bluuebunny/crossref_metadata_2025_split huggingface dataset:
Complex Transcriptional Control Links NikABCDE-Dependent Nickel Transport with Hydrogenase Expression in
Escherichia coli
Authors: Rowe Jessica L. , Starnes G. Lucas , Chivers Peter T. 
Published: September 2005

ABSTRACT

Escherichia coli
requires nickel under anaerobic growth conditions for the synthesis of catalytically active NiFe hydrogenases. Transcription of the NikABCDE nickel transporter, which is required for NiFe hydrogenase synthesis, was previously shown to be upregulated by FNR (fumarate-nit rate regulator) in the absence of oxygen and repressed by the NikR repressor in the presence of high extracellular nickel levels. We present here a detailed analysis of
nikABCDE
transcriptional regulation and show that it closely correlates with hydrogenase expression levels. We identify a nitrate-dependent mechanism for
nikABCDE
repression that is linked to the NarLX two-component system. NikR is functional under all nickel conditions tested, but its activity is modulated by the total nickel concentration present as well as by one or more components of the hydrogenase assembly pathway. Unexpectedly, NikR function is independent of NikABCDE function, suggesting that NikABCDE is a hydrogenase-specific nickel transporter, consistent with its original identification as a hydrogenase (
hyd
) mutant. Further, the results suggest that the hydrogenase assembly pathway is sequestered within the cell. A second nickel import pathway in
E. coli
is implicated in NikR function.
Purification and Characterization of the N-Terminal Domain of ExeA: a Novel ATPase Involved in the Type II Secretion Pathway of
Aeromonas hydrophila
Authors: Schoenhofen Ian C. , Li Gang , Strozen Timothy G. , Howard S. Peter 
Published: September 2005

ABSTRACT

Aeromonas hydrophila
secretes a number of degradative enzymes and toxins into the external milieu via the type II secretory pathway or secreton. ExeA is an essential component of this system and is necessary for the localization and/or multimerization of the secretin ExeD. ExeA contains two sequence motifs characteristic of the Walker superfamily of ATPases. Previous examination of substitution derivatives altered in these motifs suggested that ATP binding or hydrolysis is required for ExeAB complex formation and subsequent secretion function. To directly examine ExeA function, the N-terminal cytoplasmic domain of ExeA with the addition of a C-terminal hexahistidine tag (cytExeA) was overproduced in
Escherichia coli
and purified by metal chelate affinity and anion-exchange chromatographic techniques. Purified preparations of cytExeA exhibited ATPase activity in the presence of several divalent cations, Mg
2+
being the preferred cation, with an optimum reaction temperature of ∼37 to 42°C and an optimum pH of 7 to 8. cytExeA exhibited an apparent

K
m

for Mg-ATP of 0.22 mM and a
V
max
of 0.72 nmol min
−1
mg
−1
of protein. cytExeA displayed low specificity for nucleoside triphosphate substrates and was significantly inhibited by F-type ATPase inhibitors. Gel filtration analyses of cytExeA, ExeA, and ExeAB indicated that ExeA dimerizes and forms a very large complex with ExeB. These findings support a model whereby ExeAB utilizes energy derived from ATP hydrolysis to facilitate the correct localization and multimerization of the ExeD secretin.
Partial Restoration of Antibacterial Activity of the Protein Encoded by a Cryptic Open Reading Frame (
cyt1Ca
) from
Bacillus thuringiensis
subsp.
israelensis
by Site-Directed Mutagenesis
Authors: Itsko Mark , Manasherob Robert , Zaritsky Arieh 
Published: September 2005

