bluuebunny/crossref_metadata_2025_split huggingface dataset:
Relation of Capsular Polysaccharide Production and Colonial Cell Organization to Colony Morphology in
Vibrio parahaemolyticus
Authors: Enos-Berlage Jodi L. , McCarter Linda L. 
Published: July 2002

ABSTRACT

Vibrio parahaemolyticus
is a ubiquitous, gram-negative marine bacterium that undergoes phase variation between opaque and translucent colony morphologies. The purpose of this study was to determine the factor(s) responsible for the opaque and translucent phenotypes and to examine cell organization within both colony types. Examination of thin sections of ruthenium red-stained bacterial cells by electron microscopy revealed a thick, electron-dense layer surrounding the opaque cells that was absent in preparations from translucent strains. Extracellular polysaccharide (EPS) material was extracted from both opaque and translucent strains, and the opaque strain was shown to produce abundant levels of polysaccharide, in contrast to the translucent strain. Compositional analysis of the EPS identified four major sugars: glucose, galactose, fucose, and
N
-acetylglucosamine. Confocal scanning laser microscopy was used to investigate cell organization within opaque and translucent colonies. Cells within both types of colonies exhibited striking organization; rod-shaped cells were aligned parallel to one another and perpendicular to the agar surface throughout the depth of the colony. Cells within translucent colonies appeared more tightly packed than cells in opaque colonies. In addition, a dramatic difference in the structural integrity of these two colony types was observed. When colonies were perturbed, the cell organization of the translucent colonies was completely disrupted while the organization of the opaque colonies was maintained. To our knowledge, this study represents the first description of how cells are organized in the interior of a viable bacterial colony. We propose that the copious amount of EPS produced by the opaque strain fills the intercellular space within the colony, resulting in increased structural integrity and the opaque phenotype.
Trk1 and Trk2 Define the Major K
+
Transport System in Fission Yeast
Authors: Calero Fernando , Gómez Néstor , Ariño Joaquín , Ramos José 
Published: July 2002

ABSTRACT

The
trk1
+
gene has been proposed as a component of the K
+
influx system in the fission yeast
Schizosaccharomyces pombe
. Previous work from our laboratories revealed that
trk1
mutants do not show significantly altered content or influx of K
+
, although they are more sensitive to Na
+
. Genome database searches revealed that
S. pombe
encodes a putative gene (designated here
trk2
+
) that shows significant identity to
trk1
+
. We have analyzed the characteristics of potassium influx in
S. pombe
by using
trk1 trk2
mutants. Unlike budding yeast, fission yeast displays a biphasic transport kinetics.
trk2
mutants do not show altered K
+
transport and exhibit only a slightly reduced Na
+
tolerance. However,
trk1 trk2
double mutants fail to grow at low K
+
concentrations and show a dramatic decrease in Rb
+
influx, as a result of loss of the high-affinity transport component. Furthermore,
trk1 trk2
cells are very sensitive to Na
+
, as would be expected for a strain showing defective potassium transport. When
trk1 trk2
cells are maintained in K
+
-free medium, the potassium content remains higher than that of the wild type or
trk
single mutants. In addition, the
trk1 trk2
strain displays increased sensitivity to hygromycin B. These results are consistent with a hyperpolarized state of the plasma membrane. An additional phenotype of cells lacking both Trk components is a failure to grow at acidic pH. In conclusion, the Trk1 and Trk2 proteins define the major K
+
transport system in fission yeast, and in contrast to what is known for budding yeast, the presence of any of these two proteins is sufficient to allow growth at normal potassium levels.
The C-Terminal Portion of the Tail Fiber Protein of Bacteriophage Lambda Is Responsible for Binding to LamB, Its Receptor at the Surface of
Escherichia coli
K-12
Authors: Wang Jiang , Hofnung Maurice , Charbit Alain 
Published: July 2002

