bluuebunny/crossref_metadata_2025_split huggingface dataset:
Iron-Dependent Cytochrome
c
1
Expression Is Mediated by the Status of Heme in
Bradyrhizobium japonicum
Authors: Gao Tao , O'Brian Mark R. 
Published: July 2005

ABSTRACT

The heme prosthetic group of heme proteins contains iron, which can be a limiting nutrient. Here, we show that cytochrome
c
1
protein from
Bradyrhizobium japonicum
was strongly affected by the iron status, with low expression in cells grown under iron limitation. This control was not affected in mutants encoding the iron regulator Irr or Fur. Furthermore, cytochrome
c
1
mRNA was not influenced by the iron status, suggesting control at a posttranscriptional step. Cytochrome
c
1
protein levels were very low in mutants defective in the genes encoding δ-aminolevulinic acid (ALA) synthase and ferrochelatase, enzymes that catalyze the first and final steps of the heme biosynthetic pathway, respectively. Iron-dependent cytochrome
c
1
expression was restored in the ALA synthase mutant by supplementation of the medium with the heme precursor ALA. Supplementation with heme resulted in high levels of cytochrome
c
1
protein in the wild type and in both mutants, but expression was no longer iron dependent. Cytochrome
c
1
is synthesized as a protein precursor fused with cytochrome
b
. A plasmid-borne construct encoding only cytochrome
c
1
was expressed in an iron- and heme-dependent manner similar to that of the wild-type gene, indicating that control by those effectors is not linked to posttranslational processing of the fusion protein. Mutation of the cytochrome
c
1
cysteines involved in covalent binding to heme nearly abolished immunodetectable protein. Thus, defects in heme synthesis or heme binding abrogate cytochrome
c
1
accumulation, apparently due to protein degradation. We suggest that iron-dependent cytochrome
c
1
expression is mediated by heme availability for heme protein formation
Carboxyl-Terminal Extensions beyond the Conserved Pentapeptide Reduce Rates of Chemoreceptor Adaptational Modification
Authors: Lai Wing-Cheung , Hazelbauer Gerald L. 
Published: July 2005

ABSTRACT

Sensory adaptation in bacterial chemotaxis is mediated by covalent modification of chemoreceptors. Specific glutamyl residues are methylated and demethylated in reactions catalyzed by methyltransferase CheR and methylesterase CheB. In the well-characterized chemosensory systems of
Escherichia coli
and
Salmonella
spp., efficient modification by either enzyme is dependent on a conserved pentapeptide sequence, NWETF or NWESF, present at the extreme carboxyl terminus of high-abundance chemoreceptors. To what extent is position at the extreme carboxyl terminus important for pentapeptide-mediated enhancement of adaptational modification? Is this position equally important for enhancement of both enzyme activities? To address these questions, we created forms of high-abundance receptor Tsr or Tar carrying one, six, or eight additional amino acids extending beyond the pentapeptide at their carboxyl termini and assayed methylation, demethylation, deamidation, and ability to mediate chemotaxis. In vitro and in vivo, all three carboxyl-terminal extensions reduced pentapeptide-mediated enhancement of rates of adaptational modification. CheB-catalyzed reactions were more affected than CheR-catalyzed reactions. Effects were less severe for the complete sensory system in vivo than for the minimal system of receptor and modification enzymes in vitro. Notably, extended receptors mediated chemotaxis as efficiently as wild-type receptors, providing a striking example of robustness in chemotactic systems. This could reflect compensatory reductions of rates for both modification reactions, mitigation of effects of slower reactions by the intertwined circuitry of signaling and adaptation, or tolerance of a range of reactions rates for adaptational modification. No matter what the mechanism, the observations provide a challenging test for mathematical models of chemotaxis.
Contribution of ExsFA and ExsFB Proteins to the Localization of BclA on the Spore Surface and to the Stability of the
Bacillus anthracis
Exosporium
Authors: Sylvestre Patricia , Couture-Tosi Evelyne , Mock Michèle 
Published: July 2005

