bluuebunny/crossref_metadata_2025_split huggingface dataset:
Recognition of iron‐free siderophores by TonB‐dependent iron transporters
Authors: Schalk Isabelle J. , Yue Wyatt W. , Buchanan Susan K. 
Published: September 2004

SummaryTonB‐dependent iron transporters reside in the outer membranes of Gram‐negative bacteria, transporting ferric‐complexes into the periplasm by a mechanism requiring proton motive force and an integral inner membrane complex, TonB–ExbB–ExbD. Certain TonB‐dependent transporters contain an additional domain at the N‐terminus, which interacts with an inner membrane regulatory protein and a cytoplasmic sigma factor to induce transcription of iron transport genes when a ferric‐ligand is bound at the extracellular surface of the transporter. Transport of the ferric‐ligand is apparently not necessary for transcription induction. Recent biophysical and crystallographic experiments have shown that this subclass of TonB‐dependent iron transporters can bind iron‐free ligands, whereas only the ferric‐ligands are transported into the periplasm. This review focuses on the ligand binding properties of these transporters and includes a discussion of the biological function of the additional domain, the mechanism of transcription induction and the mechanism of ferric‐ligand transport.
Novel substitutions in the σ54‐dependent activator DctD that increase dependence on upstream activation sequences or uncouple ATP hydrolysis from transcriptional activation
Authors: Xu Hao , Kelly Mary T. , Nixon B. Tracy , Hoover Timothy R. 
Published: September 2004

Summary Sinorhizobium meliloti DctD is an activator of σ54‐RNA polymerase holoenzyme and member of the AAA+ superfamily of ATPases. DctD uses energy released from ATP hydrolysis to stimulate the isomerization of a closed promoter complex to an open complex. DctD binds to upstream activation sequences (UAS) and contacts the closed complex through DNA looping to activate transcription, but the UAS is not essential for activation if DctD is expressed at higher than normal levels. Introduction of specific substitutions within or near the conserved ESELFG motif in the C3 region of a truncated, constitutively active form of DctD produced several mutant forms of the protein that had increased dependence on the UAS for activation. Removing the DNA‐binding domain from one UAS‐dependent mutant and from one activation‐deficient mutant significantly increased transcriptional activation, indicating that the DNA‐binding domain interfered with the activities of these mutant proteins. A UAS‐dependent mutant with a P315L substitution in the C6 region was identified from a genetic screen. Alanine scanning mutagenesis of conserved amino acid residues around Pro‐315 produced two additional UAS‐dependent mutants as well as several mutants that failed to activate transcription but retained ATPase activity. In contrast to the two mutant proteins with substitutions in the C3 region, removal of the DNA‐binding domain from the mutant proteins with substitutions in the C6 region did not stimulate their activity. The residues in the C6 region that were altered are in a probable hinge region between the α/β and α‐helical subdomains of the AAA+ domain. The α‐helical subdomain contains the sensor II helix that has been implicated in other AAA+ proteins as sensing changes in the nucleotide during the hydrolysis cycle. Substitutions in the hinge region may have abolished nucleotide sensing by interfering with subdomain interactions, altering the relative orientation of the sensor II helix or interfering with oligomerization of the protein.
Expression of nitrite reductase in Nitrosomonas europaea involves NsrR, a novel nitrite‐sensitive transcription repressor
Authors: Beaumont Hubertus J. E. , Lens Sylvia I. , Reijnders Willem N. M. , Westerhoff Hans V. , Van Spanning Rob J. M. 
Published: September 2004