ABSTRACT

Bacteriophage λ adsorbs to its
Escherichia coli
K-12 host by interacting with LamB, its cell-surface receptor. We fused C-terminal portions of J, the tail fiber protein of λ, to maltose-binding protein. Solid-phase binding assays demonstrated that a purified fusion protein comprising only the last 249 residues of J could bind to LamB trimers and inhibited recognition by anti-LamB antibodies. Electron microscopy further demonstrated that the fusion protein could also bind to LamB at the surface of intact cells. This interaction prevented λ adsorption but affected only partially maltose uptake.
The
Streptococcus pneumoniae
Beta-Galactosidase Is a Surface Protein
Authors: Zähner Dorothea , Hakenbeck Regine 
Published: July 2002

ABSTRACT

The β-galactosidase gene of
Streptococcus pneumoniae
,
bgaA
, encodes a putative 2,235-amino-acid protein with the two amino acid motifs characteristic of the glycosyl hydrolase family of proteins. In addition, an N-terminal signal sequence and a C-terminal LPXTG motif typical of surface-associated proteins of gram-positive bacteria are present. Trypsin treatment of cells resulted in solubilization of the enzyme, documenting that it is associated with the cell envelope. In order to obtain defined mutants suitable for
lacZ
reporter experiments, the
bgaA
gene was disrupted, resulting in a complete absence of endogenous β-galactosidase activity. The results are consistent with β-galactosidase being a surface protein that seems not to be involved in lactose metabolism but that may play a role during pathogenesis.
A Sheep in Wolf's Clothing:
Listeria innocua
Strains with Teichoic Acid-Associated Surface Antigens and Genes Characteristic of
Listeria monocytogenes
Serogroup 4
Authors: Lan Zheng , Fiedler Franz , Kathariou Sophia 
Published: July 2002

ABSTRACT

Listeria monocytogenes
serotype 4b has been implicated in numerous food-borne epidemics and in a substantial fraction of sporadic listeriosis. A unique lineage of the nonpathogenic species
Listeria innocua
was found to express teichoic acid-associated surface antigens that were otherwise expressed only by
L. monocytogenes
of serotype 4b and the rare serotypes 4d and 4e. These
L. innocua
strains were also found to harbor sequences homologous to the gene
gtcA
, which has been shown to be essential for teichoic acid glycosylation in
L. monocytogenes
serotype 4b. Transposon mutagenesis and genetic studies revealed that the
gtcA
gene identified in this lineage of
L. innocua
was functional in serotype 4b-like glycosylation of the teichoic acids of these organisms. The genomic organization of the
gtcA
region was conserved between this lineage of
L. innocua
and
L. monocytogenes
serotype 4b. Our data are in agreement with the hypothesis that, in this lineage of
L. innocua
,
gtcA
was acquired by lateral transfer from
L. monocytogenes
serogroup 4. The high degree of nucleotide sequence conservation in the
gtcA
sequences suggests that such transfer was relatively recent. Transfer events of this type may alter the surface antigenic properties of
L. innocua
and may eventually lead to evolution of novel pathogenic lineages through additional acquisition of genes from virulent listeriae.
Role of thedinBGene Product in Spontaneous Mutation inEscherichia coliwith an Impaired Replicative Polymerase
Authors: Strauss B. S. , Roberts R. , Francis L. , Pouryazdanparast P. 
Published: July 2002

