bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Surface-Focused Biotinylation Procedure Identifies the
Yersinia pestis
Catalase KatY as a Membrane-Associated but Non-Surface-Located Protein
Authors: Myers-Morales Tanya , Cowan Clarissa , Gray Michael E. , Wulff Christine R. , Parker Carol E. , Borchers Christoph H. , Straley Susan C. 
Published: July 2007

ABSTRACT

This study identified major surface proteins of the plague bacterium
Yersinia pestis
. We applied a novel surface biotinylation method, followed by NeutrAvidin (NA) bead capture, on-bead digestion, and identification by liquid chromatography-tandem mass spectrometry (LC-MS-MS). The use of stachyose during biotinylation focused the reaction to the surface. Coupled with NA pulldown and immunoblot analysis, this method determined whether a protein was accessible to the surface. We applied the method to test the hypothesis that the catalase KatY is a surface protein of the plague bacterium
Y. pestis
. A rabbit serum recognized the catalase KatY as a major putative outer membrane-associated antigen expressed by
Y. pestis
cells grown at 37°C. Similar findings by other groups had led to speculations that this protein might be exposed to the surface and might be a candidate for evaluation as a protective antigen for an improved plague vaccine. KatY was obtained only in the total membrane fraction, and stachyose greatly reduced its biotinylation as well as that of the periplasmic maltose binding protein, indicating that KatY is not on the bacterial surface. LC-MS-MS analysis of on-bead digests representing ca. 10
9
cells identified highly abundant species, including KatY, Pal, and OmpA, as well as the lipoprotein Pcp, all of which bound in a biotin-specific manner. Pla, Lpp, and OmpX (Ail) bound to the NA beads in a non-biotin-specific manner. There was no contamination from abundant cytoplasmic proteins. We hypothesize that OmpX and Pcp are highly abundant and likely to be important for the
Y. pestis
pathogenic process. We speculate that a portion of KatY associates with the outer membrane in intact cells but that it is located on the periplasmic side. Consistent with this idea, it did not protect C57BL/6 mice against bubonic plague.
Serum Lipoproteins Promote Efficient Presentation of the Malaria Virulence Protein PfEMP1 at the Erythrocyte Surface
Authors: Frankland Sarah , Elliott Salenna R. , Yosaatmadja Francisca , Beeson James G. , Rogerson Stephen J. , Adisa Akinola , Tilley Leann 
Published: July 2007

ABSTRACTThe virulence of the malaria parasitePlasmodium falciparumis related to its ability to express a family of adhesive proteins known asP. falciparumerythrocyte membrane protein 1 (PfEMP1) at the infected red blood cell surface. The mechanism for the transport and delivery of these adhesins to the erythrocyte membrane is only poorly understood. In this work, we have used specific immune reagents in a flow cytometric assay to monitor the effects of serum components on the surface presentation of PfEMP1. We show that efficient presentation of the A4 and VAR2CSA variants of PfEMP1 is dependent on the presence of serum in the bathing medium during parasite maturation. Lipid-loaded albumin supports parasite growth but allows much less efficient presentation of PfEMP1 at the red blood cell surface. Analysis of the serum components reveals that lipoproteins, especially those of the low-density lipoprotein fraction, promote PfEMP1 presentation. Cytoadhesion of infected erythrocytes to the host cell receptors CD36 and ICAM-1 is also decreased in infected erythrocytes cultured in the absence of serum. The defect appears to be in the transfer of PfEMP1 from parasite-derived structures known as the Maurer's clefts to the erythrocyte membrane or in surface conformation rather than a down-regulation or switching of particular PfEMP1 variants.
Energy Metabolism of Bloodstream Form
Trypanosoma theileri
Authors: van Hellemond Jaap J. , Hoek Aad , Schreur Paul Wichgers , Chupin Vladimir , Özdirekcan Suat , Geysen Dirk , van Grinsven Koen W. A. , Koets Ad P. , Van den Bossche Peter , Geerts Stanny , Tielens Aloysius G. M. 
Published: July 2007

ABSTRACT

Bloodstream form
Trypanosoma theileri
degrades glucose to acetate (47%) and succinate (45%) and, therefore, does not solely rely on glycolysis for ATP production. This trypanosomatid does not use amino acids for energy metabolism. These results refute the prevailing hypothesis that substrate availability determines the type of energy metabolism of trypanosomatids.
Choline Induces Transcriptional Repression and Proteasomal Degradation of the Malarial Phosphoethanolamine Methyltransferase
Authors: Witola William Harold , Ben Mamoun Choukri 
Published: July 2007

