bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evolution of the Mycobacterial SigK Regulon
Authors: Veyrier Frédéric , Saïd-Salim Battouli , Behr Marcel A. 
Published: January 2008

ABSTRACT

Previous studies have established that members of the
Mycobacterium tuberculosis
complex exhibit variable production of the antigenic proteins MPT70 and MPT83 due to mutations in their positive regulator, SigK (sigma factor K), and their negative regulator, RskA (regulator of sigma K). To further understand this highly specific SigK-controlled regulon, we have undertaken evolutionary studies to determine the presence of homologues of SigK-regulated genes in other organisms and to predict its transcriptional network. Evolutionary analysis indicates that the positive and negative regulators are conserved across many organisms, but that the genes under their control are variable. Moreover, the addition, loss, and movement of various genes in the
mpt70/83
locus suggest that these genes are unlikely to be cotranscribed. To test predictions from sequence analysis, we have used promoter luciferase fusions and Northern blots to show that the majority of genes in this locus have their own promoters, of which a subset are SigK regulated (
mpt83
,
dipZ
,
mpt70
, and Rv0449c). Next, we have shown that the intracellular inducibility of
mpt70
and
mpt83
is a conserved property, shared between
M. tuberculosis
and
Mycobacterium marinum
. In addition, we have shown that SigK and RskA from an environmental mycobacterium isolate (
M. gilvum
PYR-GCK) complemented the regulatory activity of
M. tuberculosis
Δ
sigK rskA
. Together, our data indicate that the regulatory system SigK/RskA is conserved across the
Mycobacterium
genus, whereas the regulon under its control varies considerably across species.
Three-Dimensional Imaging of the Highly Bent Architecture of
Bdellovibrio bacteriovorus
by Using Cryo-Electron Tomography
Authors: Borgnia Mario J. , Subramaniam Sriram , Milne Jacqueline L. S. 
Published: January 2008

ABSTRACT

Bdellovibrio bacteriovorus
cells are small deltaproteobacterial cells that feed on other gram-negative bacteria, including human pathogens. Using cryo-electron tomography, we demonstrated that
B. bacteriovorus
cells are capable of substantial flexibility and local deformation of the outer and inner membranes without loss of cell integrity. These shape changes can occur in less than 2 min, and analysis of the internal architecture of highly bent cells showed that the overall distribution of molecular machines and the nucleoid is similar to that in moderately bent cells.
B. bacteriovorus
cells appear to contain an extensive internal network of short and long filamentous structures. We propose that rearrangements of these structures, in combination with the unique properties of the cell envelope, may underlie the remarkable ability of
B. bacteriovorus
cells to find and enter bacterial prey.
Role of the Nfo and ExoA Apurinic/Apyrimidinic Endonucleases in Repair of DNA Damage during Outgrowth of
Bacillus subtilis
Spores
Authors: Ibarra Juan R. , Orozco Alma D. , Rojas Juan A. , López Karina , Setlow Peter , Yasbin Ronald E. , Pedraza-Reyes Mario 
Published: January 2008

ABSTRACT

Germination and outgrowth are critical steps for returning
Bacillus subtilis
spores to life. However, oxidative stress due to full hydration of the spore core during germination and activation of metabolism in spore outgrowth may generate oxidative DNA damage that in many species is processed by apurinic/apyrimidinic (AP) endonucleases.
B. subtilis
spores possess two AP endonucleases, Nfo and ExoA; the outgrowth of spores lacking both of these enzymes was slowed, and the spores had an elevated mutation frequency, suggesting that these enzymes repair DNA lesions induced by oxidative stress during spore germination and outgrowth. Addition of H
2
O
2
also slowed the outgrowth of
nfo exoA
spores and increased the mutation frequency, and
nfo
and
exoA
mutations slowed the outgrowth of spores deficient in either RecA, nucleotide excision repair (NER), or the DNA-protective α/β-type small acid-soluble spore proteins (SASP). These results suggest that α/β-type SASP protect DNA of germinating spores against damage that can be repaired by Nfo and ExoA, which is generated either spontaneously or promoted by addition of H
2
O
2
. The contribution of RecA and Nfo/ExoA was similar to but greater than that of NER in repair of DNA damage generated during spore germination and outgrowth. However,
nfo
and
exoA
mutations increased the spontaneous mutation frequencies of outgrown spores lacking
uvrA
or
recA
to about the same extent, suggesting that DNA lesions generated during spore germination and outgrowth are processed by Nfo/ExoA in combination with NER and/or RecA. These results suggest that Nfo/ExoA, RecA, the NER system, and α/β-type SASP all contribute to the repair of and/or protection against oxidative damage of DNA in germinating and outgrowing spores.
Cel9D, an Atypical 1,4-β-
d
-Glucan Glucohydrolase from
Fibrobacter succinogenes
: Characteristics, Catalytic Residues, and Synergistic Interactions with Other Cellulases
Authors: Qi Meng , Jun Hyun-Sik , Forsberg Cecil W. 
Published: January 2008

