bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dehalorespiration with Polychlorinated Biphenyls by an Anaerobic Ultramicrobacterium
Authors: May Harold D. , Miller Greg S. , Kjellerup Birthe V. , Sowers Kevin R. 
Published: January 2008

ABSTRACT

Anaerobic microbial dechlorination is an important step in the detoxification and elimination of polychlorinated biphenyls (PCBs), but a microorganism capable of coupling its growth to PCB dechlorination has not been isolated. Here we describe the isolation from sediment of an ultramicrobacterium, strain DF-1, which is capable of dechlorinating PCBs containing double-flanked chlorines added as single congeners or as Aroclor 1260 in contaminated soil. The isolate requires
Desulfovibrio
spp. in coculture or cell extract for growth on hydrogen and PCB in mineral medium. This is the first microorganism in pure culture demonstrated to grow by dehalorespiration with PCBs and the first isolate shown to dechlorinate weathered commercial mixtures of PCBs in historically contaminated sediments. The ability of this isolate to grow on PCBs in contaminated sediments represents a significant breakthrough for the development of in situ treatment strategies for this class of persistent organic pollutants.
Novel Members of Glycoside Hydrolase Family 13 Derived from Environmental DNA
Authors: Labes Antje , Karlsson Eva Nordberg , Fridjonsson Olafur H. , Turner Pernilla , Hreggvidson Gudmundur O. , Kristjansson Jakob K. , Holst Olle , Schönheit Peter 
Published: January 2008

ABSTRACT
Starch and pullulan-modifying enzymes of the α-amylase family (glycoside hydrolase family 13) have several industrial applications. To date, most of these enzymes have been derived from isolated organisms. To increase the number of members of this enzyme family, in particular of the thermophilic representatives, we have applied a consensus primer-based approach using DNA from enrichments from geothermal habitats. With this approach, we succeeded in isolating three new enzymes: a neopullulanase and two cyclodextrinases. Both cyclodextrinases displayed significant maltogenic amylase side activity, while one showed significant neopullulanase side activity. Specific motifs and domains that correlated with enzymatic activities were identified; e.g., the presence of the N domain was correlated with cyclodextrinase activity. The enzymes exhibited stability under thermophilic conditions and showed features appropriate for biotechnological applications.
Catabolic Pathway for the Production of Skatole and Indoleacetic Acid by the Acetogen
Clostridium drakei
,
Clostridium scatologenes
, and Swine Manure
Authors: Whitehead Terence R. , Price Neil P. , Drake Harold L. , Cotta Michael A. 
Published: January 2008

ABSTRACT

Skatole (3-methylindole) is a malodorous chemical in stored swine manure and is implicated as a component of foul-tasting pork. Definitive evidence for the skatole pathway is lacking. Deuterium-labeled substrates were employed to resolve this pathway in the acetogenic bacterium
Clostridium drakei
and
Clostridium scatologenes
and to determine if a similar pathway is used by microorganisms present in stored swine manure. Indoleacetic acid (IAA) was synthesized from tryptophan by both bacteria, and skatole was synthesized from both IAA and tryptophan. Microorganisms in swine manure produced skatole and other oxidation products from tryptophan, but IAA yielded only skatole. A catabolic mechanism for the synthesis of skatole is proposed.
Glycogen Phosphorylase in
Acanthamoeba
spp.: Determining the Role of the Enzyme during the Encystment Process Using RNA Interference
Authors: Lorenzo-Morales Jacob , Kliescikova Jarmila , Martinez-Carretero Enrique , De Pablos Luis Miguel , Profotova Bronislava , Nohynkova Eva , Osuna Antonio , Valladares Basilio 
Published: January 2008

