bluuebunny/crossref_metadata_2025_split huggingface dataset:
Development and Implementation of a Single-Chain Fv Antibody for Specific Detection of
Bacillus anthracis
Spores
Authors: Mechaly A. , Zahavy E. , Fisher M. 
Published: October 2007

ABSTRACT

A single-chain Fv (scFv) antibody was developed and applied for efficient and specific detection of
Bacillus anthracis
spores. The antibody was isolated from a phage display library prepared from spleens of mice immunized with a water-soluble extract of the outer membrane of the
B. anthracis
spore (exosporium). The library (7 × 10
6
PFU) was biopanned against live, native
B. anthracis
ATCC Δ14185 spores suspended in solution, resulting in the isolation of a unique soluble scFv antibody. The antibody was affinity purified and its affinity constant (3 × 10
8
± 1 × 10
8
M
−1
) determined via flow cytometry (FCM). Preliminary characterization of scFv specificity indicated that the scFv antibody does not cross-react with representatives of some phylogenetically related
Bacillus
spores. The potential use of scFv antibodies in detection platforms was demonstrated by the successful application of the soluble purified scFv antibody in enzyme-linked immunosorbent assays, immunofluorescence assays, and FCM.
Riboflavin Biosynthesis Is Associated with Assimilatory Ferric Reduction and Iron Acquisition by
Campylobacter jejuni
Authors: Crossley Rachel A. , Gaskin Duncan J. H. , Holmes Kathryn , Mulholland Francis , Wells Jerry M. , Kelly David J. , van Vliet Arnoud H. M. , Walton Nicholas J. 
Published: October 2007

ABSTRACT

One of the pathways involved in the acquisition of the essential metal iron by bacteria involves the reduction of insoluble Fe
3+
to soluble Fe
2+
, followed by transport of Fe
2+
to the cytoplasm. Flavins have been implicated as electron donors in this poorly understood process. Ferrous iron uptake is essential for intestinal colonization by the important pathogen
Campylobacter jejuni
and may be of particular importance under low-oxygen conditions. In this study, the links among riboflavin biosynthesis, ferric reduction, and iron acquisition in
C. jejuni
NCTC11168 have been investigated. A riboflavin auxotroph was generated by inactivation of the
ribB
riboflavin biosynthesis gene (Cj0572), and the resulting isogenic
ribB
mutant only grew in the presence of exogenous riboflavin or the riboflavin precursor diacetyl but not in the presence of the downstream products flavin adenine dinucleotide and flavin mononucleotide. Riboflavin uptake was unaffected in the
ribB
mutant under iron-limited conditions but was lower in both the wild-type strain and the
ribB
mutant under iron-replete conditions. Mutation of the
fur
gene, which encodes an iron uptake regulator of
C. jejuni
, resulted in an increase in riboflavin uptake which was independent of the iron content of the medium, suggesting a role for Fur in the regulation of the as-yet-unknown riboflavin transport system. Finally, ferric reduction activity was independent of iron availability in the growth medium but was lowered in the
ribB
mutant compared to the wild-type strain and, conversely, increased in the
fur
mutant. Taken together, the findings confirm close relationships among iron acquisition, riboflavin production, and riboflavin uptake in
C. jejuni
.
Identification of New Kinetoplast DNA Replication Proteins in Trypanosomatids Based on Predicted S-Phase Expression and Mitochondrial Targeting
Authors: Li Yue , Sun Yu , Hines Jane C. , Ray Dan S. 
Published: October 2007

ABSTRACT

Trypanosomatid parasites contain an unusual form of mitochondrial DNA (kinetoplast DNA [kDNA]) consisting of a catenated network of several thousand minicircles and a smaller number of maxicircles. Many of the proteins involved in the replication and division of kDNA are likely to have no counterparts in other organisms and would not be identified by similarity to known replication proteins in other organisms. A new kDNA replication protein conserved in kinetoplastids has been identified based on the presence of posttranscriptional regulatory sequences associated with S-phase gene expression and predicted mitochondrial targeting. The
Leishmania major
protein P105 (
Lm
P105) and
Trypanosoma brucei
protein P93 (
Tb
P93) localize to antipodal sites flanking the kDNA disk, where several other replication proteins and nascent minicircles have been localized. Like some of these kDNA replication proteins, the
Lm
P105 protein is only present at the antipodal sites during S phase. RNA interference (RNAi) of
Tb
P93 expression resulted in a cessation of cell growth and the loss of kDNA. Nicked/gapped forms of minicircles, the products of minicircle replication, were preferentially lost from the population of free minicircles during RNAi, suggesting involvement of
Tb
P93 in minicircle replication. This approach should allow the identification of other novel proteins involved in the duplication of kDNA.
The Ccr4-Not Complex Regulates Skn7 through Srb10 Kinase
Authors: Lenssen Eve , Azzouz Nowel , Michel Agnès , Landrieux Emilie , Collart Martine A. 
Published: October 2007

