bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mycobacterium tuberculosis
Cells Growing in Macrophages Are Filamentous and Deficient in FtsZ Rings
Authors: Chauhan Ashwini , Madiraju Murty V. V. S. , Fol Marek , Lofton Hava , Maloney Erin , Reynolds Robert , Rajagopalan Malini 
Published: February 2006

ABSTRACT

FtsZ, a bacterial homolog of tubulin, forms a structural element called the FtsZ ring (Z ring) at the predivisional midcell site and sets up a scaffold for the assembly of other cell division proteins. The genetic aspects of FtsZ-catalyzed cell division and its assembly dynamics in
Mycobacterium tuberculosis
are unknown. Here, with an
M. tuberculosis
strain containing FtsZ
TB
tagged with green fluorescent protein as the sole source of FtsZ, we examined FtsZ structures under various growth conditions. We found that midcell Z rings are present in approximately 11% of actively growing cells, suggesting that the low frequency of Z rings is reflective of their slow growth rate. Next, we showed that SRI-3072, a reported FtsZ
TB
inhibitor, disrupted Z-ring assembly and inhibited cell division and growth of
M. tuberculosis
. We also showed that
M. tuberculosis
cells grown in macrophages are filamentous and that only a small fraction had midcell Z rings. The majority of filamentous cells contained nonring, spiral-like FtsZ structures along their entire length. The levels of FtsZ in bacteria grown in macrophages or in broth were comparable, suggesting that Z-ring formation at midcell sites was compromised during intracellular growth. Our results suggest that the intraphagosomal milieu alters the expression of
M. tuberculosis
genes affecting Z-ring formation and thereby cell division.
Identification of
Rhodospirillum rubrum
GlnB Variants That Are Altered in Their Ability To Interact with Different Targets in Response to Nitrogen Status Signals
Authors: Zhu Yu , Conrad Mary C. , Zhang Yaoping , Roberts Gary P. 
Published: February 2006

ABSTRACT

In
Rhodospirillum rubrum
, NifA, the transcriptional activator for the
nif
genes, is posttranslationally activated only by the uridylylated form of GlnB, one of three P
II
homologs in the organism. We have used the yeast two-hybrid system to detect variants of GlnB that interact better with NifA than does wild-type GlnB. When examined for physiological effects in
R. rubrum
, these GlnB* variants activated NifA in the presence of NH
4
+
, which normally blocks NifA activation completely, and in the absence of GlnD, whose uridylylation of GlnB is also normally essential for NifA activation. When these variants were tested in the two-hybrid system for their interaction with NtrB, a receptor that should interact with the nonuridylylated form of GlnB, they were uniformly weaker than wild-type GlnB in that interaction. When expressed in
R. rubrum
either as single-copy integrants or on multiple-copy plasmids, these variants were also dramatically altered in terms of their ability to regulate several other receptors involved in nitrogen metabolism, including GlnE, NtrB/NtrC, and DRAT (dinitrogenase reductase ADP-ribosyl transferase)-DRAG (dinitrogenase reductase-activating glycohydrolase). The consistent pattern throughout is that these GlnB variants partially mimic the uridylylated form of wild-type GlnB, even under nitrogen-excess conditions and in strains lacking GlnD. The results suggest that the role of uridylylation of GlnB is primarily to shift the equilibrium of GlnB from a “nitrogen-sufficient” form to a “nitrogen-deficient” form, each of which interacts with different but overlapping receptor proteins in the cell. These GlnB variants apparently shift that equilibrium through direct structural changes.
Characterization of the Bifunctional Glycosyltransferase/Acyltransferase Penicillin-Binding Protein 4 of
Listeria monocytogenes
Authors: Zawadzka-Skomiał Joanna , Markiewicz Zdzislaw , Nguyen-Distèche Martine , Devreese Bart , Frère Jean-Marie , Terrak Mohammed 
Published: February 2006

