bluuebunny/crossref_metadata_2025_split huggingface dataset:
In Vivo Expression of the Mannose-Resistant Fimbriae of
Photorhabdus temperata
K122 during Insect Infection
Authors: Meslet-Cladiere L. M. , Pimenta A. , Duchaud E. , Holland I. B. , Blight M. A. 
Published: January 2004

ABSTRACT

Photorhabdus temperata
K122 is an entomopathogenic bacterium symbiotically associated with nematodes of the family
Heterorhabditidae
. Surface fimbriae are important for the colonization of many pathogenic bacteria, and here we report the nucleotide sequence and analysis of the expression of a 12-kbp fragment encoding the mannose-resistant fimbriae of
P. temperata
(
mrf
). The
mrf
gene cluster contains 11 genes with an organization similar to that of the
mrp
locus from
Proteus mirabilis. mrfI
(encoding a putative recombinase) and
mrfA
(encoding pilin), the first gene in an apparent operon of nine other genes, are expressed from divergent promoters. The
mrfI
-
mrfA
intergenic region contains inverted repeats flanking the
mrfA
promoter. This region was shown to be capable of inversion, consistent with an ON/OFF regulation of the operon. In in vitro liquid cultures, both orientations were detected. Nevertheless, when we analyzed the expression of all of the genes in the
mrf
locus by semiquantitative reverse transcription-PCR during infection of
Galleria mellonella
(greater wax moth) larvae, expression of
mrfA
was not detected until 25 h postinfection, preceding the death of the larvae at 32 h. In contrast,
mrfJ
(a putative inhibitor of flagellar synthesis) was expressed throughout infection. Expression of
mrfI
was also detected only late in infection (25 to 30 h), indicating a possible increase in inversion frequency at this stage. In both in vitro liquid cultures and in vivo larval infections, the distal genes of the operon were expressed at substantially lower levels than
mrfA
. These results indicate the complex regulation of the
mrf
cluster during infection.
A Novel Evolutionary Lineage of Carbonic Anhydrase (ε Class) Is a Component of the Carboxysome Shell
Authors: So Anthony K.-C. , Espie George S. , Williams Eric B. , Shively Jessup M. , Heinhorst Sabine , Cannon Gordon C. 
Published: January 2004

ABSTRACT

A significant portion of the total carbon fixed in the biosphere is attributed to the autotrophic metabolism of prokaryotes. In cyanobacteria and many chemolithoautotrophic bacteria, CO
2
fixation is catalyzed by ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO), most if not all of which is packaged in protein microcompartments called carboxysomes. These structures play an integral role in a cellular CO
2
-concentrating mechanism and are essential components for autotrophic growth. Here we report that the carboxysomal shell protein, CsoS3, from
Halothiobacillus neapolitanus
is a novel carbonic anhydrase (ε-class CA) that has an evolutionary lineage distinct from those previously recognized in animals, plants, and other prokaryotes. Functional CAs encoded by
csoS3
homologues were also identified in the cyanobacteria
Prochlorococcus
sp. and
Synechococcus
sp., which dominate the oligotrophic oceans and are major contributors to primary productivity. The location of the carboxysomal CA in the shell suggests that it could supply the active sites of RuBisCO in the carboxysome with the high concentrations of CO
2
necessary for optimal RuBisCO activity and efficient carbon fixation in these prokaryotes, which are important contributors to the global carbon cycle.
Reversible Acyl-Homoserine Lactone Binding to Purified
Vibrio fischeri
LuxR Protein
Authors: Urbanowski M. L. , Lostroh C. P. , Greenberg E. P. 
Published: January 2004