ABSTRACT

The Ccr4-Not complex is a multifunctional regulatory platform composed of nine subunits that controls diverse cellular events including mRNA degradation, protein ubiquitination, and transcription. In this study, we identified the yeast
Saccharomyces cerevisiae
osmotic and oxidative stress transcription factor Skn7 as a new target for regulation by the Ccr4-Not complex. Skn7 interacts with Not1 in a two-hybrid assay and coimmunoprecipitates with Not5 in a Not4-dependent manner. Skn7-dependent expression of
OCH1
and Skn7 binding to the
OCH1
promoter are increased in
not4
Δ or
not5
Δ mutants. Skn7 purified from wild-type cells but not from
not4
Δ cells is associated with the Srb10 kinase. This kinase plays a central role in the regulation of Skn7 by Not4, since increased
OCH1
expression in
not4
Δ cells requires Srb10. These results reveal a critical role for the Ccr4-Not complex in the mechanism of activation of Skn7 that is dependent upon the Srb10 kinase.
Involvement of Sensor Kinases in the Stress Tolerance Response ofStreptococcus mutans
Authors: Biswas Indranil , Drake Laura , Erkina Dasha , Biswas Saswati 
Published: October 2007

ABSTRACTThe gram-positive bacteriumStreptococcus mutansis the primary causative agent in the formation of dental caries in humans. The ability ofS. mutansto adapt and to thrive in the hostile environment of the oral cavity suggests that this cariogenic pathogen is capable of sensing and responding to different environmental stimuli. This prompted us to investigate the role of two-component signal transduction systems (TCS), particularly the sensor kinases, in response to environmental stresses. Analysis of the annotated genome sequence ofS. mutansindicates the presence of 13 putative TCS. Further bioinformatics analysis in our laboratory has identified an additional TCS in the genome ofS. mutans. We verified the presence of the 14 sensor kinases by using PCR and Southern hybridization in 13 differentS. mutansstrains and found that not all of the sensor kinases are encoded by each strain. To determine the potential role of each TCS in the stress tolerance ofS. mutansUA159, insertion mutations were introduced into the genes encoding the individual sensor kinases. We were successful in inactivating all of the sensor kinases, indicating that none of the TCS are essential for the viability ofS. mutans. The mutantS. mutansstrains were assessed for their ability to withstand various stresses, including osmotic, thermal, oxidative, and antibiotic stress, as well as the capacity to produce mutacin. We identified three sensor kinases, Smu486, Smu1128, and Smu1516, which play significant roles in stress tolerance ofS. mutansstrain UA159.
Genomic Characterization of
Ralstonia solanacearum
Phage φRSA1 and Its Related Prophage (φRSX) in Strain GMI1000
Authors: Fujiwara Akiko , Kawasaki Takeru , Usami Shoji , Fujie Makoto , Yamada Takashi 
Published: October 2007

ABSTRACT

φRSA1 is a wide-host-range bacteriophage isolated from
Ralstonia solanacearum
. In this study, the complete nucleotide sequence of the φRSA1 genomic DNA was determined. The genome was 38,760 bp of double-stranded DNA (65.3% G+C) with 19-bp 5′-extruding cohesive ends (
cos
) and contained 51 open reading frames (ORFs). Two-thirds of the φRSA1 genomic region encodes the phage structural modules, and they are very similar to those reported for coliphage P2 and P2-like phages. A φRSA1 minireplicon with an 8.2-kbp early-expressing region was constructed. A late-expression promoter sequence motif was predicted for these φRSA1 genes as 5′ TGTTGT-(X)
13
-ACAACA. The genomic sequence similarity between φRSA1 and related phages φ52237 and φCTX was interrupted by three AT islands, one of which contained an insertion sequence element, suggesting that they were recombinational hot spots. φRSA1 was found to be integrated into at least three different strains of
R. solanacearum
, and the chromosomal integration site (
attB
) was identified as the 3′ portion of the arginine tRNA(CCG) gene. In the light of the φRSA1 gene arrangement, one possible prophage sequence previously detected on the chromosome of
R. solanacearum
strain GMI1000 was characterized as a φRSA1-related prophage (designated φRSX). φRSX was found to be integrated at the serine tRNA (GGA) gene as an
att
site, and its size was determined to be 40,713 bp. φRSX ORFs shared very high amino acid identity with their φRSA1 counterparts. The relationships and evolution of these P2-like phages are discussed.
Purification and Characterization of Serine Racemase from a Hyperthermophilic Archaeon,Pyrobaculum islandicum
Authors: Ohnishi Masato , Saito Makoto , Wakabayashi Sadao , Ishizuka Morio , Nishimura Katsushi , Nagata Yoko , Kasai Sabu 
Published: October 2007

