bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization and Use of Catabolite-Repressed Promoters from Gluconate Genes in
Corynebacterium glutamicum
Authors: Letek Michal , Valbuena Noelia , Ramos Angelina , Ordóñez Efrén , Gil José A. , Mateos Luís M. 
Published: December 2005

ABSTRACT

The genes involved in gluconate catabolism (
gntP
and
gntK
) in
Corynebacterium glutamicum
are scattered in the chromosome, and no regulatory genes are apparently associated with them, in contrast with the organization of the
gnt
operon in
Escherichia coli
and
Bacillus subtilis
. In
C. glutamicum
,
gntP
and
gntK
are essential genes when gluconate is the only carbon and energy source. Both genes contain upstream regulatory regions consisting of a typical promoter and a hypothetical cyclic AMP (cAMP) receptor protein (CRP) binding region but lack the expected consensus operator region for binding of the GntR repressor protein. Expression analysis by Northern blotting showed monocistronic transcripts for both genes. The expression of
gntP
and
gntK
is not induced by gluconate, and the
gnt
genes are subject to catabolite repression by sugars, such as glucose, fructose, and sucrose, as was detected by quantitative reverse transcription-PCR (qRT-PCR). Specific analysis of the DNA promoter sequences (P
gntK
and P
gntP
) was performed using bifunctional promoter probe vectors containing
mel
(involved in melanin production) or e
gfp2
(encoding a green fluorescent protein derivative) as the reporter gene. Using this approach, we obtained results parallel to those from qRT-PCR. An applied example of in vivo gene expression modulation of the
divIVA
gene in
C. glutamicum
is shown, corroborating the possible use of the
gnt
promoters to control gene expression.
glxR
(which encodes GlxR, the hypothetical CRP protein) was subcloned from the
C. glutamicum
chromosomal DNA and overexpressed in corynebacteria; we found that the level of
gnt
expression was slightly decreased compared to that of the control strains. The purified GlxR protein was used in gel shift mobility assays, and a specific interaction of GlxR with sequences present on P
gntP
and P
gntK
fragments was detected only in the presence of cAMP.
Genetic Characterization of a Single Bifunctional Enzyme for Fumarate Reduction and Succinate Oxidation in
Geobacter sulfurreducens
and Engineering of Fumarate Reduction in
Geobacter metallireducens
Authors: Butler Jessica E. , Glaven Richard H. , Esteve-Núñez Abraham , Núñez Cinthia , Shelobolina Evgenya S. , Bond Daniel R. , Lovley Derek R. 
Published: December 2005

ABSTRACT

The mechanism of fumarate reduction in
Geobacter sulfurreducens
was investigated. The genome contained genes encoding a heterotrimeric fumarate reductase, FrdCAB, with homology to the fumarate reductase of
Wolinella succinogenes
and the succinate dehydrogenase of
Bacillus subtilis
. Mutation of the putative catalytic subunit of the enzyme resulted in a strain that lacked fumarate reductase activity and was unable to grow with fumarate as the terminal electron acceptor. The mutant strain also lacked succinate dehydrogenase activity and did not grow with acetate as the electron donor and Fe(III) as the electron acceptor. The mutant strain could grow with acetate as the electron donor and Fe(III) as the electron acceptor if fumarate was provided to alleviate the need for succinate dehydrogenase activity in the tricarboxylic acid cycle. The growth rate of the mutant strain under these conditions was faster and the cell yields were higher than for wild type grown under conditions requiring succinate dehydrogenase activity, suggesting that the succinate dehydrogenase reaction consumes energy. An orthologous
frdCAB
operon was present in
Geobacter metallireducens
, which cannot grow with fumarate as the terminal electron acceptor. When a putative dicarboxylic acid transporter from
G. sulfurreducens
was expressed in
G. metallireducens
, growth with fumarate as the sole electron acceptor was possible. These results demonstrate that, unlike previously described organisms,
G. sulfurreducens
and possibly
G. metallireducens
use the same enzyme for both fumarate reduction and succinate oxidation in vivo.
The Scc Spirochetal Coiled-Coil Protein Forms Helix-Like Filaments and Binds to Nucleic Acids Generating Nucleoprotein Structures
Authors: Mazouni Khalil , Pehau-Arnaudet Gérard , England Patrick , Bourhy Pascale , Saint Girons Isabelle , Picardeau Mathieu 
Published: December 2005