ABSTRACT

Insecticidal crystal proteins of
Bacillus thuringiensis
belong to two unrelated toxin families: receptor-specific Cry toxins against insects and Cyt toxins that lyse a broad range of cells, including bacteria, via direct binding to phospholipids. A new
cyt
-like open reading frame (
cyt1Ca
) encoding a 60-kDa protein, has recently been discovered (C. Berry et al., Appl. Environ. Microbiol.
68:
5082-5095, 2002). Cyt1Ca displays the structure of a two-domain fusion protein: the N-terminal moiety resembles the full-length Cyt toxins, and the C-terminal moiety is similar to the receptor-binding domains of several ricin-like toxins, such as Mtx1. Neither the larvicidal activity of
cyt1Ca
expressed in
Escherichia coli
nor the hemolytic effect of His-tagged purified Cyt1Ca has been observed (R. Manasherob et al., unpublished). This was attributed to five amino acid differences between the sequences of its N-terminal moiety and Cyt1Aa. The 3′ end of
cyt1Ca
was truncated (removing the ricin-binding domain of Cyt1Ca), and six single bases were appropriately changed by site-directed mutagenesis, sequentially replacing the noncharged amino acids by charged ones, according to Cyt1Aa, to form several versions. Expression of these mutated
cyt1Ca
versions caused loss of the colony-forming ability of the corresponding
E. coli
cells to different extents compared with the original gene. In some mutants this antibacterial effect was associated by significant distortion of cell morphology and in others by generation of multiple inclusion bodies spread along the cell envelope. The described deleterious effects of mutated
cyt1Ca
versions against
E. coli
may reflect an evolutionary relationship between Cyt1Aa and Cyt1Ca.
Revisiting the Evolution of
Mycobacterium bovis
Authors: Mostowy Serge , Inwald Jackie , Gordon Steve , Martin Carlos , Warren Rob , Kremer Kristin , Cousins Debby , Behr Marcel A. 
Published: September 2005

ABSTRACT

Though careful consideration has been placed towards genetic characterization of tubercle bacillus isolates causing disease in humans, those causing disease predominantly among wild and domesticated mammals have received less attention. In contrast to
Mycobacterium tuberculosis
, whose host range is largely specific to humans,
M. bovis
and “
M bovis
-like” organisms infect a broad range of animal species beyond their most prominent host in cattle. To determine whether strains of variable genomic content are associated with distinct distributions of disease, the DNA contents of
M. bovis
or
M. bovis
-like isolates from a variety of hosts were investigated via Affymetrix GeneChip. Consistent with previous genomic analysis of the
M. tuberculosis
complex (MTC), large sequence polymorphisms of putative diagnostic and biological consequence were able to unambiguously distinguish interrogated isolates. The distribution of deleted regions indicates organisms genomically removed from
M. bovis
and also points to structured genomic variability within
M. bovis
. Certain genomic profiles spanned a variety of hosts but were clustered by geography, while others associated primarily with host type. In contrast to the prevailing assumption that
M. bovis
has broad host capacity, genomic profiles suggest that distinct MTC lineages differentially infect a variety of mammals. From this, a phylogenetic stratification of genotypes offers a predictive framework upon which to base future genetic and phenotypic studies of the MTC.
Homologies and Divergences in the Transcription Regulatory System of Two RelatedBacillus subtilisPhages
Authors: Pérez-Lago Laura , Salas Margarita , Camacho Ana 
Published: September 2005

ABSTRACTTranscription regulation relies on the molecular interplay between the RNA polymerase and regulatory factors. Phages of the φ29-like genus encode two regulatory proteins, p4 and p6. In φ29, the switch from early to late transcription is based on the synergistic binding of proteins p4 and p6 to the promoter sequence, resulting in a nucleosome-like structure able to synergize or antagonize the binding of RNAP. We show that a nucleosome-like structure of p4 and p6 is also formed in the related phage Nf and that this structure is responsible for the coordinated control of the early and late promoters. However, in spite of their homologies, the transcriptional regulators are not interchangeable, and only when all of the components of the Nf regulatory system are present is fully active transcriptional regulation of the Nf promoters achieved.
A Gene Encoding a Holin-Like Protein Involved in Spore Morphogenesis and Spore Germination inBacillus subtilis
Authors: Real Gonçalo , Pinto Sérgio M. , Schyns Ghislain , Costa Teresa , Henriques Adriano O. , Moran Charles P. 
Published: September 2005

ABSTRACTWe report here studies of expression and functional analysis of aBacillus subtilisgene,ywcE, which codes for a product with features of a holin. Primer extension analysis ofywcEtranscription revealed that a single transcript accumulated from the onset of sporulation onwards, produced from a σA-type promoter bearing the TG dinucleotide motif of “extended” −10 promoters. No primer extension product was detected in vivo during growth. However, specific runoff products were produced in vitro from theywcEpromoter by purified σA-containing RNA polymerase (EσA), and the in vivo and in vitro transcription start sites were identical. These results suggested that utilization of theywcEpromoter by EσAduring growth was subjected to repression. Studies with alacZfusion revealed that the transition-state regulator AbrB repressed the transcription ofywcEduring growth. This repression was reversed at the onset of sporulation in a Spo0A-dependent manner, but Spo0A did not appear to contribute otherwise toywcEtranscription. We foundywcEto be required for proper spore morphogenesis. Spores of theywcEmutant showed a reduced outer coat which lacked the characteristic striated pattern, and the outer coat failed to attach to the underlying inner coat. The mutant spores also accumulated reduced levels of dipicolinic acid.ywcEwas also found to be important for spore germination.
Modeling the Effects of
prl
Mutations on the
Escherichia coli
SecY Complex
Authors: Smith Margaret A. , Clemons William M. , DeMars Cathrine J. , Flower Ann M. 
Published: September 2005