ABSTRACTPyrobaculum islandicumis an anaerobic hyperthermophilic archaeon that is most active at 100°C. A pyridoxal 5′-phosphate-dependent serine racemase called Srr was purified from the organism. The correspondingsrrgene was cloned, and recombinant Srr was purified fromEscherichia coli. It showed the highest racemase activity towardl-serine, followed byl-threonine,d-serine, andd-threonine. Like rodent and plant serine racemases, Srr is bifunctional, showing highl-serine/l-threonine dehydratase activity. The sequence of Srr is 87% similar to that ofPyrobaculum aerophilumIlvA (a putative threonine dehydratase) but less than 32% similar to any other serine racemases and threonine dehydratases. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration analyses revealed that Srr is a homotrimer of a 44,000-molecular-weight subunit. Both racemase and dehydratase activities were highest at 95°C, while racemization and dehydration were maximum at pH 8.2 and 7.8, respectively. Unlike other, related Ilv enzymes, Srr showed no allosteric properties: neither of these enzymatic activities was affected by eitherl-amino acids (isoleucine and valine) or most of the metal ions. Only Fe2+and Cu2+caused 20 to 30% inhibition and 30 to 40% stimulation of both enzyme activities, respectively. ATP inhibited racemase activity by 10 to 20%. TheKmandVmaxvalues of the racemase activity of Srr forl-serine were 185 mM and 20.1 μmol/min/mg, respectively, while the corresponding values of the dehydratase activity ofl-serine were 2.2 mM and 80.4 μmol/min/mg, respectively.
Genetic and Proteomic Analyses of a Proteasome-Activating Nucleotidase A Mutant of the Haloarchaeon
Haloferax volcanii
Authors: Kirkland P. Aaron , Gil Malgorzata A. , Karadzic Ivanka M. , Maupin-Furlow Julie A. 
Published: October 2007

ABSTRACT

The halophilic archaeon
Haloferax volcanii
encodes two related proteasome-activating nucleotidase proteins, PanA and PanB, with PanA levels predominant during all phases of growth. In this study, an isogenic
panA
mutant strain of
H. volcanii
was generated. The growth rate and cell yield of this mutant strain were lower than those of its parent and plasmid-complemented derivatives. In addition, a consistent and discernible 2.1-fold increase in the number of phosphorylated proteins was detected when the
panA
gene was disrupted, based on phosphospecific fluorescent staining of proteins separated by 2-dimensional gel electrophoresis. Subsequent enrichment of phosphoproteins by immobilized metal ion and metal oxide affinity chromatography (in parallel and sequentially) followed by tandem mass spectrometry was employed to identify key differences in the proteomes of these strains as well as to add to the restricted numbers of known phosphoproteins within the
Archaea
. In total, 625 proteins (approximately 15% of the deduced proteome) and 9 phosphosites were identified by these approaches, and 31% (195) of the proteins were identified by multiple phosphoanalytical methods. In agreement with the phosphostaining results, the number of identified proteins that were reproducibly exclusive or notably more abundant in one strain was nearly twofold greater for the
panA
mutant than for the parental strain. Enriched proteins exclusive to or more abundant in the
panA
mutant (versus the wild type) included cell division (FtsZ, Cdc48), dihydroxyacetone kinase-linked phosphoenolpyruvate phosphotransferase system (EI, DhaK), and oxidoreductase homologs. Differences in transcriptional regulation and signal transduction proteins were also observed, including those differences (e.g., OsmC and BolA) which suggest that proteasome deficiency caused an up-regulation of stress responses (e.g., OsmC versus BolA). Consistent with this, components of the Fe-S cluster assembly, protein-folding, DNA binding and repair, oxidative and osmotic stress, phosphorus assimilation, and polyphosphate synthesis systems were enriched and identified as unique to the
panA
mutant. The cumulative proteomic data not only furthered our understanding of the archaeal proteasome system but also facilitated the assembly of the first subproteome map of
H. volcanii
.
Transcriptome Analysis of
Pseudomonas putida
in Response to Nitrogen Availability
Authors: Hervás Ana B. , Canosa Inés , Santero Eduardo 
Published: October 2007

