bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mutants of
Streptomyces clavuligerus
with Disruptions in Different Genes for Clavulanic Acid Biosynthesis Produce Large Amounts of Holomycin: Possible Cross-Regulation of Two Unrelated Secondary Metabolic Pathways
Authors: de la Fuente Alvaro , Lorenzana Luis M. , Martín Juan F. , Liras Paloma 
Published: November 2002

ABSTRACT

A
Streptomyces clavuligerus ccaR
::
aph
strain, which has a disruption in the regulatory gene
ccaR
, does not produce cephamycin C or clavulanic acid, but does produce a bioactive compound that was identified as holomycin by high-performance liquid chromatography (HPLC) and infrared and mass spectrometry.
S. clavuligerus
strains with disruptions in different genes of the clavulanic acid pathway fall into three groups with respect to holomycin biosynthesis. (i) Mutants with mutations in the early steps of the pathway blocked in the gene
ceaS
(
pyc
) (encoding carboxyethylarginine synthase),
bls
(encoding a β-lactam synthetase), or open reading frame 6 (ORF6; coding for an acetyltransferase of unknown function) are holomycin nonproducers. (ii) Mutants blocked in the regulatory gene
ccaR
or
claR
or blocked in the last gene of the pathway encoding clavulanic acid reductase (
car
) produce holomycin at higher levels than the wild-type strain. (iii) Mutants with disruption in
cyp
(coding for cytochrome P450), ORF12, and ORF15, genes that appear to be involved in the conversion of clavaminic acid into clavaldehyde or in secretion steps, produce up to 250-fold as much holomycin as the wild-type strain. An assay for holomycin synthetase was developed. This enzyme forms holomycin from holothin by using acetyl coenzyme A as an acetyl group donor. The holomycin synthase activities in the different clavulanic acid mutants correlate well with their production of holomycin.
A Third Transposable Element, IS
Ppu12
, from the Toluene-Xylene Catabolic Plasmid pWW0 of
Pseudomonas putida
mt-2
Authors: Williams Peter A. , Jones Rheinallt M. , Shaw Linda E. 
Published: November 2002

ABSTRACT

A 3,372-bp insertion sequence, IS
Ppu12
, has been identified on the archetypal toluene-xylene TOL catabolic plasmid pWW0 from
Pseudomonas putida
mt-2. The insertion sequence element is located on the plasmid between bases 84397 and 87768 in a region which also contains the termini and transposase genes of the catabolic transposons Tn
4651
and Tn
4653
(A. Greated, L. Lambertson, P. A. Williams, and C. M. Thomas, Environ. Microbiol., in press). IS
Ppu12
has terminal inverted repeats of 24 bp with three mismatches and contains four open reading frames, a
tnpA
homologue and three open reading frames (
lspA
,
orf1
, and
orf2)
of undetermined function. After insertion in vitro of a Km
r
cassette into IS
Ppu12
either in the intergenic region between
orf1
and
orf2
or directly into the
orf1
gene and ligation into a suicide vector, the modified IS
Ppu12
-Km transposes at high frequency, often in multiple copies, into the chromosome of a
P. putida
recipient. Inactivation of
lspA
,
orf1
, and
orf2
by introducing a 7-bp deletion into the 5′ region of each gene had no major effect upon transposition, but a similar mutation of
tnpA
completely eliminated transposition. Analysis of the literature and of strains derived from the chlorobenzoate-degrading
Pseudomonas
sp. strain B13 suggests that the promiscuity of this element has played an important role in the history of plasmid pWW0. Database comparisons and the accompanying paper (A. J. Weightman, A. W. Topping, K. E. Hill, L. L. Lee, K. Sakai, J. H. Slater, and A. W. Thomas, J. Bacteriol. 184:6581-6591, 2002) show that IS
Ppu12
is a transposable element also found in other bacteria.
Transposition of
DEH
, a Broad-Host-Range Transposon Flanked by IS
Ppu12
, in
Pseudomonas putida
Is Associated with Genomic Rearrangements and Dehalogenase Gene Silencing
Authors: Weightman Andrew J. , Topping Andrew W. , Hill Katja E. , Lee Li Ling , Sakai Kenji , Slater J. Howard , Thomas Andrew W. 
Published: November 2002