ABSTRACTWe isolated several new mutator mutations of theEscherichia colireplicative polymerasednaEsubunit alpha and used them and a previously reporteddnaEmutation to study spontaneous frameshift and base substitution mutations. Two of thesednaEstrains produce many more mutants when grown on rich (Luria-Bertani) than on minimal medium. A differential effect of the medium was not observed when thesednaEmutations were combined with a mismatch repair mutation. The selection scheme for thednaEmutations required that they be able to complement a temperature-sensitive strain. However, the ability to complement is not related to the mutator effect for at least one of the mutants. Comparison of the mutation rates for frameshift and base substitution mutations inmutSanddnaE mutSstrains suggests that the mismatch repair proteins respond differently to the two types of change. Deletion ofdinBfrom both chromosome and plasmid resulted in a four- to fivefold decrease in the rate of frameshift and base substitution mutations in adnaE mutSdouble mutant background. This reduction indicates that most mistakes in replication occur as a result of the action of the auxiliary rather than the replicative polymerase in thisdnaEmutant. Deletion ofdinBfrom strains carrying a wild-typednaEhad a measurable effect, suggesting that a fraction of spontaneous mutations occur as a result ofdinBpolymerase action even in cells with a normal replicative polymerase.
Increase of External Osmolarity Reduces Morphogenetic Defects and Accumulation of Chitin in a
gas1
Mutant of
Saccharomyces cerevisiae
Authors: Turchini Anna , Ferrario Laura , Popolo Laura 
Published: July 2002

ABSTRACT

We have performed a physiological analysis of the effects of high-osmolarity media on
gas1
Δ cells. The reductions in the duplication time, number of pluribudded cells, hypersensitivity to Calcofluor and sodium dodecyl sulfate, and chitin level indicate a partial suppression of the mutant phenotype.
GAS1
deletion was found to be lethal in the absence of the Bck1 and Slt2 (Mpk1) proteins of the cell integrity pathway.
pTAR-Encoded Proteins in Plasmid Partitioning
Authors: Kalnin Kirill , Stegalkina Svetlana , Yarmolinsky Michael 
Published: July 2002

ABSTRACT

Partition cassettes, essential for the segregational stability of low-copy-number bacterial plasmids, typically encode two autoregulated proteins and an adjacent
cis
-acting centromere analog to which one or perhaps both proteins bind. The diminutive partition region of pTAR of
Agrobacterium
spp. was reported to be exceptional, encoding only a single protein, ParA (D. R. Gallie and C. I. Kado, J. Mol. Biol. 193:465–478, 1987). However, resequencing of the region revealed two small downstream genes,
parB
and
orf-84
, of which only
parB
was found to be essential for partitioning in
A. tumefaciens
. Purified ParA exhibited a weak ATPase activity that was modestly increased by nonspecific DNA. ParB bound in vitro to repeated sequences present in a region,
parS
, that possesses centromere and operator functions and within which we identified the primary transcription start site by primer extension. In certain respects the Par proteins behave normally in the foreign host
Escherichia coli
. In
E. coli
, as in
A. tumefaciens
, ParB repressed the partition operon; ParA, inactive alone, augmented this repression. Functional similarities between the partition system of pTAR and those of other plasmids and bacteria are prominent, despite differences in size, organization, and amino acid sequence.
Characterization of the Enzymatic Component of
Clostridium perfringens
Iota-Toxin
Authors: Nagahama Masahiro , Sakaguchi Yoshihiko , Kobayashi Keiko , Ochi Sadayuki , Sakurai Jun 
Published: July 2002