ABSTRACT

During its intraerythrocytic life cycle, the malaria parasite
Plasmodium falciparum
undergoes dramatic metabolic and morphological changes and multiplies to produce up to 36 new daughter parasites. This rapid multiplication of the parasite requires an active synthesis of new membranes. The major component of these membranes, phosphatidylcholine, is synthesized via two metabolic routes, the CDP-choline pathway, which uses host choline as a precursor, and the plant-like serine decarboxylase-phosphoethanolamine methyltransferase (SDPM) pathway, which uses host serine as a precursor. Here we provide evidence indicating that the activity of the SDPM pathway is regulated by the CDP-choline precursor, choline. We show that the phosphoethanolamine methyltransferase, Pfpmt, a critical enzyme in the SDPM pathway, is down-regulated at the transcriptional level as well as targeted for degradation by the proteasome in the presence of choline. Transcript analysis revealed that Pf
PMT
transcription is repressed by choline in a dose-dependent manner. Immunoblotting, pulse-chase experiments, and immunoprecipitation studies demonstrated that Pfpmt degradation occurs not only in wild-type but also in transgenic parasites constitutively expressing Pfpmt. The proteasome inhibitor bortezomib inhibited choline-mediated Pfpmt degradation. These data provide the first evidence for metabolite-mediated transcriptional and proteasomal regulation in
Plasmodium
and will set the stage for the use of this system for conditional gene and protein expression in this organism.
Evolution of Microcin V and Colicin Ia Plasmids in
Escherichia coli
Authors: Jeziorowski Anne , Gordon David M. 
Published: July 2007

ABSTRACT

Survey results and genotypic characterization of
Escherichia coli
strains demonstrate that the bacteriocins colicin Ia and microcin V coassociate in a strain more often than would be expected by chance. When these two bacteriocins co-occur, they are encoded on the same conjugative plasmid. Plasmids encoding colicin Ia and microcin V are nonrandomly distributed with respect to the genomic background of the host strain. Characterization of microcin V and colicin Ia nucleotide variation, together with the backbone of plasmids encoding these bacteriocins, indicates that the association has evolved on multiple occasions and involves the movement of the microcin V operon, together with the genes
iroNEDCB
and
iss
, onto a nonrandom subset of colicin Ia plasmids. The fitness advantage conferred on cells encoding both colicin Ia and microcin V has yet to be determined.
Insertion Site Occupancy by
stx
2
Bacteriophages Depends on the Locus Availability of the Host Strain Chromosome
Authors: Serra-Moreno Ruth , Jofre Juan , Muniesa Maite 
Published: July 2007

ABSTRACT

Shiga toxin-producing
Escherichia coli
(STEC) is an emergent pathogen characterized by the expression of Shiga toxins, which are encoded in the genomes of lambdoid phages. These phages are infectious for other members of the
Enterobacteriaceae
and establish lysogeny when they integrate into the host chromosome. Five insertion sites, used mainly by these prophages, have been described to date. In the present study, the insertion of
stx
2
prophages in these sites was analyzed in 168 STEC strains isolated from cattle. Additionally, insertion sites were determined for
stx
2
phages which (i) converted diverse laboratory host strains, (ii) coexisted with another
stx
2
prophage, and (iii) infected a recombinant host strain lacking the most commonly used insertion site. Results show that depending on the host strain, phages preferentially use one insertion site. For the most part,
yehV
is occupied in STEC strains while
wrbA
is preferentially selected by the same
stx
phages in
E. coli
laboratory strains. If this primary insertion site is unavailable, then a secondary insertion site is selected. It can be concluded that insertion site occupancy by
stx
phages depends on the host strain and on the availability of the preferred locus in the host strain.
Flagellin Glycans from Two Pathovars of
Pseudomonas syringae
Contain Rhamnose in
d
and
l
Configurations in Different Ratios and Modified 4-Amino-4,6-Dideoxyglucose
Authors: Takeuchi Kasumi , Ono Hiroshi , Yoshida Mitsuru , Ishii Tadashi , Katoh Etsuko , Taguchi Fumiko , Miki Ryuji , Murata Katsuyoshi , Kaku Hanae , Ichinose Yuki 
Published: July 2007