ABSTRACT

Acanthamoeba
infections are difficult to treat due to often late diagnosis and the lack of effective and specific therapeutic agents. The most important reason for unsuccessful therapy seems to be the existence of a double-wall cyst stage that is highly resistant to the available treatments, causing reinfections. The major components of the
Acanthamoeba
cyst wall are acid-resistant proteins and cellulose. The latter has been reported to be the major component of the inner cyst wall. It has been demonstrated previously that glycogen is the main source of free glucose for the synthesis of cellulose in
Acanthamoeba
, partly as glycogen levels fall during the encystment process. In other lower eukaryotes (e.g.,
Dictyostelium discoideum
), glycogen phosphorylase has been reported to be the main tool used for glycogen breakdown in order to maintain the free glucose levels during the encystment process. Therefore, it was hypothesized that the regulation of the key processes involved in the
Acanthamoeba
encystment may be similar to the previously reported regulation mechanisms in other lower eukaryotes. The catalytic domain of the glycogen phosphorylase was silenced using RNA interference methods, and the effect of this phenomenon was assessed by light and electron microscopy analyses, calcofluor staining, expression zymogram assays, and Northern and Western blot analyses of both small interfering RNA-treated and control cells. The present report establishes the role of glycogen phosphorylase during the encystment process of
Acanthamoeba
. Moreover, the obtained results demonstrate that the enzyme is required for cyst wall assembly, mainly for the formation of the cell wall inner layer.
Key Function for the CCAAT-Binding Factor Php4 To Regulate Gene Expression in Response to Iron Deficiency in Fission Yeast
Authors: Mercier Alexandre , Watt Stephen , Bähler Jürg , Labbé Simon 
Published: January 2008

ABSTRACT

The fission yeast
Schizosaccharomyces pombe
responds to the deprivation of iron by inducing the expression of the
php4

+

gene, which encodes a negative regulatory subunit of the heteromeric CCAAT-binding factor. Once formed, the Php2/3/4/5 transcription complex is required to inactivate a subset of genes encoding iron-using proteins. Here, we used a pan-
S. pombe
microarray to study the transcriptional response to iron starvation and identified 86 genes that exhibit
php4

+

-dependent changes on a genome-wide scale. One of these genes encodes the iron-responsive transcriptional repressor Fep1, whose mRNA levels were decreased after treatment with the permeant iron chelator 2,2′-dipyridyl. In addition, several genes encoding the components of iron-dependent biochemical pathways, including the tricarboxylic acid cycle, mitochondrial respiration, amino acid biosynthesis, and oxidative stress defense, were downregulated in response to iron deficiency. Furthermore, Php4 repressed transcription when brought to a promoter using a yeast DNA-binding domain, and iron deprivation was required for this repression. On the other hand, Php4 was constitutively active when glutathione levels were depleted within the cell. Based on these and previous results, we propose that iron-dependent inactivation of Php4 is regulated at two distinct levels: first, at the transcriptional level by the iron-responsive GATA factor Fep1 and second, at the posttranscriptional level by a mechanism yet to be identified, which inhibits Php4-mediated repressive function when iron is abundant.
Role of
Staphylococcus aureus
Catalase in Niche Competition against
Streptococcus pneumoniae
Authors: Park Bonggoo , Nizet Victor , Liu George Y. 
Published: January 2008

ABSTRACT

Nasal colonization by
Staphylococcus aureus
is a major predisposing factor for subsequent infection. Recent reports of increased
S. aureus
colonization among children receiving pneumococcal vaccine implicate
Streptococcus pneumoniae
as an important competitor for the same niche. Since
S. pneumoniae
uses H
2
O
2
to kill competing bacteria, we hypothesized that oxidant defense could play a significant role in promoting
S. aureus
colonization of the nasal mucosa. Using targeted mutagenesis, we showed that
S. aureus
expression of catalase contributes significantly to the survival of this pathogen in the presence of
S. pneumoniae
both in vitro and in a murine model of nasal cocolonization.
Staphylococcus aureus
Pathogenicity Island DNA Is Packaged in Particles Composed of Phage Proteins
Authors: Tormo María Ángeles , Ferrer María Desamparados , Maiques Elisa , Úbeda Carles , Selva Laura , Lasa Íñigo , Calvete Juan J. , Novick Richard P. , Penadés José R. 
Published: January 2008

ABSTRACT

Staphylococcus aureus
pathogenicity islands (SaPIs) have an intimate relationship with temperate staphylococcal phages. During phage growth, SaPIs are induced to replicate and are efficiently encapsidated into special small phage heads commensurate with their size. We have analyzed by amino acid sequencing and mass spectrometry the protein composition of the specific SaPI particles. This has enabled identification of major capsid and tail proteins and a putative portal protein. As expected, all these proteins were phage encoded. Additionally, these analyses suggested the existence of a protein required for the formation of functional phage but not SaPI particles. Mutational analysis demonstrated that the phage proteins identified were involved only in the formation and possibly the function of SaPI or phage particles, having no role in other SaPI or phage functions.
Cyclic AMP-Dependent Catabolite Repression Is the Dominant Control Mechanism of Metabolic Fluxes under Glucose Limitation in
Escherichia coli
Authors: Nanchen Annik , Schicker Alexander , Revelles Olga , Sauer Uwe 
Published: January 2008