ABSTRACT

Spores of
Bacillus anthracis
, the etiological agent of anthrax, and the closely related species
Bacillus cereus
and
Bacillus thuringiensis
, possess an exosporium, which is the outermost structure surrounding the mature spore. It consists of a paracrystalline basal layer and a hair-like outer layer. To date, the structural contribution of only one exosporium component, the collagen-like glycoprotein BclA, has been described. It is the structural component of the hair-like filaments. Here, we describe two other proteins, ExsFA and ExsFB, which are probably organized in multimeric complexes with other exosporium components, including BclA. Single and double
exsF
deletion mutants were constructed and analyzed. We found that inactivation of
exsF
genes affects the BclA content of spores. BclA is produced by all mutants. However, it is partially and totally released after mother cell lysis of the Δ
exsFA
and Δ
exsFA
Δ
exsFB
mutant strains, respectively. Electron microscopy revealed that the
exsF
mutant spores have defective exosporia. The Δ
exsFA
and Δ
exsFA
Δ
exsFB
spore surfaces are partially and totally devoid of filaments, respectively. Moreover, for all mutants, the crystalline basal layer appeared unstable. This instability revealed the presence of two distinct crystalline arrays that are sloughed off from the spore surface. These results indicate that ExsF proteins are required for the proper localization of BclA on the spore surface and for the stability of the exosporium crystalline layers.
Novel Intracellular 3-Hydroxybutyrate-Oligomer Hydrolase in
Wautersia eutropha
H16
Authors: Kobayashi Teruyuki , Uchino Keiichi , Abe Tomoko , Yamazaki Yuya , Saito Terumi 
Published: July 2005

ABSTRACT

Wautersia eutropha
H16 (formerly
Ralstonia eutropha
) mobilizes intracellularly accumulated poly(3-hydroxybutyrate) (PHB) with intracellular poly(3-hydroxybutyrate) depolymerases. In this study, a novel intracellular 3-hydroxybutyrate-oligomer hydrolase (PhaZc) gene was cloned and overexpressed in
Escherichia coli
. Then PhaZc was purified and characterized. Immunoblot analysis with polyclonal antiserum against PhaZc revealed that most PhaZc is present in the cytosolic fraction and a small amount is present in the poly(3-hydroxybutyrate) inclusion bodies of
W. eutropha
. PhaZc degraded various 3-hydroxybutyrate oligomers at a high specific activity and artificial amorphous poly(3-hydroxybutyrate) at a lower specific activity. Native PHB granules and semicrystalline PHB were not degraded by PhaZc. A PhaZ deletion mutation enhanced the deposition of PHB in the logarithmic phase in nutrient-rich medium. PhaZc differs from the hydrolases of
W. eutropha
previously reported and is a novel type of intracellular 3-hydroxybutyrate-oligomer hydrolase, and it participates in the mobilization of PHB along with other hydrolases.
CitI, a Transcription Factor Involved in Regulation of Citrate Metabolism in Lactic Acid Bacteria
Authors: Martin Mauricio G. , Magni Christian , de Mendoza Diego , López Paloma 
Published: July 2005

ABSTRACT

A large variety of lactic acid bacteria (LAB) can utilize citrate under fermentative conditions. Although much information concerning the metabolic pathways leading to citrate utilization by LAB has been gathered, the mechanisms regulating these pathways are obscure. In
Weissella paramesenteroides
(formerly called
Leuconostoc paramesenteroides
), transcription of the
citMDEFCGRP
citrate operon and the upstream divergent gene
citI
is induced by the presence of citrate in the medium. Although genetic experiments have suggested that CitI is a transcriptional activator whose activity can be modulated in response to citrate availability, specific details of the interaction between CitI and DNA remained unknown. In this study, we show that CitI recognizes two A+T-rich operator sites located between
citI
and
citM
and that the DNA-binding affinity of CitI is increased by citrate. Subsequently, this citrate signal propagation leads to the activation of the
cit
operon through an enhanced recruitment of RNA polymerase to its promoters. Our results indicate that the control of CitI by the cellular pools of citrate provides a mechanism for sensing the availability of citrate and adjusting the expression of the
cit
operon accordingly. In addition, this is the first reported example of a transcription factor directly functioning as a citrate-activated switch allowing the cell to optimize the generation of metabolic energy.
ywfEinBacillus subtilisCodes for a Novel Enzyme,l-Amino Acid Ligase
Authors: Tabata Kazuhiko , Ikeda Hajime , Hashimoto Shin-ichi 
Published: July 2005