ABSTRACT

The iota
a
component (i
a
) of
Clostridium perfringens
ADP ribosylates nonmuscle β/γ actin and skeletal muscle α-actin. Replacement of Arg-295 in i
a
with alanine led to a complete loss of NAD
+
-glycohydrolase (NADase) and ADP-ribosyltransferase (ARTase); that of the residue with lysine caused a drastic reduction in NADase and ARTase activities (<0.1% of the wild-type activities) but did not completely diminish them. Substitution of alanine for Glu-378 and Glu-380 caused a complete loss of NADase and ARTase. However, exchange of Glu-378 to aspartic acid or glutamine resulted in little effect on NADase activity but a drastic reduction in ARTase activity (<0.1% of the wild-type activity). Exchange of Glu-380 to aspartic acid caused a drastic reduction in NADase and ARTase activities (<0.1% of the wild-type activities) but did not completely diminish them; that of the residue to glutamine caused a complete loss of ARTase activity. Replacement of Ser-338 with alanine resulted in 0.7 to 2.3% wild-type activities, and that of Ser-340 and Thr-339 caused a reduction in these activities of 5 to 30% wild-type activities. The kinetic analysis showed that Arg-295 and Ser-338 also play an important role in the binding of NAD
+
to i
a
, that Arg-295, Glu-380, and Ser-338 play a crucial role in the catalytic rate of NADase activity, and that these three amino acid residues and Glu-378 are essential for ARTase activity. The effect of amino acid replacement in i
a
on ARTase activity was similar to that on lethal and cytotoxic activities, suggesting that lethal and cytotoxic activities in i
a
are dependent on ARTase activity.
1-Deoxy-
d
-Xylulose 5-Phosphate Synthase, the Gene Product of Open Reading Frame (ORF) 2816 and ORF 2895 in
Rhodobacter capsulatus
Authors: Hahn Frederick M. , Eubanks Lisa M. , Testa Charles A. , Blagg Brian S. J. , Baker Jonathan A. , Poulter C. Dale 
Published: July 2002

ABSTRACT

In eubacteria, green algae, and plant chloroplasts, isopentenyl diphosphate, a key intermediate in the biosynthesis of isoprenoids, is synthesized by the methylerythritol phosphate pathway. The five carbons of the basic isoprenoid unit are assembled by joining pyruvate and
d
-glyceraldehyde 3-phosphate. The reaction is catalyzed by the thiamine diphosphate-dependent enzyme 1-deoxy-
d
-xylulose 5-phosphate synthase. In
Rhodobacter capsulatus
, two open reading frames (ORFs) carry the genes that encode 1-deoxy-
d
-xylulose 5-phosphate synthase. ORF 2816 is located in the photosynthesis-related gene cluster, along with most of the genes required for synthesis of the photosynthetic machinery of the bacterium, whereas ORF 2895 is located elsewhere in the genome. The proteins encoded by ORF 2816 and ORF 2895, 1-deoxy-
d
-xylulose 5-phosphate synthase A and B, containing a His
6
tag, were synthesized in
Escherichia coli
and purified to greater than 95% homogeneity in two steps. 1-Deoxy-
d
-xylulose 5-phosphate synthase A appears to be a homodimer with 68 kDa subunits. A new assay was developed, and the following steady-state kinetic constants were determined for 1-deoxy-
d
-xylulose 5-phosphate synthase A and B:

K
m

pyruvate
= 0.61 and 3.0 mM,

K
m

d
-glyceraldehyde 3-phosphate

= 150 and 120 μM, and
V
max
= 1.9 and 1.4 μmol/min/mg in 200 mM sodium citrate (pH 7.4). The ORF encoding 1-deoxy-
d
-xylulose 5-phosphate synthase B complemented the disrupted essential
dxs
gene in
E. coli
strain FH11.
Cloning and Analysis of a
Candida albicans
Gene That Affects Cell Surface Hydrophobicity
Authors: Singleton David R. , Masuoka James , Hazen Kevin C. 
Published: July 2002