SummaryProduction of nitric oxide (NO) and nitrous oxide (N2O) by ammonia (NH3)‐oxidizing bacteria in natural and man‐made habitats is thought to contribute to the undesirable emission of NO and N2O into the earth's atmosphere. The NH3‐oxidizing bacterium Nitrosomonas europaea expresses nitrite reductase (NirK), an enzyme that has so far been studied predominantly in heterotrophic denitrifying bacteria where it is involved in the production of these nitrogenous gases. The finding of nirK homologues in other NH3‐oxidizing bacteria suggests that NirK is widespread among this group; however, its role in these nitrifying bacteria remains unresolved. We identified a gene, nsrR, which encodes a novel nitrite (NO2–)‐sensitive transcription repressor that plays a pivotal role in the regulation of NirK expression in N. europaea. NsrR is a member of the Rrf2 family of putative transcription regulators. NirK was expressed aerobically in response to increasing concentrations of NO2– and decreasing pH. Disruption of nsrR resulted in the constitutive expression of NirK. NsrR repressed transcription from the nirK gene cluster promoter (Pnir), the activity of which correlated with NirK expression. Reconstruction of the NsrR‐Pnir system in Escherichia coli revealed that repression by NsrR was reversed by NO2– in a pH‐dependent manner. The findings are consistent with the hypothesis that N. europaea expresses NirK as a defence against the toxic NO2– that is produced during nitrification.
Versatility of the carboxy‐terminal domain of the α subunit of RNA polymerase in transcriptional activation: use of the DNA contact site as a protein contact site for MarA
Authors: Dangi Bindi , Gronenborn Angela M. , Rosner Judah L. , Martin Robert G. 
Published: September 2004

SummaryThe transcriptional activator, MarA, interacts with RNA polymerase (RNAP) to activate promoters of the mar regulon. Here, we identify the interacting surfaces of MarA and of the carboxy‐terminal domain of the α subunit of RNAP (α‐CTD) by NMR‐based chemical shift mapping. Spectral changes were monitored for a MarA‐DNA complex upon titration with α‐CTD, and for α‐CTD upon titration with MarA‐DNA. The mapping results were confirmed by mutational studies and retention chromatography. A model of the ternary complex shows that α‐CTD uses a ‘265‐like determinant’ to contact MarA at a surface distant from the DNA. This is unlike the interaction of α‐CTD with the CRP or Fis activators where the ‘265 determinant’ contacts DNA while another surface of the same α‐CTD molecule contacts the activator. These results reveal a new versatility for α‐CTD in transcriptional activation.
Cell cycle‐dependent abundance, stability and localization of FtsA and FtsQ in Caulobacter crescentus
Authors: Martin Miriam E. , Trimble Michael J. , Brun Yves V. 
Published: September 2004

SummaryCoordination between cell division and DNA replication is ensured by checkpoints that act through proteins required for cell division. Following a block in DNA replication, transcription of the cell division progression genes ftsA and ftsQ is prevented in Caulobacter crescentus. One requirement for this checkpoint is that FtsA and/or FtsQ should be limiting for division in the next cell cycle. We show that the number of FtsA and FtsQ molecules fluctuates such that their concentration is low in swarmer and stalked cells, peaks in pre‐divisional cells, and then dramatically decreases after cell division. Despite constitutive expression from an inducible promoter, FtsA and FtsQ levels still vary during the cell cycle, and the half‐life of FtsA increases from 13 min in swarmer cells to 55 min in stalked cell types, confirming cell type‐specific degradation. The post‐division degradation of FtsA and FtsQ in swarmer cells reduces their concentration to 7% and 10% of their maximal level, respectively, strongly suggesting that de novo synthesis of both proteins is required for each division cycle. The localization of FtsA and FtsQ is also cell type‐specific. FtsA and FtsQ are recruited to the midcell during a short period in late pre‐divisional cells, consistent with the demonstrated requirement of FtsA for late stages of cell division. As previously reported for FtsZ, constitutive expression of FtsA causes cell division defects. These results indicate that the tight control of FtsA, and probably FtsQ, by cell cycle transcription, proteolysis, and localization are critical for optimal cell division and the coordination of cell division with the DNA replication cycle.
Phase variable desialylation of host proteins that bind to Streptococcus pneumoniae in vivo and protect the airway
Authors: King Samantha J. , Hippe Karen R. , Gould Jane M. , Bae Deborah , Peterson Scott , Cline Robin T. , Fasching Claudine , Janoff Edward N. , Weiser Jeffrey N. 
Published: September 2004