ABSTRACTThe ATP-dependent carboxylate-amine/thiol ligase superfamily is known to contain enzymes catalyzing the formation of various types of peptide, such asd-alanyl-d-alanine, polyglutamate, and γ-peptide, but, curiously, no enzyme synthesizing α-dipeptides ofl-amino acids is known. We attempted to find such an enzyme. By in silico screening based on the consensus sequence of the superfamily followed by an in vitro assay with purified enzyme to avoid the degradation of the peptide(s) synthesized,ywfEofBacillus subtiliswas found to code for the activity formingl-alanyl-l-glutamine froml-alanine andl-glutamine with hydrolysis of ATP to ADP. No AMP was formed, supporting the idea that the enzyme belongs to the superfamily. Surprisingly, the enzyme accepted a wide variety ofl-amino acids. Among 231 combinations ofl-amino acids tested, reaction products were obtained for 111 combinations and 44 kinds of α-dipeptides were confirmed by high-performance liquid chromatography analyses, while no tripeptide or longer peptide was detected and thed-amino acids were inert. From these results, we propose thatywfEencodes a new member of the superfamily,l-amino acid ligase.
A DNA Polymerase V Homologue Encoded by TOL Plasmid pWW0 Confers Evolutionary Fitness on
Pseudomonas putida
under Conditions of Environmental Stress
Authors: Tark Mariliis , Tover Andres , Tarassova Kairi , Tegova Radi , Kivi Gaily , Hõrak Rita , Kivisaar Maia 
Published: July 2005

ABSTRACT

Plasmids in conjunction with other mobile elements such as transposons are major players in the genetic adaptation of bacteria in response to changes in environment. Here we show that a large catabolic TOL plasmid, pWW0, from
Pseudomonas putida
carries genes (
rulAB
genes) encoding an error-prone DNA polymerase Pol V homologue which increase the survival of bacteria under conditions of accumulation of DNA damage. A study of population dynamics in stationary phase revealed that the presence of pWW0-derived
rulAB
genes in the bacterial genome allows the expression of a strong growth advantage in stationary phase (GASP) phenotype of
P. putida
. When
rulAB
-carrying cells from an 8-day-old culture were mixed with Pol V-negative cells from a 1-day-old culture, cells derived from the aged culture out-competed cells from the nonaged culture and overtook the whole culture. At the same time, bacteria from an aged culture lacking the
rulAB
genes were only partially able to out-compete cells from a fresh overnight culture of the parental
P. putida
strain. Thus, in addition to conferring resistance to DNA damage, the plasmid-encoded Pol V genes significantly increase the evolutionary fitness of bacteria during prolonged nutritional starvation of a
P. putida
population. The results of our study indicate that RecA is involved in the control of expression of the pWW0-encoded Pol V.
Anagr-Like Two-Component Regulatory System inLactobacillus plantarumIs Involved in Production of a Novel Cyclic Peptide and Regulation of Adherence
Authors: Sturme Mark H. J. , Nakayama Jiro , Molenaar Douwe , Murakami Yoshiko , Kunugi Ryoko , Fujii Toshio , Vaughan Elaine E. , Kleerebezem Michiel , de Vos Willem M. 
Published: July 2005

ABSTRACTWe have analyzed a locus on the annotatedLactobacillus plantarumWCFS1 genome that showed homology to the staphylococcalagrquorum-sensing system and designated itlamforLactobacillus agr-like module. Production of thelamBDCAtranscript was shown to be growth phase dependent. Analysis of a response regulator-defective mutant (ΔlamA) in an adherence assay showed thatlamregulates adherence ofL. plantarumto a glass surface. Global transcription analysis of the wild-type and ΔlamAstrains in early, mid-, and late log phase of growth was performed using a clone-based microarray. Remarkably, only a small set of genes showed significant differences in transcription profiles between the wild-type andlamAmutant strains. The microarray analysis confirmed thatlamBDCAis autoregulatory and showed thatlamAis involved in regulation of expression of genes encoding surface polysaccharides, cell membrane proteins, and sugar utilization proteins. ThelamBDgenes encoding the putative autoinducing peptide precursor (LamD) and its processing protein (LamB) were overexpressed using the nisin-controlled expression system, and culture supernatants were analyzed by liquid chromatography/mass spectrometry (LC/MS) to identify overproduced LamD-derived peptides. In this way, a cyclic thiolactone pentapeptide that possesses a ring structure similar to those of autoinducing peptides of the staphylococcalagrsystem was identified. The peptide was designated LamD558, and its sequence (CVGIW) matched the annotated precursor peptide sequence. Time course analysis of wild-type culture supernatants by LC/MS indicated that LamD558 production was increased markedly from mid-log to late log growth phase. This is the first example of anagr-like system in nonpathogenic bacteria that encodes a cyclic thiolactone autoinducing peptide and is involved in regulation of adherence.
Characterization of a Small Heat Shock Protein, Mx Hsp16.6, ofMyxococcus xanthus
Authors: Otani Mieko , Ueki Toshiyuki , Kozuka Satoshi , Segawa Miki , Sano Keiji , Inouye Sumiko 
Published: July 2005