ABSTRACT

The apparatus responsible for translocation of proteins across bacterial membranes is the conserved SecY complex, consisting of SecY, SecE, and SecG. Prior genetic analysis provided insight into the mechanisms of protein export, as well as the interactions between the component proteins. In particular, the
prl
suppressor alleles of
secE
and
secY
, which allow export of secretory proteins with defective signal sequences, have proven particularly useful. Here, we report the isolation of novel mutations in
secE
and
secY
, as well as the phenotypic effects of combinations of
prl
mutations. These new alleles, as well as previously characterized
prl
mutations, were analyzed in light of the recently published crystal structure of the archaeal SecY complex. Our results support and expand a model of Prl suppressor activity that proposes that all of the
prlA
and
prlG
alleles either destabilize the closed state of the channel or stabilize the open form. These mutants thus allow channel opening to occur without the triggering event of signal sequence binding that is required in a wild-type complex.
The pCoo Plasmid of Enterotoxigenic
Escherichia coli
Is a Mosaic Cointegrate
Authors: Froehlich Barbara , Parkhill Julian , Sanders Mandy , Quail Michael A. , Scott June R. 
Published: September 2005

ABSTRACT

CS1 is the prototype of a class of pili of enterotoxigenic
Escherichia coli
(ETEC) associated with diarrheal disease in humans. The genes encoding this pilus are carried on a large plasmid, pCoo. We report the sequence of the complete 98,396-bp plasmid. Like many other virulence plasmids, pCoo is a mosaic consisting of regions derived from multiple sources. Complete and fragmented insertion sequences (IS) make up 24% of the total DNA and are scattered throughout the plasmid. The pCoo DNA between these IS elements has a wide range of G+C content (35 to 57%), suggesting that these regions have different ancestries. We find that the pCoo plasmid is a cointegrate of two functional replicons, related to R64 and R100, which are joined at a 1,953-bp direct repeat of IS
100
. Recombination between these repeats in the cointegrate generates the two smaller replicons which coexist with the cointegrate in the culture. Both of the smaller replicons have plasmid stability genes as well as genes that may be important in pathogenesis. Examination by PCR of 17 other unrelated CS1 ETEC strains with a variety of serotypes demonstrated that all contained at least parts of both replicons of pCoo and that strains of the O6 genotype appear to contain a cointegrate very similar to pCoo. The results suggest that this family of CS1-encoding plasmids is evolving rapidly.
Activation of thephzOperon ofPseudomonas fluorescens2-79 Requires the LuxR Homolog PhzR,N-(3-OH-Hexanoyl)-l-Homoserine Lactone Produced by the LuxI Homolog PhzI, and acis-ActingphzBox
Authors: Khan Sharik R. , Mavrodi Dmitri V. , Jog Geetanjali J. , Suga Hiroaki , Thomashow Linda S. , Farrand Stephen K. 
Published: September 2005