ABSTRACT

The opportunistic pathogenic yeast
Candida albicans
exhibits growth phase-dependent changes in cell surface hydrophobicity, which has been correlated with adhesion to host tissues. Cell wall proteins that might contribute to the cell surface hydrophobicity phenotype were released by limited glucanase digestion. These proteins were initially characterized by their rates of retention during hydrophobic interaction chromatography–high-performance liquid chromatography and used as immunogens for monoclonal antibody production. The present work describes the cloning and functional analysis of a
C. albicans
gene encoding a 38-kDa protein recognized by the monoclonal antibody 6C5-H4CA. The 6C5-H4CA antigen was resolved by two-dimensional electrophoresis, and a partial protein sequence was determined by mass spectrometry analysis of tryptic fragments. The obtained peptides were used to identify the gene sequence from the unannotated
C. albicans
DNA database. The antibody epitope was provisionally mapped by peptide display panning, and a peptide sequence matching the epitope was identified in the gene sequence. The gene sequence encodes a novel open reading frame (ORF) of unknown function that is highly similar to several other
C. albicans
ORFs and to a single
Saccharomyces cerevisiae
ORF. Knockout of the gene resulted in a decrease in measurable cell surface hydrophobicity and in adhesion of
C. albicans
to fibronectin. The results suggest that the 38-kDa protein is a hydrophobic surface protein that meditates binding to host target proteins.
Rebinding of Extracellular Adherence Protein Eap toStaphylococcus aureusCan Occur through a Surface-Bound Neutral Phosphatase
Authors: Flock Margareta , Flock Jan-Ingmar 
Published: July 2002

ABSTRACTExtracellular adherence protein Eap secreted fromStaphylococcus aureuswas previously found to enhance the adherence ofS. aureusto eukaryotic cells. This enhancement effect is due to the ability of Eap to rebind toS. aureusand to bind to eukaryotic cells and several plasma and matrix proteins. In this study we defined one potential binding target for Eap on the surface ofS. aureus, a surface-located neutral phosphatase. This phosphatase lacks an LPXTG region, but around 80% is retained on the cell surface. The soluble phosphatase can form a complex with Eap at a nonrandom molar ratio, and phosphatase activity is retained. The phosphatase can also bind to fibronectin. The cell surface-located portion presumably contributes to adherence ofS. aureusto fibronectin.
Overexpression of
yccL
(
gnsA
) and
ydfY
(
gnsB
) Increases Levels of Unsaturated Fatty Acids and Suppresses both the Temperature-Sensitive
fabA6
Mutation and Cold-Sensitive
secG
Null Mutation of
Escherichia coli
Authors: Sugai Rie , Shimizu Hisayo , Nishiyama Ken-ichi , Tokuda Hajime 
Published: July 2002

ABSTRACT

A multicopy suppressor of the cold-sensitive
secG
null mutation was isolated. The suppressor contained
sfa
and
yccL
, the former of which has been reported to be a multicopy suppressor of the
fabA6
mutation carried by a temperature-sensitive unsaturated fatty acid auxotroph. Subcloning of the suppressor gene revealed that
yccL
, renamed
gnsA
(
secG
null mutant suppressor), was responsible for the suppression of both the
secG
null mutation and the
fabA6
mutation. In contrast, the
sfa
gene did not suppress the
fabA6
mutation. The
ydfY
(
gnsB
) gene, encoding a protein which is highly similar to GnsA, also suppressed both the
secG
null mutation and the
fabA6
mutation. Although both
gnsA
and
gnsB
are linked to cold shock genes, the levels of GnsA and GnsB did not exhibit a cold shock response. A
gnsA-gnsB
double null mutant grew normally under all conditions examined; thus, the in vivo functions of
gnsA
and
gnsB
remain unresolved. However, overexpression of
gnsA
and
gnsB
stimulated proOmpA translocation of the
secG
null mutant at low temperature and caused a significant increase in the unsaturated fatty acid content of phospholipids. Taken together, these results suggest that an increase in membrane fluidity due to the increase in unsaturated fatty acids compensates for the absence of the SecG function, especially at low temperature.
Salmonella enterica
Serovar Typhimurium Swarming Mutants with Altered Biofilm-Forming Abilities: Surfactin Inhibits Biofilm Formation
Authors: Mireles Joe Robert , Toguchi Adam , Harshey Rasika M. 
Published: July 2002