ABSTRACT

Although a whole arsenal of mechanisms are potentially involved in metabolic regulation, it is largely uncertain when, under which conditions, and to which extent a particular mechanism actually controls network fluxes and thus cellular physiology. Based on
13
C flux analysis of
Escherichia coli
mutants, we elucidated the relevance of global transcriptional regulation by ArcA, ArcB, Cra, CreB, CreC, Crp, Cya, Fnr, Hns, Mlc, OmpR, and UspA on aerobic glucose catabolism in glucose-limited chemostat cultures at a growth rate of 0.1 h
−1
. The by far most relevant control mechanism was cyclic AMP (cAMP)-dependent catabolite repression as the inducer of the phosphoenolpyruvate (PEP)-glyoxylate cycle and thus low tricarboxylic acid cycle fluxes. While all other mutants and the reference
E. coli
strain exhibited high glyoxylate shunt and PEP carboxykinase fluxes, and thus high PEP-glyoxylate cycle flux, this cycle was essentially abolished in both the Crp and Cya mutants, which lack the cAMP-cAMP receptor protein complex. Most other mutations were phenotypically silent, and only the Cra and Hns mutants exhibited slightly altered flux distributions through PEP carboxykinase and the tricarboxylic acid cycle, respectively. The Cra effect on PEP carboxykinase was probably the consequence of a specific control mechanism, while the Hns effect appears to be unspecific. For central metabolism, the available data thus suggest that a single transcriptional regulation process exerts the dominant control under a given condition and this control is highly specific for a single pathway or cycle within the network.
Binding of the Major Phasin, PhaP1, fromRalstonia eutrophaH16 to Poly(3-Hydroxybutyrate) Granules
Authors: Neumann Liv , Spinozzi Francesco , Sinibaldi Raffaele , Rustichelli Franco , Pötter Markus , Steinbüchel Alexander 
Published: January 2008

ABSTRACTThe surface of polyhydroxybutyrate (PHB) storage granules in bacteria is covered mainly by proteins referred to as phasins. The layer of phasins stabilizes the granules and prevents coalescence of separated granules in the cytoplasm and nonspecific binding of other proteins to the hydrophobic surfaces of the granules. Phasin PhaP1Reuis the major surface protein of PHB granules inRalstonia eutrophaH16 and occurs along with three homologues (PhaP2, PhaP3, and PhaP4) that have the capacity to bind to PHB granules but are present at minor levels. All four phasins lack a highly conserved domain but share homologous hydrophobic regions. To identify the region of PhaP1Reuwhich is responsible for the binding of the protein to the granules, N-terminal and C-terminal fusions of enhanced green fluorescent protein with PhaP1Reuor various regions of PhaP1Reuwere generated by recombinant techniques. The fusions were localized in the cells of various recombinant strains by fluorescence microscopy, and their presence in different subcellular protein fractions was determined by immunodetection of blotted proteins. The fusions were also analyzed to determine their capacities to bind to isolated PHB granules in vitro. The results of these studies indicated that unlike the phasin ofRhodococcus ruber, there is no discrete binding motif; instead, several regions of PhaP1Reucontribute to the binding of this protein to the surface of the granules. The conclusions are supported by the results of a small-angle X-ray scattering analysis of purified PhaP1Reu, which revealed that PhaP1Reuis a planar, triangular protein that occurs as trimer. This study provides new insights into the structure of the PHB granule surface, and the results should also have an impact on potential biotechnological applications of phasin fusion proteins and PHB granules in nanobiotechnology.
Identification of Genes Associated with the Long-Gut-Persistence Phenotype of the Probiotic
Lactobacillus johnsonii
Strain NCC533 Using a Combination of Genomics and Transcriptome Analysis
Authors: Denou Emmanuel , Pridmore Raymond David , Berger Bernard , Panoff Jean-Michel , Arigoni Fabrizio , Brüssow Harald 
Published: January 2008