ABSTRACT

Multimodular penicillin-binding proteins (PBPs) are essential enzymes responsible for bacterial cell wall peptidoglycan (PG) assembly. Their glycosyltransferase activity catalyzes glycan chain elongation from lipid II substrate (undecaprenyl-pyrophosphoryl-
N
-acetylglucosamine-
N
-acetylmuramic acid-pentapeptide), and their transpeptidase activity catalyzes cross-linking between peptides carried by two adjacent glycan chains.
Listeria monocytogenes
is a food-borne pathogen which exerts its virulence through secreted and cell wall PG-associated virulence factors. This bacterium has five PBPs, including two bifunctional glycosyltransferase/transpeptidase class A PBPs, namely, PBP1 and PBP4. We have expressed and purified the latter and have shown that it binds penicillin and catalyzes in vitro glycan chain polymerization with an efficiency of 1,400 M
−1
s
−1
from
Escherichia coli
lipid II substrate. PBP4 also catalyzes the aminolysis (
d
-Ala as acceptor) and hydrolysis of the thiolester donor substrate benzoyl-Gly-thioglycolate, indicating that PBP4 possesses both transpeptidase and carboxypeptidase activities. Disruption of the gene
lmo2229
encoding PBP4 in
L. monocytogenes
EGD did not have any significant effect on growth rate, peptidoglycan composition, cell morphology, or sensitivity to β-lactam antibiotics but did increase the resistance of the mutant to moenomycin.
Transcription Profiling of the
mgrA
Regulon in
Staphylococcus aureus
Authors: Luong Thanh T. , Dunman Paul M. , Murphy Ellen , Projan Steven J. , Lee Chia Y. 
Published: February 2006

ABSTRACT

MgrA has been shown to affect multiple
Staphylococcus aureus
genes involved in virulence and antibiotic resistance. To comprehensively identify the target genes regulated by
mgrA
, we employed a microarray method to analyze the transcription profiles of
S. aureus
Newman, its isogeneic
mgrA
mutant, and an MgrA-overproducing derivative. We compared genes that were differentially expressed at exponential or early stationary growth phases. Our results showed that MgrA affected an impressive number of genes, 175 of which were positively regulated and 180 of which were negatively regulated in an
mgrA
-specific manner. The target genes included all functional categories. The microarray results were validated by real-time reverse transcription-PCR quantitation of a set of selected genes from different functional categories. Our data also indicate that
mgrA
regulates virulence factors in a fashion analogous to that of the accessory gene regulatory locus (
agr
). Accordingly, exoproteins are upregulated and surface proteins are downregulated by the regulator, suggesting that
mgrA
may function in concert with
agr
. The fact that a large number of genes are regulated by
mgrA
implies that MgrA is a major global regulator in
S. aureus
.
Cloning, Expression, and Purification of Functional Sec11a and Sec11b, Type I Signal Peptidases of the Archaeon
Haloferax volcanii
Authors: Fine Amir , Irihimovitch Vered , Dahan Idit , Konrad Zvia , Eichler Jerry 
Published: February 2006

ABSTRACT

Across evolution, type I signal peptidases are responsible for the cleavage of secretory signal peptides from proteins following their translocation across membranes. In
Archaea
, type I signal peptidases combine domain-specific features with traits found in either their eukaryal or bacterial counterparts. Eukaryal and bacterial type I signal peptidases differ in terms of catalytic mechanism, pharmacological profile, and oligomeric status. In this study, genes encoding Sec11a and Sec11b, two type I signal peptidases of the halophilic archaeon
Haloferax volcanii
, were cloned. Although both genes are expressed in cells grown in rich medium, gene deletion approaches suggest that Sec11b, but not Sec11a, is essential. For purification purposes, tagged versions of the protein products of both genes were expressed in transformed
Haloferax volcanii
, with Sec11a and Sec11b being fused to a cellulose-binding domain capable of interaction with cellulose in hypersaline surroundings. By employing an in vitro signal peptidase assay designed for use with high salt concentrations such as those encountered by halophilic archaea such as
Haloferax volcanii
, the signal peptide-cleaving activities of both isolated membranes and purified Sec11a and Sec11b were addressed. The results show that the two enzymes differentially cleave the assay substrate, raising the possibility that the Sec11a and Sec11b serve distinct physiological functions.
Involvement of the LlaKR2I Methylase in Expression of the AbiR Bacteriophage Defense System in
Lactococcus lactis
subsp.
lactis
biovar diacetylactis KR2
Authors: Yang Julie M. , DeUrraza Patricio J. , Matvienko Nadya , O'Sullivan Daniel J. 
Published: February 2006