ABSTRACT

The
Vibrio fischeri
LuxR protein is the founding member of a family of acyl-homoserine lactone-responsive quorum-sensing transcription factors. Previous genetic evidence indicates that in the presence of its quorum-sensing signal,
N
-(3-oxohexanoyl) homoserine lactone (3OC6-HSL), LuxR binds to
lux
box DNA within the promoter region of the
luxI
gene and activates transcription of the
luxICDABEG
luminescence operon. We have purified LuxR from recombinant
Escherichia coli
. Purified LuxR binds specifically and with high affinity to DNA containing a
lux
box. This binding requires addition of 3OC6-HSL to the assay reactions, presumably forming a LuxR-3OC6-HSL complex. When bound to the
lux
box at the
luxI
promoter in vitro, LuxR-3OC6-HSL enables
E. coli
RNA polymerase to initiate transcription from the
luxI
promoter. Unlike the well-characterized LuxR homolog TraR in complex with its signal (3-oxo-octanoyl-HSL), the LuxR-30C6-HSL complex can be reversibly inactivated by dilution, suggesting that 3OC6-HSL in the complex is not tightly bound and is in equilibrium with the bulk solvent. Thus, although LuxR and TraR both bind 3-oxoacyl-HSLs, the binding is qualitatively different. The differences have implications for the ways in which these proteins respond to decreases in signal concentrations or rapid drops in population density.
The
Streptococcus gordonii
Platelet BindingProtein GspB Undergoes Glycosylation Independently ofExport
Authors: Bensing Barbara A. , Gibson Bradford W. , Sullam Paul M. 
Published: January 2004

ABSTRACT

The
binding of bacteria and platelets may play a central role in the
pathogenesis of infective endocarditis. Platelet binding by
Streptococcus gordonii
strain M99 is predominantly mediated by
the 286-kDa cell wall-anchored protein GspB. This unusually large
protein lacks a typical amino-terminal signal peptide and is
translocated from the cytoplasm via a dedicated transport system. A
14-kb segment just downstream of
gspB
encodes SecA2 and SecY2,
two components of the GspB-specific transport system. The downstream
segment also encodes several putative glycosyl transferases that may be
responsible for the posttranslational modification of GspB. In this
study, we compared the abilities of M99 and two
GspB
−
mutant strains to bind various
lectins. GspB was found to have affinity for lectins that bind
N
-acetylglucosamine. We also examined variant forms of GspB
that lack a carboxy-terminal cell wall-anchoring domain and thus are
free of covalent linkage to cell wall peptidoglycan. Like native GspB,
these truncated proteins appear to be heavily glycosylated, as
evidenced by migration during sodium dodecyl sulfate-polyacrylamide gel
electrophoresis with an apparent molecular mass >100 kDa in
excess of the predicted mass, negligible staining with conventional
protein stains, and reactivity with hydrazide following periodate
oxidation. Furthermore, analysis of the carbohydrate associated with
the GspB variants by high-pH anion-exchange chromatography revealed the
presence of ∼70 to 100 monosaccharide residues per GspB
polypeptide (primarily
N
-acetylglucosamine and glucose).
Analysis of GspB in protoplasts of s
ecA2
or
secY2
mutant strains, which do not export GspB, indicates that GspB is
glycosylated in the cytoplasm of these strains. The combined data
suggest that the native GspB is a glycoprotein and that it may be
glycosylated prior to
export.
Nine Mutants of
Chlorobium tepidum
Each Unable To Synthesize a Different Chlorosome Protein Still Assemble Functional Chlorosomes
Authors: Frigaard Niels-Ulrik , Li Hui , Milks Kirstin J. , Bryant Donald A. 
Published: January 2004