ABSTRACT

Swarming motility plays an important role in surface colonization by several flagellated bacteria. Swarmer cells are specially adapted to rapidly translocate over agar surfaces by virtue of their more numerous flagella, longer cell length, and encasement of slime. The external slime provides the milieu for motility and likely harbors swarming signals. We recently reported the isolation of swarming-defective transposon mutants of
Salmonella enterica
serovar Typhimurium, a large majority of which were defective in lipopolysaccharide (LPS) synthesis. Here, we have examined the biofilm-forming abilities of the swarming mutants using a microtiter plate assay. A whole spectrum of efficiencies were observed, with LPS mutants being generally more proficient than wild-type organisms in biofilm formation. Since we have postulated that O-antigen may serve a surfactant function during swarming, we tested the effect of the biosurfactant surfactin on biofilm formation. We report that surfactin inhibits biofilm formation of wild-type
S. enterica
grown either in polyvinyl chloride microtiter wells or in urethral catheters. Other bio- and chemical surfactants tested had similar effects.
RecA-Mediated Rescue ofEscherichia coliStrains with Replication Forks Arrested at the Terminus
Authors: Maisnier-Patin Sophie , Nordström Kurt , Dasgupta Santanu 
Published: July 2002

ABSTRACTThe recombinational rescue of chromosome replication was investigated inEscherichia colistrains with the unidirectional originoriR1, from the plasmid R1, integrated withinoriCin clockwise (intR1CW) or counterclockwise (intR1CC) orientations. Only theintR1CCstrain, with replication forks arrested at the terminus, required RecA for survival. Unlike the strains with RecA-dependent replication known so far, theintR1CCstrain did not require RecBCD, RecF, RecG, RecJ, RuvAB, or SOS activation for viability. The overall levels of degradation of replicating chromosomes caused by inactivation of RecA were similar inoriCandintR1CCstrains. In theintR1CCstrain, RecA was also needed to maintain the integrity of the chromosome when the unidirectional replication forks were blocked at the terminus. This was consistent with suppression of the RecA dependence of theintR1CCstrain by inactivating Tus, the protein needed to block replication forks atTersites. Thus, RecA is essential during asymmetric chromosome replication for the stable maintenance of the forks arrested at the terminus and for their eventual passage across the termination barrier(s) independently of the SOS and some of the major recombination pathways.
Molecular Cloning and Functional Analysis of the MutY Homolog of
Deinococcus radiodurans
Authors: Li Xianghong , Lu A-Lien 
Published: July 2002

ABSTRACT

The
mutY
homolog gene (
mutY
Dr
) from
Deinococcus radiodurans
encodes a 39.4-kDa protein consisting of 363 amino acids that displays 35% identity to the
Escherichia coli
MutY (MutY
Ec
) protein. Expressed MutY
Dr
is able to complement
E. coli mutY
mutants but not
mutM
mutants to reduce the mutation frequency. The glycosylase and binding activities of MutY
Dr
with an A/G-containing substrate are more sensitive to high salt and EDTA concentrations than the activities with an A/7,8-dihydro-8-oxoguanine (GO)-containing substrate are. Like the MutY
Ec
protein, purified recombinant MutY
Dr
expressed in
E. coli
has adenine glycosylase activity with A/G, A/C, and A/GO mismatches and weak guanine glycosylase activity with a G/GO mismatch. However, MutY
Dr
exhibits limited apurinic/apyrimidinic lyase activity and can form only weak covalent protein-DNA complexes in the presence of sodium borohydride. This may be due to an arginine residue that is present in MutY
Dr
at the position corresponding to the position of MutY
Ec
Lys142, which forms the Schiff base with DNA. The kinetic parameters of MutY
Dr
are similar to those of MutY
Ec
. Although MutY
Dr
has similar substrate specificity and a binding preference for an A/GO mismatch over an A/G mismatch, as MutY
Ec
does, the binding affinities for both mismatches are slightly lower for MutY
Dr
than for MutY
Ec
. Thus, MutY
Dr
can protect the cell from GO mutational effects caused by ionizing radiation and oxidative stress.