ABSTRACT

The native lactococcal plasmid, pKR223, from
Lactococcus lactis
subsp.
lactis
biovar diacetylactis KR2 encodes two distinct bacteriophage-resistant mechanisms, the LlaKR2I restriction and modification (R/M) system and the abortive infection (Abi) mechanism, AbiR, that impedes bacteriophage DNA replication. This study completed the characterization of AbiR, revealing that it is the first Abi system to be encoded by three genes,
abiRa
,
abiRb
, and
abiRc
, arranged in an operon and that it requires the methylase gene from the LlaKR2I R/M system. An analysis of deletion and insertion clones demonstrated that the AbiR operon was toxic in
L. lactis
without the presence of the LlaKR2I methylase, which is required to protect
L. lactis
from AbiR toxicity. The novelty of the AbiR system resides in its original gene organization and the unusual protective role of the LlaKR2I methylase. Interestingly, the AbiR genetic determinants are flanked by two IS
982
elements generating a likely transposable AbiR composite. This observation not only substantiated the novel function of the LlaKR2I methylase in the AbiR system but also illustrated the evolution of the LlaKR2I methylase toward a new and separate cellular function. This unique structure of both the LlaKR2I R/M system and the AbiR system may have contributed to the evolution of the LlaKR2I methylase toward a novel role comparable to that of the cell cycle-regulated methylases that include Dam and CcrM methylases. This new role for the LlaKR2I methylase offers a unique snapshot into the evolution of the cell cycle-regulated methylases from an existing R/M system.
Brucella abortus
Synthesizes Phosphatidylcholine from Choline Provided by the Host
Authors: Comerci Diego J. , Altabe Silvia , de Mendoza Diego , Ugalde Rodolfo A. 
Published: February 2006

ABSTRACT

The
Brucella
cell envelope is characterized by the presence of phosphatidylcholine (PC), a common phospholipid in eukaryotes that is rare in prokaryotes. Studies on the composition of
Brucella abortus
2308 phospholipids revealed that the synthesis of PC depends on the presence of choline in the culture medium, suggesting that the methylation biosynthetic pathway is not functional. Phospholipid composition of
pmtA
and
pcs
mutants indicated that in
Brucella
, PC synthesis occurs exclusively via the phosphatidylcholine synthase pathway. Transformation of
Escherichia coli
with an expression vector containing the
B. abortus pcs
homologue was sufficient for PC synthesis upon induction with IPTG (isopropyl-β-
d
-thiogalactopyranoside), while no PC formation was detected when bacteria were transformed with a vector containing
pmtA.
These findings imply that
Brucella
depends on choline provided by the host cell to form PC. We could not detect any obvious associated phenotype in the PC-deficient strain under vegetative or intracellular growth conditions in macrophages. However, the
pcs
mutant strain displays a reproducible virulence defect in mice, which suggests that PC is necessary to sustain a chronic infection process.
Mutational Analysis of an Extracytoplasmic-Function Sigma Factor To Investigate Its Interactions with RNA Polymerase and DNA
Authors: Wilson Megan J. , Lamont Iain L. 
Published: February 2006

ABSTRACT

The extracytoplasmic-function (ECF) family of sigma factors comprises a large group of proteins required for synthesis of a wide variety of extracytoplasmic products by bacteria. Residues important for core RNA polymerase (RNAP) binding, DNA melting, and promoter recognition have been identified in conserved regions 2 and 4.2 of primary sigma factors. Seventeen residues in region 2 and eight residues in region 4.2 of an ECF sigma factor, PvdS from
Pseudomonas aeruginosa
, were selected for alanine-scanning mutagenesis on the basis of sequence alignments with other sigma factors. Fourteen of the mutations in region 2 had a significant effect on protein function in an in vivo assay. Four proteins with alterations in regions 2.1 and 2.2 were purified as His-tagged fusions, and all showed a reduced affinity for core RNAP in vitro, consistent with a role in core binding. Region 2.3 and 2.4 mutant proteins retained the ability to bind core RNAP, but four mutants had reduced or no ability to cause core RNA polymerase to bind promoter DNA in a band-shift assay, identifying residues important for DNA binding. All mutations in region 4.2 reduced the activity of PvdS in vivo. Two of the region 4.2 mutant proteins were purified, and each showed a reduced ability to cause core RNA polymerase to bind to promoter DNA. The results show that some residues in PvdS have functions equivalent to those of corresponding residues in primary sigma factors; however, they also show that several residues not shared with primary sigma factors contribute to protein function.
A LuxR/LuxI-Type Quorum-Sensing System in a Plant Bacterium,
Mesorhizobium tianshanense
, Controls Symbiotic Nodulation
Authors: Zheng Huiming , Zhong Zengtao , Lai Xin , Chen Wen-Xin , Li Shunpeng , Zhu Jun 
Published: February 2006