ABSTRACTA number of heat shock proteins inMyxococcus xanthuswere previously identified by two-dimensional (2D) gel electrophoresis. One of these protein was termed Mx Hsp16.6, and the gene encoding Mx Hsp16.6 was isolated. Mx Hsp16.6 consists of 147 amino acid residues and has an estimated molecular weight of 16,642, in accordance with the apparent molecular mass in the 2D gel. An α-crystallin domain, typically conserved in small heat shock proteins, was found in Mx Hsp16.6. Mx Hsp16.6 was not detected during normal vegetative growth but was immediately induced after heat shock. Expression of thehsp16.6gene was not induced by other stresses, such as starvation, oxidation, and high osmolarity. Mx Hsp16.6 was mostly localized in particles formed after heat shock and precipitated by low-speed centrifugation. Furthermore, Mx Hsp16.6 was detected in highly electron-dense particles in heat-shocked cells by immunoelectron microscopy, suggesting that it forms large complexes with heat-denatured proteins. An insertion mutation in thehsp16.6gene resulted in lower viability during heat shock and lower acquired thermotolerance. Therefore, it is likely that Mx Hsp16.6 plays critical roles in the heat shock response inM. xanthus.
Identification of an
Escherichia coli
Operon Required for Formation of the O-Antigen Capsule
Authors: Peleg Adi , Shifrin Yulia , Ilan Ophir , Nadler-Yona Chen , Nov Shani , Koby Simi , Baruch Kobi , Altuvia Shoshy , Elgrably-Weiss Maya , Abe Cecilia M. , Knutton Stuart , Saper Mark A. , Rosenshine Ilan 
Published: July 2005

ABSTRACT

Escherichia coli
produces polysaccharide capsules that, based on their mechanisms of synthesis and assembly, have been classified into four groups. The group 4 capsule (G4C) polysaccharide is frequently identical to that of the cognate lipopolysaccharide O side chain and has, therefore, also been termed the O-antigen capsule. The genes involved in the assembly of the group 1, 2, and 3 capsules have been described, but those required for G4C assembly remained obscure. We found that enteropathogenic
E. coli
(EPEC) produces G4C, and we identified an operon containing seven genes,
ymcD
,
ymcC
,
ymcB
,
ymcA
,
yccZ
,
etp
, and
etk
, which are required for formation of the capsule. The encoded proteins appear to constitute a polysaccharide secretion system. The G4C operon is absent from the genomes of enteroaggregative
E. coli
and uropathogenic
E. coli. E. coli
K-12 contains the G4C operon but does not express it, because of the presence of IS
1
at its promoter region. In contrast, EPEC, enterohemorrhagic
E. coli
, and
Shigella
species possess an intact G4C operon.
Cystic Fibrosis Sputum Supports Growth and Cues Key Aspects of
Pseudomonas aeruginosa
Physiology
Authors: Palmer Kelli L. , Mashburn Lauren M. , Singh Pradeep K. , Whiteley Marvin 
Published: July 2005

ABSTRACT

The opportunistic human pathogen
Pseudomonas aeruginosa
causes persistent airway infections in patients with cystic fibrosis (CF). To establish these chronic infections,
P. aeruginosa
must grow and proliferate within the highly viscous sputum in the lungs of CF patients. In this study, we used Affymetrix GeneChip microarrays to investigate the physiology of
P. aeruginosa
grown using CF sputum as the sole source of carbon and energy. Our results indicate that CF sputum readily supports high-density
P. aeruginosa
growth. Furthermore, multiple signals, which reduce swimming motility and prematurely activate the
Pseudomonas
quinolone signal cell-to-cell signaling cascade in
P. aeruginosa
, are present in CF sputum.
P. aeruginosa
factors critical for lysis of the common CF lung inhabitant
Staphylococcus aureus
were also induced in CF sputum and increased the competitiveness of
P. aeruginosa
during polymicrobial growth in CF sputum.
Use of Suppression-Subtractive Hybridization To Identify Genes in the
Burkholderia cepacia
Complex That Are Unique to
Burkholderia cenocepacia
Authors: Bernier Steve P. , Sokol Pamela A. 
Published: July 2005