ABSTRACT

Pseudomonas putida
strain PP3 produces two hydrolytic dehalogenases encoded by
dehI
and
dehII
, which are members of different
deh
gene families. The 9.74-kb
DEH
transposon containing
dehI
and its cognate regulatory gene,

dehR
I

, was isolated from strain PP3 by using the TOL plasmid pWW0.
DEH
was fully sequenced and shown to have a composite transposon structure, within which
dehI
and

dehR
I

were divergently transcribed and were flanked on either side by 3.73-kb identical direct repeats. The flanking repeat unit, designated IS
Ppu12
, had the structure of an insertion sequence in that it was bordered by 24-bp near-perfect inverted repeats and contained four open reading frames (ORFs), one of which was identified as
tnpA
, putatively encoding an ISL
3
family transposase. A putative lipoprotein signal peptidase was encoded by an adjacent ORF,
lspA
, and the others, IS
Ppu12 orf1
and
orf2
, were tentatively identified as a truncated cation efflux transporter gene and a PbrR family regulator gene, respectively. The
orf1
-
orf2
intergenic region contained an exact match with a previously described active, outward-orientated promoter, Pout. Transposition of
DEH
-IS
Ppu12
was investigated by cloning the whole transposon into a suicide plasmid donor, pAWT34, and transferring the construct to various recipients. In this way
DEH
-IS
Ppu12
was shown to transpose in a broad range of
Proteobacteria
. Transposition of IS
Ppu12
independently from
DEH
, and inverse transposition, whereby the vector DNA and IS
Ppu12
inserted into the target genome without the
deh
genes, were also observed to occur at high frequencies in
P. putida
PaW340. Transposition of a second
DEH
-IS
Ppu12
derivative introduced exogenously into
P. putida
PP3 via the suicide donor pAWT50 resulted in silencing of resident
dehI
and
dehII
genes in about 10% of transposition transconjugants and provided a genetic link between transposition of IS
Ppu12
and dehalogenase gene silencing. Database searches identified IS
Ppu12
-related sequences in several bacterial species, predominantly associated with plasmids and xenobiotic degradative genes. The potential role of IS
Ppu12
in gene silencing and activation, as well as the adaptation of bacteria to degrade xenobiotic compounds, is discussed.
Transcriptional Phase Variation of a Type III Restriction-Modification System in
Helicobacter pylori
Authors: de Vries Nicolette , Duinsbergen Dirk , Kuipers Ernst J. , Pot Raymond G. J. , Wiesenekker Patricia , Penn Charles W. , van Vliet Arnoud H. M. , Vandenbroucke-Grauls Christina M. J. E. , Kusters Johannes G. 
Published: November 2002

ABSTRACT

Phase variation is important in bacterial pathogenesis, since it generates antigenic variation for the evasion of immune responses and provides a strategy for quick adaptation to environmental changes. In this study, a
Helicobacter pylori
clone, designated MOD525, was identified that displayed phase-variable
lacZ
expression. The clone contained a transcriptional
lacZ
fusion in a putative type III DNA methyltransferase gene (
mod
, a homolog of the gene JHP1296 of strain J99), organized in an operon-like structure with a putative type III restriction endonuclease gene (
res
, a homolog of the gene JHP1297), located directly upstream of it. This putative type III restriction-modification system was common in
H. pylori
, as it was present in 15 out of 16 clinical isolates. Phase variation of the
mod
gene occurred at the transcriptional level both in clone MOD525 and in the parental
H. pylori
strain 1061. Further analysis showed that the
res
gene also displayed transcriptional phase variation and that it was cotranscribed with the
mod
gene. A homopolymeric cytosine tract (C tract) was present in the 5′ coding region of the
res
gene. Length variation of this C tract caused the
res
open reading frame (ORF) to shift in and out of frame, switching the
res
gene on and off at the translational level. Surprisingly, the presence of an intact
res
ORF was positively correlated with active transcription of the downstream
mod
gene. Moreover, the C tract was required for the occurrence of transcriptional phase variation. Our finding that translation and transcription are linked during phase variation through slipped-strand mispairing is new for
H. pylori
.
The Nitrate Reductase and Nitrite Reductase Operons and the
narT
Gene of
Staphylococcus carnosus
Are Positively Controlled by the Novel Two-Component System NreBC
Authors: Fedtke I. , Kamps A. , Krismer B. , Götz F. 
Published: November 2002