ABSTRACT

The ability of rhizobia to symbiotically fix nitrogen from the atmosphere when forming nodules on their plant hosts requires various signal transduction pathways. LuxR-LuxI-type quorum-sensing systems have been shown to be one of the players in a number of rhizobium species. In this study, we found that
Mesorhizobium tianshanense
, a moderate-growth
Rhizobium
that forms nodules on a number of licorice plants, produces multiple
N
-acyl homoserine lactone (AHL)-like molecules. A simple screen for AHL synthase genes using an
M. tianshanense
genomic expression library in
Escherichia coli
, coupled with a sensitive AHL detector, uncovered a LuxI-type synthase, MrtI, and a LuxR-type regulator, MrtR, in
M. tianshanense
. Deletions of the
mrtI
or
mrtR
locus completely abolished AHL production in
M. tianshanense
. Using
lacZ
transcriptional fusions, we found that expression of the quorum-sensing regulators is autoinduced, as
mrtI
gene expression requires MrtR and cognate AHLs and
mrtR
expression is dependent on AHLs. Compared with the wild-type strains, quorum-sensing-deficient mutants showed a marked reduction in the efficiency of root hair adherence and, more importantly, were defective in nodule formation on their host plant,
Glycyrrhiza uralensis
. These data provide strong evidence that quorum sensing plays a critical role in the
M. tianshanense
symbiotic process.
The Cytoplasmic C-Terminal Domain of the
Escherichia coli
KdpD Protein Functions as a K
+
Sensor
Authors: Rothenbücher Marina C. , Facey Sandra J. , Kiefer Dorothee , Kossmann Marina , Kuhn Andreas 
Published: February 2006

ABSTRACT

The KdpD protein is a K
+
sensor kinase located in the cytoplasmic membrane of
Escherichia coli
. It contains four transmembrane stretches and two short periplasmic loops of 4 and 10 amino acid residues, respectively. To determine which part of KdpD functions as a K
+
sensor, genetic variants were constructed with truncations or altered arrangements of the transmembrane segments. All KdpD constructs were tested by complementation of an
E. coli kdpD
deletion strain for their ability to grow at a K
+
concentration of 0.1 mM in the medium. A soluble protein composed of the C-terminal cytoplasmic domain was able to complement the
kdpD
deletion strain. In addition, analysis of the β-galactosidase activity of an
E. coli
strain which carries a transcriptional fusion of the upstream region of the
kdpFABC
operon and a promoterless
lacZ
gene revealed that this soluble KdpD mutant responds to changes in the K
+
concentration in the extracellular medium. The results suggest that the sensing and response functions are both located in the C-terminal domain and might be modulated by the N-terminal domain as well as by membrane anchoring.
Mechanism of Growth Inhibition by Free Bile Acids in Lactobacilli and Bifidobacteria
Authors: Kurdi Peter , Kawanishi Koji , Mizutani Kanako , Yokota Atsushi 
Published: February 2006

ABSTRACT

The effects of the free bile acids (FBAs) cholic acid (CA), deoxycholic acid (DCA), and chenodeoxycholic acid on the bioenergetics and growth of lactobacilli and bifidobacteria were investigated. It was found that these FBAs reduced the internal pH levels of these bacteria with rapid and stepwise kinetics and, at certain concentrations, dissipated ΔpH. The bile acid concentrations that dissipated ΔpH corresponded with the MICs for the selected bacteria. Unlike acetate, propionate, and butyrate, FBAs dissipated the transmembrane electrical potential (ΔΨ). In
Bifidobacterium breve
JCM 1192, the synthetic proton conductor pentachlorophenol (PCP) dissipated ΔpH with a slow and continuous kinetics at a much lower concentration than FBAs did, suggesting the difference in mode of action between FBAs and true proton conductors. Membrane damage assessed by the fluorescence method and a viability decrease were also observed upon exposure to CA or DCA at the MIC but not to PCP or a short-chain fatty acid mixture. Loss of potassium ion was observed at CA concentrations more than 2 mM (0.4× MIC), while leakage of other cellular components increased at CA concentrations more than 4 mM (0.8× MIC). Additionally, in experiments with membrane phospholipid vesicles extracted from
Lactobacillus salivarius
subsp.
salicinius
JCM 1044, CA and DCA at the MIC collapsed the ΔpH with concomitant leakage of intravesicular fluorescent pH probe, while they did not show proton conductance at a lower concentration range (e.g., 0.2× MIC). Taking these observations together, we conclude that FBAs at the MIC disturb membrane integrity and that this effect can lead to leakage of proton (membrane ΔpH and ΔΨ dissipation), potassium ion, and other cellular components and eventually cell death.
Genetic Variation in the
Vibrio vulnificus
Group 1 Capsular Polysaccharide Operon
Authors: Chatzidaki-Livanis Maria , Jones Melissa K. , Wright Anita C. 
Published: February 2006