ABSTRACT

Chlorosomes of the green sulfur bacterium
Chlorobium tepidum
comprise mostly bacteriochlorophyll
c
(BChl
c
), small amounts of BChl
a
, carotenoids, and quinones surrounded by a lipid-protein envelope. These structures contain 10 different protein species (CsmA, CsmB, CsmC, CsmD, CsmE, CsmF, CsmH, CsmI, CsmJ, and CsmX) but contain relatively little total protein compared to other photosynthetic antenna complexes. Except for CsmA, which has been suggested to bind BChl
a
, the functions of the chlorosome proteins are not known. Nine mutants in which a single
csm
gene was inactivated were created; these mutants included genes encoding all chlorosome proteins except CsmA. All mutants had BChl
c
contents similar to that of the wild-type strain and had growth rates indistinguishable from or within ∼90% (CsmC
−
and CsmJ
−
) of those of the wild-type strain. Chlorosomes isolated from the mutants lacked only the protein whose gene had been inactivated and were generally similar to those from the wild-type strain with respect to size, shape, and BChl
c
, BChl
a
, and carotenoid contents. However, chlorosomes from the
csmC
mutant were about 25% shorter than those from the wild-type strain, and the BChl
c
absorbance maximum was blue-shifted about 8 nm, indicating that the structure of the BChl
c
aggregates in these chlorosomes is altered. The results of the present study establish that, except with CsmA, when the known chlorosome proteins are eliminated individually, none of them are essential for the biogenesis, light harvesting, or structural organization of BChl
c
and BChl
a
within the chlorosome. These results demonstrate that chlorosomes are remarkably robust structures that can tolerate considerable changes in protein composition.
Genetic Analysis of Disulfide Isomerization in
Escherichia coli
: Expression of DsbC Is Modulated by RNase E-Dependent mRNA Processing
Authors: Zhan Xiaoming , Gao Junjun , Jain Chaitanya , Cieslewicz Michael J. , Swartz James R. , Georgiou George 
Published: January 2004

ABSTRACT

We designed a selection strategy for the isolation of
Escherichia coli
mutants exhibiting enhanced protein disulfide isomerase activity. The folding of a variant of tissue plasminogen activator (v-tPA), a protein containing nine disulfide bonds, in the bacterial periplasm is completely dependent on the level of disulfide isomerase activity of the cell. Mutations that increase this activity mediate the formation of catalytically active v-tPA, which in turn cleaves a
p
-aminobenzoic acid (PABA)-peptide adduct to release free PABA and thus allows the growth of an auxotrophic strain. Following chemical mutagenesis, a total of eight
E. coli
mutants exhibiting significantly higher disulfide isomerization activity, not only with v-tPA but also with two other unrelated protein substrates, were isolated. This phenotype resulted from significantly increased expression of the bacterial disulfide isomerase DsbC. In seven of the eight mutants, the upregulation of DsbC was found to be related to defects in RNA processing by RNase E, the
rne
gene product. Specifically, the genetic lesions in five mutants were shown to be allelic to
rne
, while an additional two mutants exhibited impaired RNase E activity due to lesions in other loci. The importance of mRNA stability on the expression of DsbC is underscored by the short half-life of the
dsbC
transcript, which was found to be only 0.8 min at 37°C in wild-type cells but was two- to threefold longer in some of the stronger mutants. These results (i) confirm the central role of DsbC in disulfide bond isomerization in the bacterial periplasm and (ii) suggest a critical role for RNase E in regulating DsbC expression.
Mutational Analysis of the
Myxococcus xanthus
Ω4400 Promoter Region Provides Insight into Developmental Gene Regulation by C Signaling
Authors: Yoder Deborah R. , Kroos Lee 
Published: January 2004