ABSTRACT

The analysis of the genome of
Leptospira
spp., a group of bacteria of the phylum of spirochetes with several unique evolutionary and morphological features, has allowed the identification of a gene encoding a coiled-coil protein, called Scc, which is completely unrelated to any other eukaryotic or prokaryotic protein. Since coiled-coil proteins are often key elements of the cytoskeleton, we analyzed the protein Scc, which is a 24-kDa protein composed of a N-terminal coiled-coil domain, a proline-rich intermediate domain, and an acidic tail. The gene
scc
is located in an operon which also contains the genes encoding the initiation factor IF3 and the two ribosomal proteins L20 and L35. In this study, we showed that the presence of the coiled-coil domain was responsible for the polymerization of Scc in helix-like structures, in an ATP-independent manner, in both
Escherichia coli
living cells and in vitro. Analysis of the Scc polymers by electron microscopy showed filaments with a width of 6 to 10 nm, similar to that of eukaryotic intermediate filaments. Scc was also found to bind both RNA and double-stranded DNA without detectable sequence specificity. By electron microscopy, we showed that Scc polymer assembly was affected by the presence of nucleic acids, giving rise to rod-shaped structures with a width ranging from 45 to 155 nm. Finally,
Leptospira biflexa
cells depleted in Scc form small colonies, but the morphology of their helicoidal cell body was not affected. These results provide the first insight into a unique DNA binding filament-forming coiled-coil protein that could play an important role in the subcellular architecture of the spirochetal microorganism.
Characterization of the Opposing Roles of H-NS and TraJ in Transcriptional Regulation of the F-Plasmid
tra
Operon
Authors: Will William R. , Frost Laura S. 
Published: December 2005

ABSTRACT

The transfer (
tra
) operon of the conjugative F plasmid of
Escherichia coli
is a polycistronic 33-kb operon which encodes most of the proteins necessary for F-plasmid transfer. Here, we report that transcription from P
Y
, the
tra
operon promoter, is repressed by the host nucleoid-associated protein, H-NS. Electrophoretic mobility shift assays indicate that H-NS binds preferentially to the
tra
promoter region, while Northern blot and transcriptional fusion analyses indicate that transcription of
traY
, the first gene in the
tra
operon, is derepressed in an
hns
mutant throughout growth. The plasmid-encoded regulatory protein TraJ is essential for transcription of the
tra
operon in wild-type
Escherichia coli
; however, TraJ is not necessary for plasmid transfer or
traY
operon transcription in an
hns
mutant. This indicates that H-NS represses transcription from P
Y
directly and not indirectly via its effects on TraJ levels. These results suggest that TraJ functions to disrupt H-NS silencing at P
Y
, allowing transcription of the
tra
operon.
Characterization of Dissimilatory Fe(III) versus NO3−Reduction in the Hyperthermophilic ArchaeonPyrobaculum aerophilum
Authors: Feinberg Lawrence F. , Holden James F. 
Published: December 2005