ABSTRACT

In
Staphylococcus carnosus
, the
nreABC
(for nitrogen regulation) genes were identified and shown to link the nitrate reductase operon (
narGHJI
) and the putative nitrate transporter gene
narT
. An
nreABC
deletion mutant, m1, was dramatically affected in nitrate and nitrite reduction and growth. Transcription of
narT
,
narGHJI
, and the nitrite reductase (
nir
) operon was severely reduced even when cells were cultivated anaerobically without nitrate or nitrite.
nreABC
transcripts were detected when cells were grown aerobically or anaerobically with or without nitrate or nitrite. NreA is a GAF domain-containing protein of unknown function. In vivo and in vitro studies showed that NreC is phosphorylated by NreB and that phospho-NreC specifically binds to a GC-rich palindromic sequence to enhance transcription initiation. This binding motif was found at the
narGHJI
,
nir
, and
narT
promoters but not at the
moeB
promoter. NreB is a cytosolic protein with four N-terminal cysteine residues. The second cysteine residue was shown to be important for NreB function. In vitro autophosphorylation of NreB was not affected by nitrate, nitrite, or molybdate. The
nir
promoter activity was iron dependent. The data provide evidence for a global regulatory system important for aerobic and anaerobic metabolism, with NreB and NreC forming a classical two-component system and NreB acting as a sensor protein with oxygen as the effector molecule.
Degradation of a
Caulobacter
Soluble Cytoplasmic Chemoreceptor Is ClpX Dependent
Authors: Potocka Isabel , Thein Melanie , Østerås Magne , Jenal Urs , Alley M. R. K. 
Published: November 2002

ABSTRACT

In order to determine whether ClpXP-mediated proteolysis is a common mechanism used to regulate the chemotaxis machinery during the cell cycle of
Caulobacter crescentus
, we have characterized a soluble cytoplasmic chemoreceptor, McpB. The
mcpB
gene lies adjacent to the major chemotaxis operon, which encodes 12 chemotaxis proteins, including the membrane chemoreceptor McpA. Like McpA, McpB possesses a C-terminal CheBR docking motif and three potential methylation sites, which we suggest are methylated. The McpB protein is degraded via a ClpX-dependent pathway during the swarmer-to-stalked cell transition, and a motif, which is 3 amino acids N-terminal to the McpB CheBR docking site, is required for proteolysis. Analysis of the degradation signal in McpB and McpA reveals a common motif present in the other four chemoreceptors that possess CheBR docking sites. A green fluorescent protein (GFP) fusion bearing 58 amino acids from the C terminus of McpA, which contains this motif, is degraded, suggesting that the C-terminal sequence is sufficient to confer ClpXP protease susceptibility.
Transcriptional Response of
Pasteurella multocida
to Defined Iron Sources
Authors: Paustian Michael L. , May Barbara J. , Cao Dongwei , Boley Daniel , Kapur Vivek 
Published: November 2002

ABSTRACT

Pasteurella multocida
was grown in iron-free chemically defined medium supplemented with hemoglobin, transferrin, ferritin, and ferric citrate as iron sources. Whole-genome DNA microarrays were used to monitor global gene expression over seven time points after the addition of the defined iron source to the medium. This resulted in a set of data containing over 338,000 gene expression observations. On average, 12% of
P. multocida
genes were differentially expressed under any single condition. A majority of these genes encoded
P. multocida
proteins that were involved in either transport and binding or were annotated as hypothetical proteins. Several trends are evident when the data from different iron sources are compared. In general, only two genes (
ptsN
and
sapD
) were expressed at elevated levels under all of the conditions tested. The results also show that genes with increased expression in the presence of hemoglobin did not respond to transferrin or ferritin as an iron source. Correspondingly, genes with increased expression in the transferrin and ferritin experiments were expressed at reduced levels when hemoglobin was supplied as the sole iron source. Finally, the data show that genes that were most responsive to the presence of ferric citrate did not follow a trend similar to that of the other iron sources, suggesting that different pathways respond to inorganic or organic sources of iron in
P. multocida
. Taken together, our results demonstrate that unique subsets of
P. multocida
genes are expressed in response to different iron sources and that many of these genes have yet to be functionally characterized.
Leaderless mRNAs Bind 70S Ribosomes More Strongly than 30S Ribosomal Subunits in
Escherichia coli
Authors: O'Donnell Sean M. , Janssen Gary R. 
Published: November 2002

ABSTRACT
By primer extension inhibition assays, 70S ribosomes bound with higher affinity, or stability, than did 30S subunits to leaderless mRNAs containing AUG or GUG start codons. Addition of translation initiation factors affected ribosome binding to leaderless mRNAs. Our results suggest that translation of leaderless mRNAs might initiate through a pathway involving 70S ribosomes or 30S subunits lacking IF3.
cis
-Acting Sequences of
Bacillus subtilis pyrG
mRNA Essential for Regulation by Antitermination
Authors: Meng Qi , Switzer Robert L. 
Published: November 2002