ABSTRACT

We have previously shown differences in virulence between species of the
Burkholderia cepacia
complex using the alfalfa infection model and the rat agar bead chronic infection model.
Burkholderia cenocepacia
strains were more virulent in these two infection models than
Burkholderia multivorans
and
Burkholderia stabilis
strains. In order to identify genes that may account for the increased virulence of
B. cenocepacia
, suppression-subtractive hybridization was performed between
B. cenocepacia
K56-2 and
B. multivorans
C5393 and between
B. cenocepacia
K56-2 and
B. stabilis
LMG14294. Genes identified included DNA modification/phage-related/insertion sequences and genes involved in cell membrane/surface structures, resistance, transport, metabolism, regulation, secretion systems, as well as genes of unknown function. Several of these genes were present in the ET12 lineage of
B. cenocepacia
but not in other members of the
B. cepacia
complex. Virulence studies in a chronic lung infection model determined that the hypothetical YfjI protein, which is unique to the ET12 clone, contributes to lung pathology. Other genes specific to
B. cenocepacia
and/or the ET12 lineage were shown to play a role in biofilm formation and swarming or swimming motility.
Orientation within the Exosporium and Structural Stability of the Collagen-Like Glycoprotein BclA of
Bacillus anthracis
Authors: Boydston Jeremy A. , Chen Ping , Steichen Christopher T. , Turnbough Charles L. 
Published: July 2005

ABSTRACT

Bacillus anthracis
spores, which cause anthrax, are enclosed by an exosporium consisting of a basal layer and an external hair-like nap. The filaments of the nap are composed of BclA, a glycoprotein containing distinct N-terminal (NTD) and C-terminal (CTD) domains separated by an extended collagen-like central region. In this study, we used immunogold electron microscopy to show that the CTD of BclA forms the distal end of each filament of the hair-like nap, indicating that the NTD is attached to the basal layer. Ten randomly chosen anti-BclA monoclonal antibodies, raised against spores or exosporium, reacted with the CTD, consistent with its exterior location. We showed that recombinant BclA (rBclA), encoded by the
B. anthracis
Sterne strain and synthesized in
Escherichia coli
, forms a collagen-like triple helix as judged by collagenase sensitivity and circular dichroism spectroscopy. In contrast, native BclA in spores was resistant to collagenase digestion. Thermal denaturation studies showed that the collagen-like region of rBclA exhibited a melting temperature (
T

m

) of 37°C, like mammalian collagen. However, rBclA trimers exhibited
T

m

values of 84°C and 95°C in buffer with and without sodium dodecyl sulfate, respectively. CTD trimers exhibited the same
T

m

values, indicating that the high temperature and detergent resistances of rBclA were due to strong CTD interactions. We observed that CTD trimers are resistant to many proteases and readily form large crystalline sheets. Structural data indicate that the CTD is composed of multiple beta strands. Taken together, our results suggest that BclA and particularly its CTD form a rugged shield around the spore.
Staphylococcus aureus
Develops an Alternative,
ica-
Independent Biofilm in the Absence of the
arlRS
Two-Component System
Authors: Toledo-Arana Alejandro , Merino Nekane , Vergara-Irigaray Marta , Débarbouillé Michel , Penadés José R. , Lasa Iñigo 
Published: July 2005

ABSTRACT

The biofilm formation capacity of
Staphylococcus aureus
clinical isolates is considered an important virulence factor for the establishment of chronic infections. Environmental conditions affect the biofilm formation capacity of
S. aureus
, indicating the existence of positive and negative regulators of the process. The majority of the screening procedures for identifying genes involved in biofilm development have been focused on genes whose presence is essential for the process. In this report, we have used random transposon mutagenesis and systematic disruption of all
S. aureus
two-component systems to identify negative regulators of
S. aureus
biofilm development in a chemically defined medium (
H
ussain-
H
astings-
W
hite
m
odified medium [HHWm]). The results of both approaches coincided in that they identified
arlRS
as a repressor of biofilm development under both steady-state and flow conditions. The
arlRS
mutant exhibited an increased initial attachment as well as increased accumulation of poly-
N
-acetylglucosamine (PNAG). However, the biofilm formation of the
arlRS
mutant was not affected when the
icaADBC
operon was deleted, indicating that PNAG is not an essential compound of the biofilm matrix produced in HHWm. Disruption of the major autolysin gene,
atl
, did not produce any effect on the biofilm phenotype of an
arlRS
mutant. Epistatic experiments with global regulators involved in staphylococcal-biofilm formation indicated that
sarA
deletion abolished, whereas
agr
deletion reinforced, the biofilm development promoted by the
arlRS
mutation.