SummaryMost clinical isolates of Streptococcus pneumoniae consist of heterogeneous populations of at least two colony phenotypes, opaque and transparent, selected for in the bloodstream and nasopharynx, respectively. Microarray analysis revealed 24 orfs that demonstrated differences in expression greater than twofold between variants of independent strains. Twenty‐one of these showed increased expression in the transparent variants, including 11 predicted to be involved in sugar metabolism. A single genomic region contains seven of these loci including the gene that encodes the neuraminidase, NanA. In contrast to previous studies, there was no contribution of NanA to adherence of S. pneumoniae to epithelial cells or colonization in an animal model. However, we observed NanA‐dependent desialylation of human airway components that bind to the organism and may mediate bacterial clearance. Targets of desialylation included human lactoferrin, secretory component, and IgA2 that were shown to be present on the surface of the pneumococcus in vivo during pneumococcal pneumonia. The efficiency of desialylation was increased in the transparent variants and enhanced for host proteins binding to the surface of S. pneumoniae. Because deglycosylation affects the function of many host proteins, NanA may contribute to a protease‐independent mechanism to modify bound targets and facilitate enhanced survival of the bacterium.
HmsP, a putative phosphodiesterase, and HmsT, a putative diguanylate cyclase, control Hms‐dependent biofilm formation in Yersinia pestis
Authors: Kirillina Olga , Fetherston Jacqueline D. , Bobrov Alexander G. , Abney Jennifer , Perry Robert D. 
Published: September 2004

SummaryThe Hms+ phenotype of Yersinia pestis promotes the binding of haemin or Congo red (CR) to the cell surface at temperatures below 34°C. We previously demonstrated that temperature regulation of the Hms+ phenotype is not controlled at the level of transcription. Instead, HmsH, HmsR and HmsT are degraded upon a temperature shift from 26°C to 37°C. We used random transposon mutagenesis to identify new genes involved in the temperature‐regulated expression of the Hms phenotype. One of these genes, which we designated hmsP, encodes a putative phosphodiesterase with a conserved EAL motif. Mutations in hmsP caused formation of red colonies on CR plates at 26°C and 37°C. Strains complemented with hmsP+ on a plasmid form white colonies at both temperatures. We used a crystal violet assay and confocal laser scanning microscopy to demonstrate Hms‐dependent biofilm formation by Y. pestis cells. Y. pestis Hms+ strains grown at 26°C but not at 37°C form a biofilm on borosilicate glass surfaces. Strains that either overexpress HmsT (a GGDEF domain protein) or have a mutation in hmsP produced an extremely thick biofilm. Alanine substitutions for each of the GGEE residues (amino acids 296–299) of HmsT as well as the E506 and L508 residues of HmsP caused a loss of function. We propose that HmsT and HmsP together control the amount of biofilm produced in Y. pestis. Degradation of HmsT at 37°C may be a critical factor in controlling the temperature‐dependent expression of the Hms biofilm.
Post‐proteomic identification of a novel phage‐encoded streptodornase, Sda1, in invasive M1T1 Streptococcus pyogenes
Authors: Aziz Ramy K. , Ismail Shehab A. , Park Hee‐Won , Kotb Malak 
Published: September 2004

SummaryThe M1T1 strain remains the most frequently isolated strain from group A streptococcal (GAS) infection cases worldwide. We previously reported that M1T1 differs from the fully sequenced M1 SF370 strain. To better understand the reason for the persistence and increased virulence of M1T1, we analysed its secreted proteome and identified two virulence proteins that are not present in the sequenced M1 SF370 strain: streptococcal pyrogenic exotoxin A (SpeA) and a streptodornase D (SdaD) homologue. In the present study, we determined the nucleotide sequence of the M1T1 streptodornase and found that its deduced amino acid sequence is highly similar to other streptococcal streptodornases, and is most closely related to the SdaD of GAS strain M49. M1T1 Sda shares two highly conserved domains with several DNases and putative DNases in streptococci; however, it possesses a unique C‐terminal amino acid sequence. Thus, we named the protein Sda1, and we detected the presence of the sda1 gene in 16 M1T1 clinical isolates. The cloned and expressed Sda1 degrades both streptococcal and mammalian DNA at physiological pH. Amino acid similarity analyses of known GAS deoxyribonucleases suggest that Sda1 may be a chimeric protein created through recombination events. Moreover, a natural mutation that resulted in longer Sda1 and SdaD as compared to other GAS DNases was found to confer increased activity on the protein. Analysis of the sequences flanking sda1 determined that it is carried by a prophage or a prophage‐like element inserted in the tRNA‐Ser gene of M1T1 GAS. Ongoing studies in our laboratory aim to determine the contribution of Sda1 to the virulence of this globally disseminated M1T1 strain.
Widespread distribution of a lexA‐regulated DNA damage‐inducible multiple gene cassette in the Proteobacteria phylum
Authors: Abella Marc , Erill Ivan , Jara Mónica , Mazón Gerard , Campoy Susana , Barbé Jordi 
Published: September 2004