ABSTRACT

Myxococcus xanthus
utilizes extracellular signals during development to coordinate cell movement, differentiation, and changes in gene expression. One of these signals, the C signal, regulates the expression of many genes, including Ω4400, a gene identified by an insertion of Tn
5 lac
into the chromosome. Expression of Tn
5 lac
Ω4400 is reduced in
csgA
mutant cells, which fail to perform C signaling, and the promoter region has several sequences similar to sequences found in the regulatory regions of other C-signal-dependent genes. One such gene, Ω4403, depends absolutely on the C signal for expression, and its promoter region has been characterized previously by mutational analysis. To determine if the similar sequences within the Ω4400 and Ω4403 regulatory regions function in the same way, deletion analysis and site-directed mutagenesis of the Ω4400 promoter region were performed. A 7-bp sequence centered at −49 bp, termed a C box, is identical in the Ω4400 and Ω4403 promoter regions, yet mutations in the individual base pairs affected expression from the two promoters very differently. Also, a single-base-pair change within a similar 5-bp element, which is centered at −61 bp in both promoter regions, had very different effects on the activities of the two promoters. Further mutational analysis showed that two regions are important for Ω4400 expression; one region, from −63 to −31 bp, is required for Ω4400 expression, and the other, from −86 to −81 bp, exerts a two- to fourfold effect on expression and is at least partially responsible for the C signal dependence of the Ω4400 promoter. Mutations in
sigD
and
sigE
, which are genes that encode σ factors, abolished and reduced Ω4400 expression, respectively. Expression of Ω4400 in
actB
or
actC
mutants correlated well with the altered levels of C signal produced in these mutants. Our results provide the first detailed analysis of an
M. xanthus
regulatory region that depends partially on C signaling for expression and indicate that similar DNA sequences in the Ω4400 and Ω4403 promoter regions function differently.
A Novel Putative Enterococcal Pathogenicity Island Linked to theespVirulence Gene ofEnterococcus faeciumand Associated with Epidemicity
Authors: Leavis Helen , Top Janetta , Shankar Nathan , Borgen Katrine , Bonten Marc , van Embden Jan , Willems Rob J. L. 
Published: January 2004

ABSTRACTEnterococcus faecalisharbors a virulence-associated surface protein encoded by theespgene. This gene has been shown to be part of a 150-kb putative pathogenicity island. A gene similar toesphas recently been found inEnterococcus faeciumisolates recovered from hospitalized patients. In the present study we analyzed the polymorphism in theespgene ofE. faecium, and we investigated the association ofespwith neighboring chromosomal genes. Theespgene showed considerable sequence heterogeneity in the regions encoding the nonrepeat N- and C-terminal domains of the Esp protein as well as differences in the number of repeats. DNA sequencing of chromosomal regions flanking theespgene ofE. faeciumrevealed seven open reading frames, representing putative genes implicated in virulence, regulation of transcription, and antibiotic resistance. These flanking regions were invariably associated with the presence or absence of theespgene inE. faecium, indicating thatespinE. faeciumis part of a distinct genetic element. Because of the presence of virulence genes in this gene cluster, the lower G+C content relative to that of the genome, and the presence ofespinE. faeciumisolates associated with nosocomial outbreaks and clinically documented infections, we conclude that this genetic element constitutes a putative pathogenicity island, the first one described inE. faecium. Except for the presence ofespandaraC, this pathogenicity island is completely different from theesp-containing pathogenicity island previously disclosed inE. faecalis.
ATP/ADP Translocases: a Common Feature of Obligate Intracellular Amoebal Symbionts Related to Chlamydiae and Rickettsiae
Authors: Schmitz-Esser Stephan , Linka Nicole , Collingro Astrid , Beier Cora L. , Neuhaus H. Ekkehard , Wagner Michael , Horn Matthias 
Published: January 2004