ABSTRACTThe hyperthermophilic archaeonPyrobaculum aerophilumused 20 mM Fe(III) citrate, 100 mM poorly crystalline Fe(III) oxide, and 10 mM KNO3as terminal electron acceptors. The two forms of iron were reduced at different rates but with equal growth yields. The insoluble iron was reduced when segregated spatially by dialysis tubing, indicating that direct contact with the iron was not necessary for growth. When partitioned, there was no detectable Fe(III) or Fe(II) outside of the tubing after growth, suggesting that an electron shuttle, not a chelator, may be used as an extracellular mediator of iron reduction. The addition of 25 and 50% (vol vol−1) cell-free spent insoluble iron media to fresh media led to growth without a lag phase. Liquid chromatography analysis of spent media showed that cultures grown in iron, especially insoluble iron, produced soluble extracellular compounds that were absent or less abundant in spent nitrate medium. NADH-dependent ferric reductase activity increased approximately 100-fold, while nitrate reductase activity decreased 10-fold in whole-cell extracts from iron-grown cells relative to those from nitrate-grown cells, suggesting that dissimilatory iron reduction was regulated. A novel 2,6-anthrahydroquinone disulfonate oxidase activity was more than 580-fold higher in iron-grown cells than in nitrate-grown cells. The activity was primarily (>95%) associated with the membrane cellular fraction, but its physiological function is unknown. Nitrate-grown cultures produced two membrane-bound,c-type cytochromes that are predicted to be monoheme and part of nitrite reductase and abc1complex using genome analyses. Only one cytochrome was present in cells grown on Fe(III) citrate whose relative abundance was unchanged.
Synthesis, Stability, and Function of Cytadhesin P1 and Accessory Protein B/C Complex of
Mycoplasma pneumoniae
Authors: Waldo Robert H. , Krause Duncan C. 
Published: December 2005

ABSTRACT

The genes MPN141 and MPN142 encode the major adhesin P1 and the cytadherence-related B/C proteins (P90/P40), respectively, in
Mycoplasma pneumoniae
. Using reverse transcriptase PCR we found open reading frames MPN140 to MPN142 constitute a polycistronic transcriptional unit. Cytadherence mutant IV-22 has a frameshift mutation in MPN141 and lacks the P1, B, or C proteins. Recombinant MPN141 and/or MPN142 were introduced into mutant IV-22 by transposon delivery in several configurations, and the levels of the P1, B, and C proteins were assessed by immunoblotting. MPN142 in mutant IV-22 has a wild-type nucleotide sequence, yet the introduction of recombinant MPN141 alone to mutant IV-22, although it restored P1 levels, failed to restore levels of B or C. In contrast, recombinant MPN141 and MPN142 delivered in
cis
or in
trans
were sufficient to restore all three proteins. Taken together, our data indicated that some but not all synthesis of B or C is dependent on coupling to the translation of P1 immediately upstream of MPN142 and demonstrated that proteins B and C are not stable in the absence of P1. The linkage of MPN141 and MPN142 at the levels of transcription, translation, and protein stability, in addition to their previously demonstrated colocalization and the requirement of B and/or C for P1 function, reinforces the conclusion that these proteins constitute a multiprotein complex that functions in receptor binding.
The β-Oxidation Systems ofEscherichia coliandSalmonella entericaAre Not Functionally Equivalent
Authors: Iram Surtaj Hussain , Cronan John E. 
Published: December 2005