SummaryThe SOS response comprises a set of cellular functions aimed at preserving bacterial cell viability in front of DNA injuries. The SOS network, negatively regulated by the LexA protein, is found in many bacterial species that have not suffered major reductions in their gene contents, but presents distinctly divergent LexA‐binding sites across the Bacteria domain. In this article, we report the identification and characterization of an imported multiple gene cassette in the Gamma Proteobacterium Pseudomonas putida that encodes a LexA protein, an inhibitor of cell division (SulA), an error‐prone polymerase (DinP) and the alpha subunit of DNA polymerase III (DnaE). We also demonstrate that these genes constitute a DNA damage‐inducible operon that is regulated by its own encoded LexA protein, and we establish that the latter is a direct derivative of the Gram‐positive LexA protein. In addition, in silico analyses reveal that this multiple gene cassette is also present in many Proteobacteria families, and that both its gene content and LexA‐binding sequence have evolved over time, ultimately giving rise to the lexA lineage of extant Gamma Proteobacteria.
Identification and characterization of the Pasteurella multocida toxin translocation domain
Authors: Baldwin Michael R. , Lakey Jeremy H. , Lax Alistair J. 
Published: September 2004

SummaryThe Pasteurella multocida toxin (PMT) is a potent mitogen which enters the cytosol of eukaryotic cells via a low pH membrane translocation event. In common with the Escherichia coli cytotoxic necrotizing factor 1 (CNF1), the core of the PMT translocation domain is composed of two predicted hydrophobic helices (H1 – residues 402–423, H2 – 437–457) linked by a hydrophilic loop (PMT‐TL – 424–436). The peptide loop contains three acidic residues (D425, D431 and E434), which may play a role equivalent to D373, D379 and E382/383 in CNF1. To test this hypothesis, a series of point mutants was generated in which acidic residues were mutated into the permanently charged positive residue lysine. Individual mutation of D425, D431 and E434 each caused a four‐ to sixfold reduction in toxin activity. Interestingly, mutation of D401 located immediately outside the predicted helix–loop–helix motif completely abolished toxin activity. Individual mutations did not affect cell binding nor greatly altered toxin structure, but did prevent translocation of the surface‐bound proteins into the cytosol after a low pH pulse. Moreover, we demonstrate using an in vitro assay that PMT undergoes a pH‐dependent membrane insertion.
A network of net‐workers: report of the Euresco conference on ‘Bacterial Neural Networks’ held at San Feliu (Spain) from 8 to 14 May 2004
Authors: Hellingwerf Klaas J. 
Published: September 2004

SummaryIn May 2004, over 100 bacteriologists from 19 different countries discussed recent progress in identification and understanding of individual signal transfer mechanisms in bacteria and in the mutual interactions between these systems to form a functional living cell. The meeting was held in San Feliu and supported by ESF and EMBO. In part through the extensive sequencing efforts of the past few years, the bulk of the bacterial signal transfer systems have been resolved and their detailed characterization is revealing such characteristics as signal specificity, signalling rate constants, molecular interaction affinities, subcellular localization, etc., which should provide a solid basis to a computational extension of this field of studies. In parallel, the new genomics techniques are providing tools to characterize the way a collection of such systems interact in an individual cell, to give rise to ‘life’. Systems theory provides rational and convenient ways to bring order to the wide range of observables thus obtained. Ultimately, the performance of engineered design will have to prove whether or not we know enough about the processes involved.
Clinical Paratuberculosis as a Cause for Higher Culling Rate in Cows with Left Displaced Abomasum and Diarrhoea in a Holstein Dairy Herd
Authors: Kasravi R. , Nowrouzian I. 
Published: September 2004

