDa protein is not known, preliminary evidence indicates that it is necessary for the correct maturation of AIDA‐I. The AIDA‐l precursor exhibits significant homology with the virG(icsA) protein of Shigella flexneri which seems to be involved in the intercellular spread of invasive Shigella organisms.
PCR‐amplified length polymorphisms in tRNA intergenic spacers for categorizing staphylococci
Authors: Welsh John , McClelland Michael 
Published: October 2006

SummaryThe intergenic spacers between some adjacent tRNA genes were shown to be polymorphic in length when closely related Staphylococcus species were compared. A simple procedure was developed to detect and sequence these tRNA intergenic length polymorphisms (tRNA‐ILPs). A comparison of homologous tRNA gene sequences flanking these ILPs in three Staphylococcus species was used to derive primers for high‐stringency amplification of the ILPs by the polymerase chain reaction (PCR). The detection of tRNA‐ILPs by PCR allowed the classification of virtuatly all strains from the five species of Staphylococcus that were examined. The procedure used to identify, sequence and derive primers for PCR detection of tRNA‐ILPs in Staphylococcus should be applicable to many other genera of eubacteria. These primers could be used on uncultured material such as clinical samples.
Early events in yeast mitochondrial protein targeting
Authors: Verner Keith 
Published: October 2006

SummaryProtein import into mitochondria involves a number of complex steps occurring in the cytosol, on the mitochondrial surface, and inside the organelle. Once an initial interaction between mitochondrial proteins and their specific receptors occurs, the proteins are transported into the organelle in a series of reactions involving (in the case of a protein to be translocated into the mitochondrial matrix) the mitochondrial membrane potential, ATP hydrolysis and an undetermined number of membrane components. Inside the organelle, mitochondrial proteins are processed and sorted to their final intramitochondrial destinations. The earliest steps in the import process take place in the cytosol and include the synthesis of the mitochondrial proteins themselves, their interaction with cytosolic factors, and perhaps the establishment of cotranslational import complexes on the mitochondrial surface. These early events are important because it is during this phase that the system as a whole is most sensitive to cytosolic conditions that may exert control over the entire import process.
Differential regulation of two genes encoding lysyl‐tRNA synthetases in Escherichia coli: lysU‐constitutive mutations compensate for a lysS null mutation
Authors: Kawakami K. , Ito K. , Nakamura Y. 
Published: October 2006

SummaryLysyl‐tRNA synthetases are synthesized in Escherichia coli from two distinct genes, lysS and lysU, which are regulated differentially. A strain which is null for lysS, the constitutive gene, was created by gene disruption (lysS1) and exhibited cold‐sensitive lethality. Hence, lysS is dispensable at high temperatures. This cold sensitivity was suppressed by a multi‐copy plasmid carrying lysU, the inducible gene. These data are interpreted as indicating that lysS is functionally replaceable by lysU for cell growth, and that the cold sensitivity of lysS1 is caused by insufficient expression of lysU at low temperatures. To investigate the mechanism of lysU expression, cold‐resistant bypass mutations were isolated from lysS1, and named als (for abandonment of lysS.). Two als mutations which were linked to lysU contain IS2 insertions upstream of the lysU promoter. They caused a 16–19‐fold increase in the lysU‐mRNA level. Furthermore, deletion mutations created immediately upstream of the lysU promoter restored growth of lysS1. These results suggest that transcription of lysU is negatively controlled by a cis‐element located upstream of the promoter.
PhoA gene fusions in Legionella pneumophila generated in vivo using a new transposon, MudphoA
Authors: Albano Mark A. , Arroyo Juan , Eisenstein Barry I. , Engleberg N. Cary 
Published: October 2006

SummaryTo enable effective use of phoA gene fusions in Legionella pneumophila, we constructed MudphoA, a derivative of the mini‐Mu phage Mu dII4041, which is capable of generating gene fusions to the Escherichia coli alkaline phosphatase gene (EC 3.1.3.1). Although an existing fusion‐generating transposon, TnphoA, has been a useful tool for studying secreted proteins in other bacteria, this transposon and other Tn5 derivatives transpose inefficiently in Legionella pneumophila, necessitating the construction of a more effective vector for use in this pathogen. Using MudphoA we generated fusions to an E. coli gene encoding a periplasmic protein and to an L. pneumophila gene encoding an outer membrane protein; both sets of fusions resulted in alkaline phosphatase activity. We have begun to use MudphoA to mutate secreted proteins of L. pneumophila specifically, since this subset of bacterial proteins is most likely to be involved in host‐bacterial interactions. This modified transposon may be useful for studies of other bacteria that support transposition of Mu, but not Tn5, derivatives.
Identification and molecular analysis of glgS, a novel growth‐phase‐regulated and rpoS‐dependent gene involved in glycogen synthesis in Escherichia coli
Authors: Hengge‐Aronis Regjne , Fischer Daniela 
Published: October 2006

SummaryThe putative stationary‐phase sigma factor (σS) encoded by rpoS is essential for giycogen synthesis, but is not required for the transcription of glgC and glgA, which encode ADP‐glucose‐pyrophosphorylase and glycogen synthase, respectively. Using a mini‐Mu random chromosomal library and a screen for glycogen overproduction, we identified a novel gene (glgS) involved in glycogen synthesis. glgS maps at 66.6 min (3247 kb) on the chromosome and constitutes a mono‐cistronic operon. It encodes a hydrophilic and highly charged small protein, with a molecular weight of 7886, which is strongly expressed in minicells. Experiments with single‐copy chromosomal glgS:: lacZ gene fusions indicated that glgS expression is controlled by σS as well as by cAMP. Two transcriptional start sites were mapped in the upstream regulatory region of glgS. The glgS p1 transcript was absent in a cya mutant, whereas an rpoS mutant did not synthesize the glgS p2 transcript. Although glycogen synthesis is strongly stimulated by overproduction of GlgS and is inhibited by a glgS null mutation, glgS does not affect the expression of the glgCAP operon. Its potential role in the metabolic control of glycogen synthesis is discussed. Also, evidence is presented to show that the amount of glycogen accumulated in vivo in early stationary‐phase cells is mainly determined by σS‐controlled gene expression and allosteric activation of GlgC, whereas the absolute levels of expression of glgCAP as well as the intracellular concentration of cAMP are of minor importance.