ABSTRACT

Expression of the
Bacillus subtilis pyrG
gene, which encodes CTP synthetase, is repressed by cytidine nucleotides. Regulation involves a termination-antitermination mechanism acting at a transcription terminator located within the 5′ untranslated
pyrG
leader sequence. Deletion and substitution mutagenesis of a series of
pyrG
′
-lacZ
transcriptional fusions integrated into the
B. subtilis
chromosome demonstrated that only the terminator stem-loop and two specific 4- to 6-nucleotide RNA sequences were required for derepression of
pyrG
by starvation for cytidine nucleotides. The first sequence, GGGC/U, comprises the first four nucleotides at the 5′ end of the
pyrG
transcript, and the second, GCUCCC, forms the first six nucleotides of the 5′ strand of the terminator stem. All of the nucleotides lying between the two required RNA sequences can be deleted without loss of regulation. We propose that an as-yet-unidentified regulatory protein binds to these two RNA segments and prevents termination of transcription in the
pyrG
leader region when intracellular CTP levels are low.
The Viable but Nonculturable State and Starvation Are Different Stress Responses of
Enterococcus faecalis
, as Determined by Proteome Analysis
Authors: Heim Sabina , Del Mar Lleo Maria , Bonato Barbara , Guzman Carlos A. , Canepari Pietro 
Published: November 2002

ABSTRACT

The protein expression patterns of exponentially growing, starved, and viable but nonculturable (VBNC)
Enterococcus faecalis
cells were analyzed to establish whether differences exist between the VBNC state and other stress responses. The results indicate that the protein profile of VBNC cells differs from that of either starved or exponentially growing bacteria. This demonstrates that the VBNC state is a distinct physiological phase within the life cycle of
E. faecalis
, which is activated in response to multiple environmental stresses.
The Atheroprotective Effect of Dietary Soy Isoflavones in Apolipoprotein E−/− Mice Requires the Presence of Estrogen Receptor-α
Authors: Adams Michael R. , Golden Deborah L. , Register Thomas C. , Anthony Mary S. , Hodgin Jeffrey B. , Maeda Nobuyo , Williams J. Koudy 
Published: November 2002

Objective—Although the mechanisms by which dietary soy inhibits atherosclerosis are unclear, one line of evidence implicates an important role for its phytoestrogenic isoflavones. We sought to determine whether soy isoflavones exert atheroprotective effects through estrogen receptor–dependent processes and, if so, which estrogen receptor subtype (ie, α or β) is involved.Methods and Results—We compared the effects of diets rich in soy protein that were either isoflavone depleted (0.04 mg/g protein isolate) or isoflavone-replete, or Soy(+IF) (1.72 mg/g protein isolate) in apolipoprotein E–deficient (ee) mice that had been crossed with estrogen receptor-α– and -β–deficient mice to produce double-knockout ααeeand ββeemice and (estrogen receptor) wild-type controls (AAeeand BBee). Both male and ovariectomized female mice were studied (n=10 to 17 per treatment group; total n=201). After 16 weeks, atherosclerosis was assessed by quantifying the aortic content of esterified cholesterol. Atherosclerosis was reduced 20% to 27% (P<0.05) by Soy(+IF) in ββee, BBee, and AAeemice but was unaffected in ααeemice. The inhibitory effect of Soy(+IF) was unrelated to sex, total plasma cholesterol, VLDL, LDL, and HDL cholesterol.Conclusions—The results indicate a necessary role for estrogen receptor-α–dependent processes in mediating the atheroprotective effects of dietary soy isoflavones.
PRESERVING BODIES, SAVING SOULS: RELIGIOUS INCONGRUITY IN A NORTHERN MALAGASY MINING TOWN
Authors: Walsh Andrew 
Published: November 2002

AbstractThis article discusses the manifestation of incongruent religious ideals in a northern Malagasy sapphire mining community. It focuses particularly on how members of an Evangelical Christian congregation and long-time resident animists of the place interpret one another's practices, and on how these interpretations reflect a fundamental disparity in how they understand the material world that they share. It argues that the conflict that has emerged between them can only be understood in light of this significant disparity.
Benefits at 3 yrs of an asthma education programme coupled with regular reinforcement
Authors: Ignacio-García J-M. , Pinto-Tenorio M. , Chocrón-Giraldez M.J. , Cabello-Rueda F. , López-Cozar Gil A.I. , Ignacio-García J-M. , de Ramón-Garrido E. 
Published: November 2002