ABSTRACT

Flagellins from
Pseudomonas syringae
pv. glycinea race 4 and
Pseudomonas syringae
pv. tabaci 6605 have been found to be glycosylated. Glycosylation of flagellin is essential for bacterial virulence and is also involved in the determination of host specificity. Flagellin glycans from both pathovars were characterized, and common sites of glycosylation were identified on six serine residues (positions 143, 164, 176, 183, 193, and 201). The structure of the glycan at serine 201 (S201) of flagellin from each pathovar was determined by sugar composition analysis, mass spectrometry, and
1
H and
13
C nuclear magnetic resonance spectroscopy. These analyses showed that the S201 glycans from both pathovars were composed of a common unique trisaccharide consisting of two rhamnosyl (Rha) residues and one modified 4-amino-4,6-dideoxyglucosyl (Qui4N) residue, β-
d
-Qui
p
4N(3-hydroxy-1-oxobutyl)2Me-(1→3)-α-
l
-Rha
p
-(1→2)-α-
l
-Rha
p
. Furthermore, mass analysis suggests that the glycans on each of the six serine residues are composed of similar trisaccharide units. Determination of the enantiomeric ratio of Rha from the flagellin proteins showed that flagellin from
P. syringae
pv. tabaci 6605 consisted solely of
l
-Rha, whereas
P. syringae
pv. glycinea race 4 flagellin contained both
l
-Rha and
d
-Rha at a molar ratio of about 4:1. Taking these findings together with those from our previous study, we conclude that these flagellin glycan structures may be important for the virulence and host specificity of
P. syringae
.
Structure-Function Analysis of the C-Terminal Domain of LcrV from
Yersinia pestis
Authors: Hamad Mohamad A. , Nilles Matthew L. 
Published: July 2007

ABSTRACT

LcrV, a multifunctional protein, acts as a positive regulator of effector protein secretion for the type III secretion system (T3SS) in
Yersinia pestis
by interaction with the negative regulator LcrG. In this study, LcrV was analyzed to identify regions required for LcrG interaction. Random-linker insertion mutagenesis, deletion analysis, and site-directed mutagenesis of hydrophobic amino acids between residues 290 and 311 allowed the isolation of an LcrV mutant (LcrV L291R F308R) defective for LcrG interaction. The new residues identified in LcrG interaction lie in helix 12 of LcrV; residues in helix 7 of LcrV are known to be involved in LcrG interaction. Helix 7 and helix 12 of LcrV interact to form an intramolecular coiled coil; these new results suggest that the intramolecular coiled coil in LcrV is required for LcrG interaction and activation of the T3SS.
The Two-Component Response Regulator RcsB Regulates Type 1 Piliation inEscherichia coli
Authors: Schwan William R. , Shibata Satoshi , Aizawa Shin-Ichi , Wolfe Alan J. 
Published: July 2007

ABSTRACTThe ability ofEscherichia colicells to produce type 1 pili depends upon the orientation of thefimApromoter. The orientation depends upon the ratios of the FimB and FimE recombinases. Here, we report that the two-component response regulator RcsB influences the piliation state by controllingfimBandfimEtranscription.
Cell-to-Cell Heterogeneity in Growth Rate and Gene Expression in
Methylobacterium extorquens
AM1
Authors: Strovas Tim J. , Sauter Linda M. , Guo Xiaofeng , Lidstrom Mary E. 
Published: July 2007

ABSTRACT

Cell-to-cell heterogeneity in gene expression and growth parameters was assessed in the facultative methylotroph
Methylobacterium extorquens
AM1. A transcriptional fusion between a well-characterized methylotrophy promoter (

P
mxaF

) and

gfp
uv

(encoding a variant of green fluorescent protein [GFPuv]) was used to assess single-cell gene expression. Using a flowthrough culture system and laser scanning microscopy, data on fluorescence and cell size were obtained over time through several growth cycles for cells grown on succinate or methanol. Cells were grown continuously with no discernible lag between divisions, and high cell-to-cell variability was observed for cell size at division (2.5-fold range), division time, and growth rate. When individual cells were followed over multiple division cycles, no direct correlation was observed between the growth rate before a division and the subsequent growth rate or between the cell size at division and the subsequent growth rate. The cell-to-cell variability for GFPuv fluorescence from the

P
mxaF

promoter was less, with a range on the order of 1.5-fold. Fluorescence and growth rate were also followed during a carbon shift experiment, in which cells growing on succinate were shifted to methanol. Variability of the response was observed, and the growth rate at the time of the shift from succinate to methanol was a predictor of the response. Higher growth rates at the time of the substrate shift resulted in greater decreases in growth rates immediately after the shift, but full induction of