ABSTRACTThephzoperon ofPseudomonas fluorescens2-79, which produces phenazine-1-carboxylate, is preceded by two genes,phzRandphzI, that are homologs of quorum-sensing gene pairs of theluxR-luxIfamily. DeletingphzRandphzIfrom strain 2-79 led to loss of production of the antibiotics, as well as a suite of six acyl-homoserine lactones (acyl-HSLs) that includes four 3-hydroxy- derivatives and two alkanoyl-HSLs. Strain 2-79 accumulatesN-(3-hydroxy-hexanoyl)-l-HSL to levels 20 and 30 times those ofN-(hexanoyl)-l-HSL andN-(3-hydroxy-octanoyl)-HSL, the next most abundant species produced by this isolate. Expression of a clone ofphzIinEscherichia coliandP. fluorescens1855 resulted in the synthesis of all six acyl-HSLs. Maximal activation ofphzAandphzRfused tolacZanduidAreporters, respectively, required PhzR and the acyl-HSL signals. PhzR-mediated expression of thephzA::lacZfusion responded with highest sensitivity and greatest magnitude to pureN-(3-hydroxy-hexanoyl)-l-HSL. When exposed to organic extracts of culture supernatants containing the six acyl-HSLs at their normal levels, the reporter responded strongly toN-(3-hydroxy-hexanoyl)-l-HSL but did not respond to any of the other five acyl-HSLs. The transcriptional start sites for the divergently orientedphzAandphzRgenes were mapped by primer extension analysis. An 18-bp almost perfect inverted repeat, thephzbox, is located between thephzIandphzRpromoters. Disrupting this repeat abolished PhzR-dependent activation ofphzAandphzR. We conclude that PhzI of strain 2-79 synthesizes 3-OH acyl-HSLs and thatP. fluorescens2-79 usesN-(3-hydroxy-hexanoyl)-HSL as its quorum-sensing signal. We also conclude that PhzR, with its quormone, activates expression ofphzAandphzRand that this activation requires an intactphzbox sequence located in the divergent promoter region.
Deletion of the Gene
rpoZ
, Encoding the ω Subunit of RNA Polymerase, in
Mycobacterium smegmatis
Results in Fragmentation of the β′ Subunit in the Enzyme Assembly
Authors: Mathew Renjith , Ramakanth Madhugiri , Chatterji Dipankar 
Published: September 2005

ABSTRACT

A deletion mutation in the gene
rpoZ
of
Mycobacterium smegmatis
causes reduced growth rate and a change in colony morphology. During purification of RNA polymerase from the mutant strain, the β′ subunit undergoes fragmentation but the fragments remain associated with the enzyme and maintain it in an active state until the whole destabilized assembly breaks down in the final step of purification. Complementation of the mutant strain with an integrated copy of the wild-type
rpoZ
brings back the wild-type colony morphology and improves the growth rate and activity of the enzyme, and the integrity of the β′ subunit remains unaffected.
The MutS C Terminus Is Essential for Mismatch Repair Activity In Vivo
Authors: Calmann Melissa A. , Nowosielska Anetta , Marinus M. G. 
Published: September 2005

ABSTRACT

An
Escherichia coli
K-12 strain was constructed with a chromosomal deletion (
mutS
Δ
800
) in the
mutS
gene that produced the removal of the C-terminal 53 amino acids which are not present in the MutS crystal structure. This strain has a MutS null phenotype for mutation avoidance, antirecombination, and sensitivity to cytotoxic agents in a
dam
mutant background.
Prevalence of Surface Swarming Behavior in
Salmonella
Authors: Kim Wook , Surette Michael G. 
Published: September 2005

ABSTRACT

Swarming behavior among 167
Salmonella
sp. isolates, representing all eight groups, was assessed. Only eight strains failed to swarm under standard conditions. Four of the defective strains swarmed on alternate carbon sources, and four harbored general defects in motility or lipopolysaccharide. Thus, swarming may represent an evolutionarily conserved behavior in
Salmonella
spp.
Requirements for Heterologous Production of a Complex Metalloenzyme: the Membrane-Bound [NiFe] Hydrogenase
Authors: Lenz Oliver , Gleiche Andrea , Strack Angelika , Friedrich Bärbel 
Published: September 2005

ABSTRACT

By taking advantage of the tightly clustered genes for the membrane-bound [NiFe] hydrogenase of
Ralstonia eutropha
H16, broad-host-range recombinant plasmids were constructed carrying the entire membrane-bound hydrogenase (MBH) operon encompassing 21 genes. We demonstrate that the complex MBH biosynthetic apparatus is actively produced in hydrogenase-free hosts yielding fully assembled and functional MBH protein.
Iron Starvation Leads to Oxidative Stress in
Anabaena
sp. Strain PCC 7120
Authors: Latifi Amel , Jeanjean Robert , Lemeille Sylvain , Havaux Michel , Zhang Cheng-Cai 
Published: September 2005

ABSTRACT

We establish here that iron deficiency causes oxidative stress in the cyanobacterium
Anabaena
sp. strain PCC 7120. Iron starvation leads to a significant increase in reactive oxygen species, whose effect can be abolished by treatment with the antioxidant tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine 1-oxyl). Oxidative stress induced by iron starvation could be a common feature of photosynthetic bacteria.
Yersinia pestis
Is Viable with Endotoxin Composed of Only Lipid A
Authors: Tan Li , Darby Creg 
Published: September 2005