ABSTRACT

Vibrio vulnificus
produces human disease associated with raw-oyster consumption or wound infections, but fatalities are limited to persons with chronic underlying illness. Capsular polysaccharide (CPS) is required for virulence, and CPS expression correlates with opaque (Op) colonies that show “phase variation” to avirulent translucent (Tr) phenotypes with reduced CPS. The results discussed here confirmed homology of a
V. vulnificus
CPS locus to the group 1 CPS operon in
Escherichia coli
. However, two distinct
V. vulnificus
genotypes or alleles were associated with the operon, and they diverged at sequences encoding hypothetical proteins and also at unique, intergenic repetitive DNA elements. Phase variation was examined under conditions that promoted high-frequency transition of Op to Tr forms. Recovery of Tr isolates in these experiments showed multiple genotypes, which were designated TR1, TR2, and TR3: CPS operons of TR1 isolates were identical to the Op parent, and cells remained phase variable but expressed reduced CPS. TR2 and TR3 showed deletion mutations in one (
wzb
) or multiple genes, respectively, and deletion mutants were acapsular and locked in the Tr phase. Complementation in
trans
restored the Op phenotype in strains with the
wzb
deletion mutation. Allelic variation in repetitive elements determined the locations, rates, and extents of deletion mutations. Thus, different mechanisms are responsible for reversible phase variation in CPS expression versus genetic deletions in the CPS operon of
V. vulnificus
. Repetitive-element-mediated deletion mutations were highly conserved within the species and are likely to promote survival in estuarine environments.
Identification and Characterization of an ATP-Dependent Hexokinase with Broad Substrate Specificity from the Hyperthermophilic Archaeon
Sulfolobus tokodaii
Authors: Nishimasu Hiroshi , Fushinobu Shinya , Shoun Hirofumi , Wakagi Takayoshi 
Published: February 2006

ABSTRACT

As a new member of the glucose-phosphorylating enzymes, the ATP-dependent hexokinase from the hyperthermophilic crenarchaeon
Sulfolobus tokodaii
was purified, identified, and characterized. Our results revealed that the enzyme differs from other known enzymes in primary structure and its broad substrate specificity for both phosphoryl donors and acceptors.
Effects of Streptomycin Resistance Mutations on Posttranslational Modification of Ribosomal Protein S12
Authors: Carr Jennifer F. , Hamburg Daisy-Malloy , Gregory Steven T. , Limbach Patrick A. , Dahlberg Albert E. 
Published: February 2006

ABSTRACT

Ribosomal protein S12 contains a highly conserved aspartic acid residue that is posttranslationally β-methylthiolated. Using mass spectrometry, we have determined the modification states of several S12 mutants of
Thermus thermophilus
and conclude that β-methylthiolation is not a determinant of the streptomycin phenotype.
Demyelination in rheumatic diseases
Authors: Theodoridou A , Settas L 
Published: February 2006

Abstract
Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) characterised by multifocal areas of demyelination in the white matter of the brain and spinal cord. Autoantibodies, for example antinuclear antibodies, can also be present. MS and other demyelinating processes, such as transverse myelitis and optic neuritis (which may be clinically isolated cases or be part of the clinical spectrum of MS), are sometimes difficult to differentiate from CNS involvement in systemic autoimmune diseases like systemic lupus erythematosus (SLE), antiphospholipid syndrome (APS), Sjoegren's syndrome (SS), and Adamantiades-Behcet disease (BD). An acute isolated neurological syndrome presents the biggest diagnostic problem, since it is common in MS, but can also be the only feature or first manifestation in SLE, APS, SS, and BD. Indeed, the clinical presentation and lesions evidenced by magnetic resonance imaging may be similar.