ABSTRACT

This work describes a regulatory network of
Pseudomonas putida
controlled in response to nitrogen availability. We define NtrC as the master nitrogen regulator and suggest that it not only activates pathways for the assimilation of alternative nitrogen sources but also represses carbon catabolism under nitrogen-limited conditions, possibly to prevent excessive carbon and energy flow in the cell.
The Major Magnetosome Proteins MamGFDC Are Not Essential for Magnetite Biomineralization inMagnetospirillum gryphiswaldensebut Regulate the Size of Magnetosome Crystals
Authors: Scheffel André , Gärdes Astrid , Grünberg Karen , Wanner Gerhard , Schüler Dirk 
Published: October 2007

ABSTRACTMagnetospirillum gryphiswaldenseand related magnetotactic bacteria form magnetosomes, which are membrane-enclosed organelles containing crystals of magnetite (Fe3O4) that cause the cells to orient in magnetic fields. The characteristic sizes, morphologies, and patterns of alignment of magnetite crystals are controlled by vesicles formed of the magnetosome membrane (MM), which contains a number of specific proteins whose precise roles in magnetosome formation have remained largely elusive. Here, we report on a functional analysis of the small hydrophobic MamGFDC proteins, which altogether account for nearly 35% of all proteins associated with the MM. Although their high levels of abundance and conservation among magnetotactic bacteria had suggested a major role in magnetosome formation, we found that the MamGFDC proteins are not essential for biomineralization, as the deletion of neithermamC, encoding the most abundant magnetosome protein, nor the entiremamGFDCoperon abolished the formation of magnetite crystals. However, cells lackingmamGFDCproduced crystals that were only 75% of the wild-type size and were less regular than wild-type crystals with respect to morphology and chain-like organization. The inhibition of crystal formation could not be eliminated by increased iron concentrations. The growth of mutant crystals apparently was not spatially constrained by the sizes of MM vesicles, as cells lackingmamGFDCformed vesicles with sizes and shapes nearly identical to those formed by wild-type cells. However, the formation of wild-type-size magnetite crystals could be gradually restored by in-transcomplementation with one, two, and three genes of themamGFDCoperon, regardless of the combination, whereas the expression of all four genes resulted in crystals exceeding the wild-type size. Our data suggest that the MamGFDC proteins have partially redundant functions and, in a cumulative manner, control the growth of magnetite crystals by an as-yet-unknown mechanism.
Substitutions at Auxiliary Operator O3 Enhance Repression by Nitrate-Responsive Regulator NarL at Synthetic
lac
Control Regions in
Escherichia coli
K-12
Authors: Stewart Valley , Bledsoe Peggy J. 
Published: October 2007

ABSTRACT

We constructed monocopy
lac
operon control regions in which the operators O1-
lac
and O3-
lac
were replaced by NarL and NarP binding sites from the
nirB
or
napF
operon control regions. The results support the hypothesis that DNA-bound dimers of phospho-NarL can participate in higher-order cooperative interactions.
c-KIT overexpression, without gene amplification and mutation, in paediatric renal tumours
Authors: Jones Chris , Rodriguez-Pinilla Maria , Lambros Maryou , Bax Dorine , Messahel Boo , Vujanic Gordan M , Reis-Filho Jorge S , Pritchard-Jones Kathy 
Published: October 2007

Aims:To investigate the presence and prognostic relevance of KIT expression in paediatric renal tumours, and to determine whether receptor overexpression is associated with gene amplification and/or mutation.Methods:Immunohistochemistry without antigen retrieval for CD117 was carried out on tissue microarrays consisting of 274 Wilms’ tumours, 13 clear cell sarcomas of the kidney (CCSK), 10 mesoblastic nephromas (MN), and 7 rhabdoid tumours of the kidney (RTK). In addition, gene copy number was investigated by chromogenic in situ hybridisation (CISH), and overexpressing tumours were sequenced forKITmutations in exons 9, 11, 13 and 17.Results:Only 8/200 (4.0%) Wilms’ tumours exhibited any degree of moderate–strong KIT staining in any of their assessable cell types. This small group of KIT-positive tumours had a shorter time to relapse (p = 0.0044, log-rank test). There were no positive MNs or RTKs; however 3/11 (27.3%) CCSKs were strongly positive, with an additional two cases weakly reactive. No cases exhibited gene amplification or mutation.Conclusions:KIT overexpression in rare in Wilms’ tumours, although does appear to confer a worse prognosis, in particular for patients primarily treated with preoperative chemotherapy. CCSKs are associated with an increased expression of KIT, however, in the absence of gene amplification and/or activating mutation. The potential of anti-KIT therapeutic strategies in the treatment of paediatric renal tumours appears to be limited.
Implications of a congenitally abnormal valve: a study of 1025 consecutively excised aortic valves
Authors: Collins M J , Butany J , Borger M A , Strauss B H , David T E 
Published: October 2007