SummaryThe present report identifies a statistically significant (P < 0.001) higher postoperative culling rate in cows with left displaced abomasum (LDA) exhibiting diarrhoea, compared with cows with LDA and normal faeces from a large Holstein dairy herd (odds ratio = 34). Paratuberculosis was histopathologically confirmed in 77.0% of cows with LDA exhibiting diarrhoea.
Early and Cost‐Effective Identification of High Risk/Priority Control Areas in Foot‐and‐Mouth Disease Epidemics
Authors: Rivas A. L. , Schwager S. J. , Smith S. , Magri A. 
Published: September 2004

SummaryGeo‐referenced data from the 2001 Uruguayan foot‐and‐mouth disease (FMD) epidemic were explored to assess whether spatial analysis could lead to cost‐benefit based policies. Four variables were analysed: (i) location and size of 4022 individual rural land parcels, of which 574 were infected over 60 days, (ii) animal density, (iii) percentage of dairy farms per county, and (iv) road density. Each variable was categorized into two to five classes (e.g. small/medium/large) and the proportion of cases per class reported at days 1–3 of the epidemic was compared with that reported at days 4–6. A higher proportion of cases was found at days 4–6 than at days 1–3 in areas with: small and medium size land parcels, high animal density, >20% farms specialized in dairy production, and high road density (P < 0.03 for each). Each of these classes showed a greater proportion of cases at days 7–60 than the proportion of the total territory covered by each class's area (early case concentration ratios: 1.14–1.37). Land parcel clusters were indicated by Moran's I‐test (P < 0.01). A new region was constructed by intersecting the four spatial classes associated with higher proportions of cases at days 4–6. At days 7–60, this region included 50.4% of all cases and represented 30.6% of the territory under study (final case concentration ratio: 1.65). The final area per case in this region was at least 33% lower and covered at least 45% less territory than any of the four single‐variable approaches. Bio‐statistical, multivariate spatial analysis of early cases may greatly increase the efficiency of epidemiologic policy.
Influence of Methods for Trichinella Detection in Pigs from Endemic and Non‐endemic European Region
Authors: Nöckler K. , Hamidi A. , Fries R. , Heidrich J. , Beck R. , Marinculic A. 
Published: September 2004

SummaryA total of 1401 German and 226 Croatian pigs raised either indoors or outdoors were tested for Trichinella infection by direct and indirect detection methods. A 10 g sample of diaphragm were examined for muscle larvae by the artificial digestion method; the species was determined by multiplex polymerase chain reaction (PCR). For detection of anti‐Trichinella IgG, serum samples diluted 1:100, and meat juice samples diluted 1:10, were tested by enzyme‐linked immunosorbent assay. All German pigs and those Croatian pigs raised indoors proved to be Trichinella‐negative by all methods. Muscle larvae were detected in a total of eleven of the Croatian pigs, which were raised on small outdoor farms. For eight isolates, PCR results demonstrated that recovered larvae were Trichinella spiralis. Anti‐Trichinella‐IgG was detected in serum and meat juice of digestion positive animals when the worm burdens exceeded 0.38 larvae per gram of muscle. Positive results in Croatian pigs indicate a higher risk of infection for outdoor farming in areas where Trichinella is endemic. Results of direct and indirect detection were compared and are discussed with special regard to specificity and sensitivity of methods.
Splenectomy in severe haemophilia
Authors: Caldicott David , Bonnin Robert , Lloyd John , Day Peter 
Published: September 2004

‘I am disposed to regard the following rule as imperative: that no cutting operation be undertaken unless the patient's life be in danger for want of that particular operation.’−A Treatize on Haemophilia Sometimes Called the Hereditary Haemorrhagic Diathesis J. W. Legg, 18721
The management of any surgical condition in a haemophilia patient is a challenging problem for the surgeon. It is particularly difficult if the patient presents in extremis, with no apparent cause for their collapse. We report a case of successful management of spontaneous splenic rupture in a severe haemophiliac, and review the literature associated with this unusual condition.