P
mxaF

-

gfp
uv

was achieved faster. These results demonstrate that in
M. extorquens
, physiological heterogeneity at the single-cell level plays an important role in determining the population response to the metabolic shift examined.
The BclB Glycoprotein of
Bacillus anthracis
Is Involved in Exosporium Integrity
Authors: Thompson Brian M. , Waller Lashanda N. , Fox Karen F. , Fox Alvin , Stewart George C. 
Published: July 2007

ABSTRACT

Anthrax is a highly fatal disease caused by the gram-positive, endospore-forming, rod-shaped bacterium
Bacillus anthracis
. Spores, rather than vegetative bacterial cells, are the source of anthrax infections. Spores of
B. anthracis
are enclosed by a prominent loose-fitting structure called the exosporium. The exosporium is composed of a basal layer and an external hair-like nap. Filaments of the hair-like nap are made up largely of a single collagen-like glycoprotein called BclA. A second glycoprotein, BclB, has been identified in the exosporium layer. The specific location of this glycoprotein within the exosporium layer and its role in the biology of the spore are unknown. We created a mutant strain of
B. anthracis
ΔSterne that carries a deletion of the
bclB
gene. The mutant was found to possess structural defects in the exosporium layer of the spore (visualized by electron microscopy, immunofluorescence, and flow cytometry) resulting in an exosporium that is more fragile than that of a wild-type spore and is easily lost. Immunofluorescence studies also indicated that the mutant strain produced spores with increased levels of the BclA glycoprotein accessible to the antibodies on the surface. The resistance properties of the mutant spores were unchanged from those of the wild-type spores. A
bclB
mutation did not affect spore germination or kinetics of spore survival within macrophages. BclB plays a key role in the formation and maintenance of the exosporium structure in
B. anthracis
.
MglA Regulates
Francisella tularensis
subsp.
novicida
(
Francisella novicida
) Response to Starvation and Oxidative Stress
Authors: Guina Tina , Radulovic Dragan , Bahrami Arya J. , Bolton Diana L. , Rohmer Laurence , Jones-Isaac Kendan A. , Chen Jinzy , Gallagher Larry A. , Gallis Byron , Ryu Soyoung , Taylor Greg K. , Brittnacher Mitchell J. , Manoil Colin , Goodlett David R. 
Published: July 2007

ABSTRACT

MglA is a transcriptional regulator of genes that contribute to the virulence of
Francisella tularensis
, a highly infectious pathogen and the causative agent of tularemia. This study used a label-free shotgun proteomics method to determine the
F. tularensis
subsp.
novicida
(
F. novicida
) proteins that are regulated by MglA. The differences in relative protein amounts between wild-type
F. novicida
and the
mglA
mutant were derived directly from the average peptide precursor ion intensity values measured with the mass spectrometer by using a suite of mathematical algorithms. Among the proteins whose relative amounts changed in an
F. novicida mglA
mutant were homologs of oxidative and general stress response proteins. The
F. novicida mglA
mutant exhibited decreased survival during stationary-phase growth and increased susceptibility to killing by superoxide generated by the redox-cycling agent paraquat. The
F. novicida mglA
mutant also showed increased survival upon exposure to hydrogen peroxide, likely due to increased amounts of the catalase KatG. Our results suggested that MglA coordinates the stress response of
F. tularensis
and is likely essential for bacterial survival in harsh environments.
Global View of the
Clostridium thermocellum
Cellulosome Revealed by Quantitative Proteomic Analysis
Authors: Gold Nicholas D. , Martin Vincent J. J. 
Published: July 2007