ABSTRACT

Lactobacillus johnsonii
strains NCC533 and ATCC 33200 (the type strain of this species) differed significantly in gut residence time (12 versus 5 days) after oral feeding to mice. Genes affecting the long gut residence time of the probiotic strain NCC533 were targeted for analysis. We hypothesized that genes specific for this strain, which are expressed during passage of the bacterium through the gut, affect the phenotype. When the DNA of the type strain was hybridized against a microarray of the sequenced NCC533 strain, we identified 233 genes that were specific for the long-gut-persistence isolate. Whole-genome transcription analysis of the NCC533 strain using the microarray format identified 174 genes that were strongly and consistently expressed in the jejunum of mice monocolonized with this strain. Fusion of the two microarray data sets identified three gene loci that were both expressed in vivo and specific to the long-gut-persistence isolate. The identified genes included LJ1027 and LJ1028, two glycosyltransferase genes in the exopolysaccharide synthesis operon; LJ1654 to LJ1656, encoding a sugar phosphotransferase system (PTS) transporter annotated as mannose PTS; and LJ1680, whose product shares 30% amino acid identity with immunoglobulin A proteases from pathogenic bacteria. Knockout mutants were tested in vivo. The experiments revealed that deletion of LJ1654 to LJ1656 and LJ1680 decreased the gut residence time, while a mutant with a deleted exopolysaccharide biosynthesis cluster had a slightly increased residence time.
Comparative Genomics and an Insect Model Rapidly Identify Novel Virulence Genes of
Burkholderia mallei
Authors: Schell Mark A. , Lipscomb Lyla , DeShazer David 
Published: January 2008

ABSTRACT

Burkholderia pseudomallei
and its host-adapted deletion clone
Burkholderia mallei
cause the potentially fatal human diseases melioidosis and glanders, respectively. The antibiotic resistance profile and ability to infect via aerosol of these organisms and the absence of protective vaccines have led to their classification as major biothreats and select agents. Although documented infections by these bacteria date back over 100 years, relatively little is known about their virulence and pathogenicity mechanisms. We used in silico genomic subtraction to generate their virulome, a set of 650 putative virulence-related genes shared by
B. pseudomallei
and
B. mallei
but not present in five closely related nonpathogenic
Burkholderia
species. Although most of these genes are clustered in putative operons, the number of targets for mutant construction and verification of reduced virulence in animal models is formidable. Therefore,
Galleria mellonella
(wax moth) larvae were evaluated as a surrogate host; we found that
B. pseudomallei
and
B. mallei
, but not other phylogenetically related bacteria, were highly pathogenic for this insect. More importantly, four previously characterized
B. mallei
mutants with reduced virulence in hamsters or mice had similarly reduced virulence in
G. mellonella
larvae. Site-specific inactivation of selected genes in the computationally derived virulome identified three new potential virulence genes, each of which was required for rapid and efficient killing of larvae. Thus, this approach may provide a means to quickly identify high-probability virulence genes in
B. pseudomallei, B. mallei
, and other pathogens.
Genetic Interaction Screens with Ordered Overexpression and Deletion Clone Sets Implicate the
Escherichia coli
GTPase YjeQ in Late Ribosome Biogenesis
Authors: Campbell Tracey L. , Brown Eric D. 
Published: January 2008

ABSTRACT

The
Escherichia coli
protein YjeQ is a circularly permuted GTPase that is broadly conserved in bacteria. An emerging body of evidence, including cofractionation and in vitro binding to the ribosome, altered polysome profiles after YjeQ depletion, and stimulation of GTPase activity by ribosomes, suggests that YjeQ is involved in ribosome function. The growth of strains lacking YjeQ in culture is severely compromised. Here, we probed the cellular function of YjeQ with genetic screens of ordered
E. coli
genomic libraries for suppressors and enhancers of the slow-growth phenotype of a Δ
yjeQ
strain. Screening for suppressors using an ordered library of 374 clones overexpressing essential genes and genes associated with ribosome function revealed that two GTPases, Era and initiation factor 2, ameliorated the growth and polysome defects of the Δ
yjeQ
strain. In addition, seven bona fide enhancers of slow growth were identified (Δ
tgt
, Δ
ksgA
, Δ
ssrA
, Δ
rimM
, Δ
rluD
, Δ
trmE/mnmE
, and Δ
trmU/mnmA
) among 39 deletions (in genes associated with ribosome function) that we constructed in the Δ
yjeQ
genetic background. Taken in context, our work is most consistent with the hypothesis that YjeQ has a role in late 30S subunit biogenesis.
Quorum Sensing Controls Biofilm Formation in
Vibrio cholerae
through Modulation of Cyclic Di-GMP Levels and Repression of
vpsT
Authors: Waters Christopher M. , Lu Wenyun , Rabinowitz Joshua D. , Bassler Bonnie L. 
Published: January 2008