ABSTRACTBased on its genome sequence, the pathway of β-oxidative fatty acid degradation inSalmonella entericaserovar Typhimurium LT2 has been thought to be identical to the well-characterizedEscherichia coliK-12 system. We report that wild-type strains ofS. entericagrow on decanoic acid, whereas wild-typeE. colistrains cannot. Mutant strains (carryingfadR) of both organisms in which the genes of fatty acid degradation (fad) are expressed constitutively are readily isolated. TheS. enterica fadRstrains grow more rapidly than the wild-type strains on decanoic acid and also grow well on octanoic and hexanoic acids (which do not support growth of wild-type strains). By contrast,E. coli fadRstrains grow well on decanoic acid but grow only exceedingly slowly on octanoic acid and fail to grow at all on hexanoic acid. The two wild-type organisms also differed in the ability to grow on oleic acid when FadR was overexpressed. Under these superrepression conditions,E. colifailed to grow, whereasS. entericagrew well. Exchange of the wild-typefadRgenes between the two organisms showed this to be a property ofS. entericarather than of the FadR proteins per se. This difference in growth was attributed toS. entericahaving higher cytosolic levels of the inducing ligands, long-chain acyl coenzyme As (acyl-CoAs). The most striking results were the differences in the compositions of CoA metabolites of strains grown with octanoic acid or oleic acid.S. entericacleanly converted all of the acid to acetyl-CoA, whereasE. coliaccumulated high levels of intermediate-chain-length products. Exchange of homologous genes between the two organisms showed that theS. entericaFadE and FadBA enzymes were responsible for the greater efficiency of β-oxidation relative to that ofE. coli.
Accumulation of Homolanthionine and Activation of a Novel Pathway for Isoleucine Biosynthesis in Corynebacterium glutamicumMcbR Deletion Strains
Authors: Krömer Jens Olaf , Heinzle Elmar , Schröder Hartwig , Wittmann Christoph 
Published: December 2005

ABSTRACTIn the present work, the metabolic consequences of the deletion of the methionine and cysteine biosynthesis repressor protein (McbR) inCorynebacterium glutamicum, which releases almost all enzymes of methionine biosynthesis and sulfate assimilation from transcriptional regulation (D. A. Rey, A. Pühler, and J. Kalinowski, J. Biotechnol.103:51-65, 2003), were studied.C. glutamicumATCC 13032 ΔmcbRshowed no overproduction of methionine. Metabolome analysis revealed drastic accumulation of a single metabolite, which was not present in the wild type. It was identified by isotopic labeling studies and gas chromatography/mass spectrometry asl-homolanthionine {S-[(3S)-3-amino-3-carboxypropyl]-l-homocysteine}. The accumulation of homolanthionine to an intracellular concentration of 130 mM in the ΔmcbRstrain was accompanied by an elevated intracellular homocysteine level. It was shown that cystathionine-γ-synthase (MetB) produced homolanthionine as a side reaction. MetB showed higher substrate affinity for cysteine (Km= 260 μM) than for homocysteine (Km= 540 μM). The cell is able to cleave homolanthionine at low rates via cystathionine-β-lyase (MetC). This cleavage opens a novel threonine-independent pathway for isoleucine biosynthesis via 2-oxobutanoate formed by MetC. In fact, the deletion mutant exhibited an increased intracellular isoleucine level. Metabolic flux analysis ofC. glutamicum ΔmcbRrevealed that only 24% of theO-acetylhomoserine at the entry of the methionine pathway is utilized for methionine biosynthesis; the dominating fraction is either stored as homolanthionine or redirected towards the formation of isoleucine. Deletion ofmetBcompletely prevents homolanthionine accumulation, which is regarded as an important step in the development ofC. glutamicumstrains for biotechnological methionine production.
The Genome Sequence ofMethanosphaera stadtmanaeReveals Why This Human Intestinal Archaeon Is Restricted to Methanol and H2for Methane Formation and ATP Synthesis
Authors: Fricke Wolfgang F. , Seedorf Henning , Henne Anke , Krüer Markus , Liesegang Heiko , Hedderich Reiner , Gottschalk Gerhard , Thauer Rudolf K. 
Published: December 2005