ABSTRACT

Lipopolysaccharide (LPS) is the major outer membrane component of gram-negative bacteria. The minimal LPS structure for viability of
Escherichia coli
and
Salmonella enterica
serovar Typhimurium is lipid A glycosylated with 3-deoxy-D-manno-octulosonic acid (Kdo) residues. Here we show that another member of the
Enterobacteriaceae
,
Yersinia pestis
, can survive without Kdo in its LPS.
Characterization of Toxigenic
Corynebacterium ulcerans
Strains Isolated from Humans and Domestic Cats in the United Kingdom
Authors: De Zoysa Aruni , Hawkey Peter M. , Engler Kathy , George Robert , Mann Gina , Reilly William , Taylor David , Efstratiou Androulla 
Published: September 2005

ABSTRACT

In the United Kingdom there has been a marked increase in the number of human infections caused by toxigenic
Corynebacterium ulcerans
. During 2002 and 2003 the organism was also isolated from several domestic cats with bilateral nasal discharge. As
C. ulcerans
has never previously been isolated from cats, the 16S rRNA gene from three cat isolates was sequenced to confirm their species identities. Fifty clinical isolates from the United Kingdom isolated from 1986 to 2003 and seven cat isolates were characterized by ribotyping to determine whether the ribotypes of the cat isolates were genotypically related to those found for human clinical isolates. For comparison, the genotypes of 11 overseas isolates and 13 isolates from H. R. Carne's collection isolated between 1933 and 1979 were also determined. Strains isolated from domestic cats were found to exhibit the predominant ribotypes observed among human clinical isolates, suggesting that
C. ulcerans
isolated from cats could be a potential reservoir for human infection.
Genetic Analysis of Noroviruses in Chiba Prefecture, Japan, between 1999 and 2004
Authors: Okada Mineyuki , Ogawa Tomoko , Kaiho Ikuo , Shinozaki Kuniko 
Published: September 2005

ABSTRACT
Noroviruses (NVs) are common pathogens that consist of genetically divergent viruses that induce gastroenteritis in humans and animals. Between September 1999 and June 2004, 1,898 samples obtained from patients showing sporadic or outbreak gastroenteritis in Chiba Prefecture, Japan, were tested for NVs by reverse transcription-PCR. NVs were detected in 603 samples. Approximately 80% were positive for genogroup GII, 13% were positive for genogroup GI, and the remaining 7% were positive for both genogroups. Phylogenetic analysis showed that the GI and GII genogroups could be further divided into 13 and 16 genotypes (including new genotypes), respectively. The GII-4 genotype, which included five small genetic clusters (subtypes), was the most common in this study and was detected in approximately 40% of positive samples. The P2 regions of 10 strains belonging to each of the five GII-4 subtypes showed 5 to 18% amino acid diversity. The amino acid
substitutions accumulated in the protruding (P) region during the 5-year study period. Our data suggest that highly variable NV strains
are circulating in Chiba Prefecture, with a high rate of genetic change observed during the 5-year study period.
Performances of VITEK 2 Colorimetric Cards for Identification of Gram-Positive and Gram-Negative Bacteria
Authors: Wallet Frédéric , Loïez Caroline , Renaux Emilie , Lemaitre Nadine , Courcol René J. 
Published: September 2005

ABSTRACT
The purpose of this study was to evaluate the new VITEK 2 identification
cards that use colorimetric reading to identify gram-positive and
gram-negative bacteria (GP and GN cards, respectively) in comparison to
fluorimetric cards (ID-GPC and ID-GNB, respectively). A total of 580
clinical isolates and stock collection strains belonging to 116 taxa
were included in the study. Of the 249 gram-positive strains tested
with both the ID-GPC and GP cards, 218 (87.5%) and 235 (94.4%) strains
were correctly identified (to the genus and species level),
respectively. Of the 331 gram-negative strains tested with the ID-GNB
and GN cards, 295 (89.1%) and 321 (97%) strains were correctly
identified, respectively. Another focus of the study was to apply the
percentages of correct identifications obtained in this study to the
list of bacteria isolated in our laboratory (32,739 isolates) in the
year 2004. We obtained 97.9% correct identifications with the
colorimetric cards and 93.9% with fluorescent
cards.