Background:An increasing proportion of patients with congenitally abnormal aortic valves (AV) present for AV replacement.Aims:To review morphological changes in a large contemporary patient population undergoing AV replacement.Methods:A detailed review was conducted for all 1025 patients who underwent AV replacement from 2002 to 2005, including the clinical indication for surgery, the type of native AV disease, the pathological changes observed in each valve and the need for related surgery.Results:Tricuspid (TAV), bicuspid (BAV) and unicuspid (UAV) aortic valves were observed in 64.5%, 31.9% and 3.0% of all patients respectively. A decreased number of cusps was associated with increasing predilection for male gender (83.9%, 73.4%, 59.2% for UAV, BAV, TAV respectively), a younger patient age at surgery (41.6 (14.3), 61.3 (12.8), 67.5 (12.9) years), and an increased occurrence of pathological changes in the cusps, including calcification of both the cusp and the base, ossification and ulceration. UAV and BAV were also associated with increasing replacement of the ascending aorta due to dilatation and aneurysm formation (54.8, 38.8%, 16.6%). The incidence of infective endocarditis and rheumatic heart disease was 3.8% and 11.2% of all excised valves respectively.Conclusion:UAV and BAV were increasingly likely to affect men, fail at an earlier age, and show an increasing incidence of pathological changes in the cusps and ascending aorta than TAV. These results suggest that TAV, BAV and UAV may represent a phenotypic continuum of a similar disease process.
Hepatic fibrosis and cirrhosis in the Fontan circulation: a detailed morphological study
Authors: Kendall T J , Stedman B , Hacking N , Haw M , Vettukattill J J , Salmon A P , Cope R , Sheron N , Millward-Sadler H , Veldtman G R , Iredale J P 
Published: October 2007

Aims:To describe the histological features of the liver in patients with a Fontan circulation.Methods:Specimens from liver biopsies carried out as part of preoperative assessment prior to extracardiac cavopulmonary conversion of an older style Fontan were examined and scored semi-quantitatively for pertinent histological features. To support the use of the scoring, biopsy specimens were also ranked by eye for severity to allow correlation with assigned scores.Results:Liver biopsy specimens from 18 patients with a Fontan circulation were assessed. All specimens showed sinusoidal fibrosis. In 17 cases there was at least fibrous spur formation, with 14 showing bridging fibrosis and 2 showing frank cirrhosis. In 17 cases at least some of the dense or sinusoidal fibrosis was orcein positive, although a larger proportion of the dense fibrous bands were orcein positive compared with the sinusoidal component. All specimens showed marked sinusoidal dilatation, and 14 showed bile ductular proliferation; 1 showed minimal iron deposition, and 1 showed mild lobular lymphocytic inflammation. There was no cholestasis or evidence of hepatocellular damage. Similar appearances were observed in 2 patients with severe tricuspid regurgitation.Discussion:The histological features of the liver in patients with a Fontan circulation are similar to those described in cardiac sclerosis. Sinusoidal dilatation and sinusoidal fibrosis are marked in the Fontan series. The presence of a significant amount of orcein negative sinusoidal fibrosis suggests there may be a remediable component, although the dense fibrous bands are predominantly orcein positive, suggesting chronicity and permanence. No inflammation or hepatocellular damage is evident, suggesting that fibrosis may be mediated by a non-inflammatory mechanism.
A knowledge unique: the life of the pioneering explorer and palaeontologist, Dorothea Bate (1878–1951)
Authors: Shindler Karolyn 
Published: October 2007

Abstract
Dorothea Bate is recognized as a pioneer of archaeozoology – the study of animal bones from archaeological sites. She also worked extensively on cave sites. Her research in Cyprus, Crete, Majorca and Menorca, as well as pre World War II Palestine, showed her holistic approach to the use of animal bones in order to deduce dates, climate and environment of the sites. She was the first woman to work as a scientist at the Natural History Museum, then known as the British Museum (Natural History), and her research reports on the fossil faunas are still being used today.