ABSTRACTMethanosphaera stadtmanaehas the most restricted energy metabolism of all methanogenic archaea. This human intestinal inhabitant can generate methane only by reduction of methanol with H2and is dependent on acetate as a carbon source. We report here the genome sequence ofM. stadtmanae, which was found to be composed of 1,767,403 bp with an average G+C content of 28% and to harbor only 1,534 protein-encoding sequences (CDS). The genome lacks 37 CDS present in the genomes of all other methanogens. Among these are the CDS for synthesis of molybdopterin and for synthesis of the carbon monoxide dehydrogenase/acetyl-coenzyme A synthase complex, which explains whyM. stadtmanaecannot reduce CO2to methane or oxidize methanol to CO2and why this archaeon is dependent on acetate for biosynthesis of cell components. Four sets ofmtaABCgenes coding for methanol:coenzyme M methyltransferases were found in the genome ofM. stadtmanae. These genes exhibit homology tomtagenes previously identified inMethanosarcinaspecies. TheM. stadtmanaegenome also contains at least 323 CDS not present in the genomes of all other archaea. Seventy-three of these CDS exhibit high levels of homology to CDS in genomes of bacteria and eukaryotes. These 73 CDS include 12 CDS which are unusually long (>2,400 bp) with conspicuous repetitive sequence elements, 13 CDS which exhibit sequence similarity on the protein level to CDS encoding enzymes involved in the biosynthesis of cell surface antigens in bacteria, and 5 CDS which exhibit sequence similarity to the subunits of bacterial type I and III restriction-modification systems.
Microarrays Reveal that Each of the Ten Dominant Lineages ofStaphylococcus aureusHas a Unique Combination of Surface-Associated and Regulatory Genes
Authors: Lindsay Jodi A. , Moore Catrin E. , Day Nicholas P. , Peacock Sharon J. , Witney Adam A. , Stabler Richard A. , Husain Sarah E. , Butcher Philip D. , Hinds Jason 
Published: December 2005

ABSTRACTStaphylococcus aureusis the most common cause of hospital-acquired infection. In healthy hosts outside of the health care setting,S. aureusis a frequent colonizer of the human nose but rarely causes severe invasive infection such as bacteremia, endocarditis, or osteomyelitis. To identify genes associated with community-acquired invasive isolates, regions of genomic variability, and theS. aureuspopulation structure, we compared 61 community-acquired invasive isolates ofS. aureusand 100 nasal carriage isolates from healthy donors using a microarray spotted with PCR products representing every gene from the sevenS. aureussequencing projects. The core genes common to all strains were identified, and 10 dominant lineages ofS. aureuswere clearly discriminated. Each lineage carried a unique combination of hundreds of “core variable” (CV) genes scattered throughout the chromosome, suggesting a common ancestor but early evolutionary divergence. Many CV genes are regulators of virulence genes or known or predicted to be expressed on the bacterial surface and to interact with the host during nasal colonization and infection. Within each lineage, isolates showed substantial variation in the carriage of mobile genetic elements and their associated virulence and resistance genes, indicating frequent horizontal transfer. However, we were unable to identify any association between lineage or gene and invasive isolates. We suggest that theS. aureusgene combinations necessary for invasive disease may also be necessary for nasal colonization and that community-acquired invasive disease is strongly dependent on host factors.
Whole-Genome Plasticity among
Mycobacterium avium
Subspecies: Insights from Comparative Genomic Hybridizations
Authors: Wu Chia-wei , Glasner Jeremy , Collins Michael , Naser Saleh , Talaat Adel M. 
Published: December 2005