ABSTRACT

A metabolic isotope-labeling strategy was used in conjunction with nano-liquid chromatography-electrospray ionization mass spectrometry peptide sequencing to assess quantitative alterations in the expression patterns of subunits within cellulosomes of the cellulolytic bacterium
Clostridium thermocellum
, grown on either cellulose or cellobiose. In total, 41 cellulosomal proteins were detected, including 36 type I dockerin-containing proteins, which count among them all but three of the known docking components and 16 new subunits. All differential expression data were normalized to the scaffoldin CipA such that protein per cellulosome was compared for growth between the two substrates. Proteins that exhibited higher expression in cellulosomes from cellulose-grown cells than in cellobiose-grown cells were the cell surface anchor protein OlpB, exoglucanases CelS and CelK, and the glycoside hydrolase family 9 (GH9) endoglucanase CelJ. Conversely, lower expression in cellulosomes from cells grown on cellulose than on cellobiose was observed for the GH8 endoglucanase CelA; GH5 endoglucanases CelB, CelE, CelG; and hemicellulases XynA, XynC, XynZ, and XghA. GH9 cellulases were the most abundant group of enzymes per CipA when cells were grown on cellulose, while hemicellulases were the most abundant group on cellobiose. The results support the existing theory that expression of scaffoldin-related proteins is coordinately regulated by a catabolite repression type of mechanism, as well as the prior observation that xylanase expression is subject to a growth rate-independent type of regulation. However, concerning transcriptional control of cellulases, which had also been previously shown to be subject to catabolite repression, a novel distinction was observed with respect to endoglucanases.
Uncovering New Metabolic Capabilities of
Bacillus subtilis
Using Phenotype Profiling of Rifampin-Resistant
rpoB
Mutants
Authors: Perkins Amy E. , Nicholson Wayne L. 
Published: July 2007

ABSTRACT

RNA polymerase is a central macromolecular machine controlling the flow of information from genotype to phenotype, and insights into global transcriptional regulation can be gained by studying mutational perturbations in the enzyme. Mutations in the RNA polymerase β subunit gene
rpoB
causing resistance to rifampin (Rif
r
) in
Bacillus subtilis
were previously shown to lead to alterations in the expression of a number of global phenotypes known to be under transcriptional control, such as growth, competence for transformation, sporulation, and germination (H. Maughan, B. Galeano, and W. L. Nicholson, J. Bacteriol. 186:2481-2486, 2004). To better understand the global effects of
rpoB
mutations on metabolism, wild-type and 11 distinct congenic Rif
r
mutant strains of
B. subtilis
were tested for utilization of 95 substrates by use of Biolog GP2 MicroPlates. A number of alterations of substrate utilization patterns were observed in the Rif
r
mutants, including the utilization of novel substrates previously unknown in
B. subtilis
, such as gentiobiose, β-methyl-
d
-glucoside, and
d
-psicose. The results indicate that combining global metabolic profiling with mutations in RNA polymerase provides a system-wide approach for uncovering previously unknown metabolic capabilities and further understanding global transcriptional control circuitry in
B. subtilis
.
The ExpR/Sin Quorum-Sensing System Controls Succinoglycan Production in
Sinorhizobium meliloti
Authors: Glenn Sarah A. , Gurich Nataliya , Feeney Morgan A. , González Juan E. 
Published: July 2007

ABSTRACT

Sinorhizobium meliloti
is a gram-negative soil bacterium capable of forming a symbiotic nitrogen-fixing relationship with its plant host,
Medicago sativa
. Various bacterially produced factors are essential for successful nodulation. For example, at least one of two exopolysaccharides produced by
S. meliloti
(succinoglycan or EPS II) is required for nodule invasion. Both of these polymers are produced in high- and low-molecular-weight (HMW and LMW, respectively) fractions; however, only the LMW forms of either succinoglycan or EPS II are active in nodule invasion. The production of LMW succinoglycan can be generated by direct synthesis or through the depolymerization of HMW products by the action of two specific endoglycanases, ExsH and ExoK. Here, we show that the ExpR/Sin quorum-sensing system in
S. meliloti
is involved in the regulation of genes responsible for succinoglycan biosynthesis as well as in the production of LMW succinoglycan. Therefore, quorum sensing, which has been shown to regulate the production of EPS II, also plays an important role in succinoglycan biosynthesis.
Characterization of Tunisian
Mycobacterium tuberculosis
Rifampin-Resistant Clinical Isolates
Authors: Soudani Alya , Hadjfredj Sondess , Zribi Mériem , Masmoudi Afef , Messaoud Taieb , Tiouri Hanéne , Fendri Chedlia 
Published: July 2007

ABSTRACT

Analysis of the gene encoding the β-subunit of
Mycobacterium tuberculosis
RNA polymerase (
rpoB
) has demonstrated a small region that harbors the mutations most frequently associated with rifampin resistance. In this study, we determined the occurrence of rifampin resistance in 544 Tunisian clinical
M. tuberculosis
strains isolated in a university hospital between 2004 and 2006 by using the standard-proportion agar method, the INNO-LiPA Rif.TB assay, and DNA sequencing.