The benefits at 3 yrs of an asthma self-management education programme coupled with educational reinforcement were assessed at follow-up visits in 63 adults with chronic asthma.Changes in asthma-related morbidity parameters, lung function and use of different classes of drugs before intervention and after 1, 2 and 3 yrs of the asthma education programme were compared using Friedman one-way analysis of variance.Improvements in the number of days off work or school, general practitioner consultations, admissions to emergency services, hospital admissions and nocturnal awakenings, as well as increases in forced expiratory volume in one second (FEV1), were significant. Comparison of data obtained at 1 yr and 2–3 yrs showed significant differences in the number of asthma-associated sleep disruptions, days off work or school and unscheduled visits to the general practitioner, as well as FEV1, but significant differences between the data obtained at 2 and 3 yrs were not observed. The percentage of patients using oral steroids had decreased significantly at 3 yrs.In adults with chronic asthma, an asthma self-management education programme coupled with educational reinforcement was effective at decreasing asthma morbidity, improving lung function and decreasing consumption of oral steroids.
The anti-IgE antibody omalizumab improves asthma-related quality of life in patients with allergic asthma
Authors: Buhl R. , Hanf G. , Solèr M. , Bensch G. , Wolfe J. , Everhard F. , Champain K. , Fox H. , Thirlwell J. 
Published: November 2002

The aim of the present study was to determine the effect of treatment with omalizumab, an anti-immunoglobulin E antibody, on asthma-related quality of life (AQoL) in patients with moderate-to-severe allergic asthma.A total of 546 patients with allergic asthma were randomised to double-blind subcutaneous treatment with either placebo or omalizumab for 52 weeks. A constant beclomethasone dipropionate dose was maintained during the first 16 weeks (steroid-stable phase). This was followed by a 12-week steroid-reduction phase. The core study was followed by a 24-week double-blind extension phase. AQoL was evaluated at baseline and at the end of the steroid-stable (week 16), steroid-reduction (week 28) and extension phases (week 52) using the Juniper Asthma Quality of Life Questionnaire (AQLQ).Baseline AQLQ scores were comparable for the two treatment groups. Relative to placebo, omalizumab-treated patients demonstrated statistically significant improvements from baseline across all four AQLQ domains, as well as overall AQoL score, at weeks 16 (except environmental exposure), 28 and 52. Patients on omalizumab were also more likely to achieve clinically significant improvements in AQoL during the course of the study. Overall, almost 70% of patients and investigators rated treatment with omalizumab as “excellent/good”, compared with ∼40% of placebo recipients.Clinical studies show that omalizumab enhances disease control whilst reducing corticosteroid consumption in patients with allergic asthma. The results of the present study show that these changes are paralleled by improvements in asthma-related quality of life that are meaningful to such patients.
Single-cell analysis: a novel approach to tumour necrosis factor-α synthesis and secretion in sarcoidosis
Authors: Pantelidis P. , McGrath D.S. , Southcott A.M. , du Bois R.M. 
Published: November 2002

Tumour necrosis factor (TNF)-α is thought to be a key early cytokine in the pathogenesis of sarcoidosis, despite conflicting data. Largely the product of mononuclear phagocyte activation, it is unclear whether TNF-α production at disease sites is a feature of all mononuclear phagocytes that accumulate there or whether it is secreted by a subset of these cells. Using the reverse haemolytic plaque assay, the aims of this study were to determine if the upregulation of TNF-α could be confirmed and to investigate whether this was monocyte or macrophage specific. The reverse haemolytic plaque assay allows the measurement of cytokine production at a single cell level.A greater number of alveolar macrophages produced TNF-α compared to autologous monocytes in sarcoidosis but not in controls and, based on cell size, it was confirmed that this was the product of more mature macrophages and that the secretion of TNF-α by monocytes and macrophages was heterogeneous: not all monocytes and macrophages secrete TNF-α. No differences in the average levels of TNF-α secretion by peripheral blood monocytes or alveolar macrophages were observed.This study has demonstrated that a subset of mononuclear phagocytes, mature macrophages, are responsible for tumour necrosis factor secretion and this could have implications for targeted management in sarcoidosis in the future.