ABSTRACT

Infection with
Mycobacterium avium
subsp.
paratuberculosis
causes Johne's disease in cattle and is also implicated in cases of Crohn's disease in humans. Another closely related strain,
M. avium
subsp.
avium
, is a health problem for immunocompromised patients. To understand the molecular pathogenesis of
M. avium
subspecies, we analyzed the genome contents of isolates collected from humans and domesticated or wildlife animals. Comparative genomic hybridizations indicated distinct lineages for each subspecies where the closest genomic relatedness existed between
M. avium
subsp.
paratuberculosis
isolates collected from human and clinical cow samples. Genomic islands (
n
= 24) comprising 846 kb were present in the reference
M. avium
subsp.
avium
strain but absent from 95% of
M. avium
subsp.
paratuberculosis
isolates. Additional analysis identified a group of 18
M. avium
subsp.
paratuberculosis
-associated islands comprising 240 kb that were absent from most of the
M. avium
subsp.
avium
isolates. Sequence analysis of DNA regions flanking the genomic islands identified three large inversions in addition to several small inversions that could play a role in regulation of gene expression. Analysis of genes encoded in the genomic islands reveals factors that are probably important for various mechanisms of virulence. Overall,
M. avium
subsp.
avium
isolates displayed a higher level of genomic diversity than
M. avium
subsp.
paratuberculosis
isolates. Among
M. avium
subsp.
paratuberculosis
isolates, those from wildlife animals displayed the highest level of genomic rearrangements that were not observed in other isolates. The presented findings will affect the future design of diagnostics and vaccines for Johne's and Crohn's diseases and provide a model for genomic analysis of closely related bacteria.
DNA Sequence of a ColV Plasmid and Prevalence of Selected Plasmid-Encoded Virulence Genes among Avian
Escherichia coli
Strains
Authors: Johnson Timothy J. , Siek Kylie E. , Johnson Sara J. , Nolan Lisa K. 
Published: December 2005

ABSTRACT

ColV plasmids have long been associated with the virulence of
Escherichia coli
, despite the fact that their namesake trait, ColV production, does not appear to contribute to virulence. Such plasmids or their associated sequences appear to be quite common among avian pathogenic
E. coli
(APEC) and are strongly linked to the virulence of these organisms. In the present study, a 180-kb ColV plasmid was sequenced and analyzed. This plasmid, pAPEC-O2-ColV, possesses a 93-kb region containing several putative virulence traits, including
iss
,
tsh
, and four putative iron acquisition and transport systems. The iron acquisition and transport systems include those encoding aerobactin and salmochelin, the
sit
ABC iron transport system, and a putative iron transport system novel to APEC,
eit
. In order to determine the prevalence of the virulence-associated genes within this region among avian
E. coli
strains, 595 APEC and 199 avian commensal
E. coli
isolates were examined for genes of this region using PCR. Results indicate that genes contained within a portion of this putative virulence region are highly conserved among APEC and that the genes of this region occur significantly more often in APEC than in avian commensal
E. coli
. The region of pAPEC-O2-ColV containing genes that are highly prevalent among APEC appears to be a distinguishing trait of APEC strains.
Quorum-Sensing Signal Synthesis by theYersinia pestisAcyl-Homoserine Lactone Synthase YspI
Authors: Kirwan J. Paul , Gould Ty A. , Schweizer Herbert P. , Bearden Scott W. , Murphy Robert C. , Churchill Mair E. A. 
Published: December 2005

ABSTRACTThe acyl-homoserine lactone molecular species (AHLs) produced by theYersinia pestisAHL synthase YspI were identified by biochemical and physical/chemical techniques. Bioassays of extracts from culture supernatants of the recombinant YspI and wild-typeYersinia pestisshowed similar profiles of AHLs. Analysis by liquid chromatography-mass spectrometry revealed that the predominant AHLs wereN-3-oxooctanoyl-l-homoserine lactone andN-3-oxo-hexanoyl-l-homoserine lactone.
Autorepression of RctB, an Initiator of
Vibrio cholerae
Chromosome II Replication
Authors: Egan Elizabeth S. , Duigou Stephane , Waldor Matthew K. 
Published: December 2005

ABSTRACT

The RctB protein binds to the origin of replication of
Vibrio cholerae
chromosome II (chrII) and is required for

oriCII
Vc

-based replication. Here, we found that RctB acts as an autorepressor, inhibiting
rctB
transcription. Integration host factor promotes
rctB
transcription, while Dam and DnaA, factors required for replication of both
V. cholerae
chromosomes, influence RctB autorepression. Thus, RctB appears to regulate chrII replication as both an initiator and a transcription repressor, and its synthesis is modulated by factors that govern replication of both chromosomes.