ABSTRACT

Two chemical signaling systems, quorum sensing (QS) and 3′,5′-cyclic diguanylic acid (c-di-GMP), reciprocally control biofilm formation in
Vibrio cholerae
. QS is the process by which bacteria communicate via the secretion and detection of autoinducers, and in
V. cholerae
, QS represses biofilm formation. c-di-GMP is an intracellular second messenger that contains information regarding local environmental conditions, and in
V. cholerae
, c-di-GMP activates biofilm formation. Here we show that HapR, a major regulator of QS, represses biofilm formation in
V. cholerae
through two distinct mechanisms. HapR controls the transcription of 14 genes encoding a group of proteins that synthesize and degrade c-di-GMP. The net effect of this transcriptional program is a reduction in cellular c-di-GMP levels at high cell density and, consequently, a decrease in biofilm formation. Increasing the c-di-GMP concentration at high cell density to the level present in the low-cell-density QS state restores biofilm formation, showing that c-di-GMP is epistatic to QS in the control of biofilm formation in
V. cholerae
. In addition, HapR binds to and directly represses the expression of the biofilm transcriptional activator,
vpsT
. Together, our results suggest that
V. cholerae
integrates information about the vicinal bacterial community contained in extracellular QS autoinducers with the intracellular environmental information encoded in c-di-GMP to control biofilm formation.
Analysis of the
Campylobacter jejuni
FlgR Response Regulator Suggests Integration of Diverse Mechanisms To Activate an NtrC-Like Protein
Authors: Joslin Stephanie N. , Hendrixson David R. 
Published: January 2008

ABSTRACT

Flagellar motility in
Campylobacter jejuni
mediates optimal interactions with human or animal hosts. σ
54
and the FlgSR two-component system are necessary for the expression of many
C. jejuni
flagellar genes. The FlgR response regulator is homologous to the NtrC family of transcriptional activators. These regulators usually contain an N-terminal receiver domain, a central domain that interacts with σ
54
and hydrolyzes ATP, and a DNA-binding C-terminal domain. Most often, phosphorylation of the receiver domain influences its inherent ability to either positively or negatively control the activity of the regulator. In this study, we performed genetic and biochemical analyses to understand how FlgR activity is controlled to culminate in the expression of σ
54
-dependent flagellar genes. Our data suggest that the FlgR receiver domain has the capacity for both positive and negative regulation in controlling the activation of the protein. Analysis of the C-terminal domain of FlgR revealed that it lacks a DNA-binding motif and is not required for σ
54
-dependent flagellar gene expression. Further analysis of FlgR lacking the C-terminal domain indicates that this protein is partially functional in the absence of the cognate sensor kinase, FlgS, but its activity is still dependent on the phosphorylated residue in the receiver domain, D51. We hypothesize that the C-terminal domain may not function to bind DNA but may ensure the specificity of the phosphorylation of FlgR by FlgS. Our results demonstrate that FlgR activation mechanisms are unusual among characterized NtrC-like proteins and emphasize that various means are utilized by the NtrC family of proteins to control the transcription of target genes.
Analysis of Tryptophan Residues in the Staphylococcal Multidrug Transporter QacA Reveals Long-Distance Functional Associations of Residues on Opposite Sides of the Membrane
Authors: Hassan Karl A. , Souhani Talal , Skurray Ronald A. , Brown Melissa H. 
Published: January 2008

ABSTRACT
Tryptophan residues can possess a multitude of functions within a multidrug transport protein, e.g., mediating interactions with substrates or distal parts of the protein, or fulfilling a structural requirement, such as guiding the depth of membrane insertion. In this study, the nine tryptophan residues of the staphylococcal QacA multidrug efflux protein were individually mutated to alanine and phenylalanine, and the functional consequences of these changes were determined. Phenylalanine substitutions for each tryptophan residue were functionally tolerated. However, alanine modifications revealed an important functional role for three tryptophan residues, W58, W149, and W173, each of which is well conserved among QacA-related transport proteins in the major facilitator superfamily. The most functionally compromising mutation, an alanine substitution for W58, likely to be located at the extracellular interface of transmembrane segment 2, abolished all detectable QacA-mediated resistance and transport function. Second-site suppressor analyses identified several mutations that rescued the function of the W58A QacA mutant. Remarkably, all of these suppressor mutations were shown to be located in cytoplasmic loops between transmembrane helices 2 and 3 or 12 and 13, demonstrating novel functional associations between amino acid positions on opposite sides of the membrane and in distal N- and C-terminal regions of the QacA protein.