ABSTRACT

ATP/ADP translocases catalyze the highly specific transport of ATP across a membrane in an exchange mode with ADP. Such unique transport proteins are employed by plant plastids and have among the prokaryotes so far only been identified in few obligate intracellular bacteria belonging to the
Chlamydiales
and the
Rickettsiales
. In this study, 12 phylogenetically diverse bacterial endosymbionts of free-living amoebae and paramecia were screened for the presence of genes encoding ATP/ADP transport proteins. The occurrence of ATP/ADP translocase genes was found to be restricted to endosymbionts related to rickettsiae and chlamydiae. We showed that the ATP/ADP transport protein of the
Parachlamydia
-related endosymbiont of
Acanthamoeba
sp. strain UWE25, a recently identified relative of the important human pathogens
Chlamydia trachomatis
and
Chlamydophila pneumoniae
, is functional when expressed in the heterologous host
Escherichia coli
and demonstrated the presence of transcripts during the chlamydial developmental cycle. These findings indicate that the interaction between
Parachlamydia
-related endosymbionts and their amoeba hosts concerns energy parasitism similar to the interaction between pathogenic chlamydiae and their human host cells. Phylogenetic analysis of all known ATP/ADP translocases indicated that the genes encoding ATP/ADP translocases originated from a chlamydial ancestor and were, after an ancient gene duplication, transferred horizontally to rickettsiae and plants.
Quorum Sensing-Controlled Biofilm Development in
Serratia liquefaciens
MG1
Authors: Labbate Maurizio , Queck Shu Yeong , Koh Kai Shyang , Rice Scott A. , Givskov Michael , Kjelleberg Staffan 
Published: January 2004

ABSTRACT

Serratia liquefaciens
MG1 contains an
N-
acylhomoserine lactone-mediated quorum-sensing system that is known to regulate swarming motility colonization. In this study, we describe for
S. liquefaciens
MG1 the development of a novel biofilm consisting of cell aggregates and differentiated cell types, such as cell chains and long filamentous cells. Furthermore, quorum sensing is shown to be crucial for normal biofilm development and for elaborate differentiation. A mutant of
S. liquefaciens
MG1 that was incapable of synthesizing extracellular signal formed a thin and nonmature biofilm lacking cell aggregates and differentiated cell chains. Signal-based complementation of this mutant resulted in a biofilm with the wild-type architecture. Two quorum-sensing-regulated genes (
bsmA
and
bsmB
) involved in biofilm development were identified, and we propose that these genes are engaged in fine-tuning the formation of cell aggregates at a specific point in biofilm development.
Regulation of IcsP, the Outer Membrane Protease of the
Shigella
Actin Tail Assembly Protein IcsA, by Virulence Plasmid Regulators VirF and VirB
Authors: Wing Helen J. , Yan Arthur W. , Goldman Seth R. , Goldberg Marcia B. 
Published: January 2004

ABSTRACT

The
Shigella
outer membrane protease IcsP removes the actin assembly protein IcsA from the bacterial surface, and consequently modulates
Shigella
actin-based motility and cell-to-cell spread. Here, we demonstrate that IcsP expression is undetectable in mutants lacking either of two transcriptional activators, VirF and VirB. In wild-type
Shigella
spp.,
virB
expression is entirely dependent on VirF; therefore, to circumvent this regulatory cascade, we independently expressed VirF or VirB in
Shigella
strains lacking both activators and measured both IcsP levels and transcription from the
icsP
promoter. Our results show that VirB significantly enhanced
icsP
transcription, even in the absence of VirF. In contrast, when VirF was induced in the absence of VirB, VirF had variable effects. The regulation of
icsP
is distinctly different from the regulation of the gene encoding its major substrate,
icsA
, which is activated by VirF and not VirB. We propose that the different pathways regulating
icsA
and
icsP
may be critical to the modulation of IcsA-mediated actin-based motility by IcsP.
The NeuC Protein ofEscherichia coliK1 Is a UDPN-Acetylglucosamine 2-Epimerase
Authors: Vann Willie F. , Daines Dayle A. , Murkin Andrew S. , Tanner Martin E. , Chaffin Donald O. , Rubens Craig E. , Vionnet Justine , Silver Richard P. 
Published: January 2004

ABSTRACTThe K1 capsule is an essential virulence determinant ofEscherichia colistrains that cause meningitis in neonates. Biosynthesis and transport of the capsule, an α-2,8-linked polymer of sialic acid, are encoded by the 17-kbkpsgene cluster. We deletedneuC, a K1 gene implicated in sialic acid synthesis, from the chromosome of EV36, a K-12-K1 hybrid, by allelic exchange. Exogenously added sialic acid restored capsule expression to the deletion strain (ΔneuC), confirming that NeuC is necessary for sialic acid synthesis. The deduced amino acid sequence of NeuC showed similarities to those of UDP-N-acetylglucosamine (GlcNAc) 2-epimerases from both prokaryotes and eukaryotes. The NeuC homologue from serotype IIIStreptococcus agalactiaecomplements ΔneuC. We cloned theneuCgene into an intein expression vector to facilitate purification. We demonstrated by paper chromatography that the purifiedneuCgene product catalyzed the formation of [2-14C]acetamidoglucal and [N-14C]acetylmannosamine (ManNAc) from UDP-[14C]GlcNAc. The formation of reaction intermediate 2-acetamidoglucal with the concomitant release of UDP was confirmed by proton and phosphorus nuclear magnetic resonance spectroscopy. NeuC could not use GlcNAc as a substrate. These data suggest thatneuCencodes an epimerase that catalyzes the formation of ManNAc from UDP-GlcNAc via a 2-acetamidoglucal intermediate. The unexpected release of the glucal intermediate and the extremely low rate of ManNAc formation likely were a result of the in vitro assay conditions, in which a key regulatory molecule or protein was absent.
The Lactococcal Abortive Phage Infection System AbiP Prevents both Phage DNA Replication and Temporal Transcription Switch
Authors: Domingues Susana , Chopin Alain , Ehrlich S. Dusko , Chopin Marie-Christine 
Published: January 2004

ABSTRACTWe describe here a new lactococcal abortive phage infection system, designated AbiP. AbiP is effective against some lactococcal phages of one prevalent group, 936, but not against phages from the other two groups (c6A and P335). It was identified in theLactococcus lactissubsp.cremorisstrain IL420, on the native plasmid pIL2614. AbiP is encoded by a single gene, expressed in an operon with a second gene. In this work,abiPis shown to affect both the replication and transcription of phage DNA. In AbiP+cells, phage DNA replication is arrested approximately 10 min after infection. Levels of middle and late phage transcripts are lower in AbiP+than in AbiP−cells, probably due to the smaller amount of phage DNA. By contrast, early phage transcripts are more abundant in AbiP+than in AbiP−cells, suggesting that the switch-off, which occurs 15 min after infection in AbiP−cells, is prevented in AbiP+cells.
Control of Glucose- and NaCl-Induced Biofilm Formation by
rbf
in
Staphylococcus aureus
Authors: Lim Yong , Jana Malabendu , Luong Thanh T. , Lee Chia Y. 
Published: January 2004

ABSTRACT

Both
Staphylococcus aureus
and
S. epidermidis
are capable of forming biofilm on biomaterials. We used Tn
917
mutagenesis to identify a gene,
rbf
, affecting biofilm formation in
S. aureus
NCTC8325-4. Sequencing revealed that Rbf contained a consensus region signature of the AraC/XylS family of regulators, suggesting that Rbf is a transcriptional regulator. Insertional duplication inactivation of the
rbf
gene confirmed that the gene was involved in biofilm formation on polystyrene and glass. Phenotypic analysis of the wild type and the mutant suggested that the
rbf
gene mediates the biofilm formation of
S. aureus
at the multicellular aggregation stage rather than at initial attachment. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis demonstrated that the mutation resulted in the loss of an ∼190-kDa protein. Biofilm production by the mutant could be restored by complementation with a 2.5-kb DNA fragment containing the
rbf
gene. The
rbf
-specific mutation affected the induction of biofilm formation by glucose and a high concentration of NaCl but not by ethanol. The mutation did not affect the transcription of the
ica
genes previously shown to be required for biofilm formation. Taken together, our results suggest that the
rbf
gene is involved in the regulation of the multicellular aggregation step of
S. aureus
biofilm formation in response to glucose and salt and that this regulation may be mediated through the 190-kDa protein.