ABSTRACT

The increasing demands of renewable energy have led to the critical emphasis on novel enzymes to enhance cellulose biodegradation for biomass conversion. To identify new cellulases in the ruminal bacterium
Fibrobacter succinogenes
, a cell extract of cellulose-grown cells was separated by ion-exchange chromatography and cellulases were located by zymogram analysis and identified by peptide mass fingerprinting. An atypical family 9 glycoside hydrolase (GH9), Cel9D, with less than 20% identity to typical GH9 cellulases, was identified. Purified recombinant Cel9D enhanced the production of reducing sugar from acid swollen cellulose (ASC) and Avicel by 1.5- to 4-fold when mixed separately with each of four other glucanases, although it had low activity on these substrates. Cel9D degraded ASC and cellodextrins with a degree of polymerization higher than 2 to glucose with no apparent endoglucanase activity, and its activity was restricted to β-1→4-linked glucose residues. It catalyzed the hydrolysis of cellulose by an inverting mode of reaction, releasing glucose from the nonreducing end. Unlike many GH9 cellulases, calcium ions were not required for its function. Cel9D had increased
k
cat
/K

m

values for cello-oligosaccharides with higher degrees of polymerization. The
k
cat
/K

m

value for cellohexaose was 2,300 times higher than that on cellobiose. This result indicates that Cel9D is a 1,4-β-
d
-glucan glucohydrolase (EC 3.2.1.74) in the GH9 family. Site-directed mutagenesis of Cel9D identified Asp166 and Glu612 as the candidate catalytic residues, while Ser168, which is not present in typical GH9 cellulases, has a crucial structural role. This enzyme has an important role in crystalline cellulose digestion by releasing glucose from accessible cello-oligosaccharides.
Variation in the Group BStreptococcusCsrRS Regulon and Effects on Pathogenicity
Authors: Jiang Sheng-Mei , Ishmael Nadeeza , Hotopp Julie Dunning , Puliti Manuela , Tissi Luciana , Kumar Nikhil , Cieslewicz Michael J. , Tettelin Hervé , Wessels Michael R. 
Published: January 2008

ABSTRACTCsrRS (or CovRS) is a two-component regulatory system that controls expression of multiple virulence factors in the important human pathogen group BStreptococcus(GBS). We now report global gene expression studies in GBS strains 2603V/R and 515 and their isogeniccsrRandcsrSmutants. Together with data reported previously for strain NEM316, the results reveal a conserved 39-gene CsrRS regulon. In vitro phosphorylation-dependent binding of recombinant CsrR to promoter regions of both positively and negatively regulated genes suggests that direct binding of CsrR can mediate activation as well as repression of target gene expression. Distinct patterns of gene regulation incsrRversuscsrSmutants in strain 2603V/R compared to 515 were associated with different hierarchies of relative virulence of wild-type,csrR, andcsrSmutants in murine models of systemic infection and septic arthritis. We conclude that CsrRS regulates a core group of genes including important virulence factors in diverse strains of GBS but also displays marked variability in the repertoire of regulated genes and in the relative effects of CsrS signaling on CsrR-mediated gene regulation. Such variation is likely to play an important role in strain-specific adaptation of GBS to particular host environments and pathogenic potential in susceptible hosts.
Induction and Loss of Ti Plasmid Conjugative Competence in Response to the Acyl-Homoserine Lactone Quorum-Sensing Signal
Authors: Su Shengchang , Khan Sharik R. , Farrand Stephen K. 
Published: January 2008

ABSTRACTConjugative transfer of the Ti plasmids ofAgrobacterium tumefaciensis controlled by a quorum-sensing system composed of TraR and its signalN-(3-oxo-octanoyl)-l-homoserine lactone. This system is, in turn, controlled by the conjugative opines produced by crown gall tumors induced on plants by the bacteria. Using nonpolartraImutants, we examined the kinetics of induction of conjugative transfer in response to exogenous acyl-homoserine lactone. In the absence of the antiactivator TraM, onset of induction of transfer requires about 30 min, 15 to 20 min of which is needed for expression and construction of the conjugative apparatus. TraM delays the onset of conjugation by 30 min. While the rate of development of conjugative competence was not significantly affected by levels of TraR, maximum efficiencies of transfer were correlated with amounts of the activator in the donors. Donors harboring Ti plasmids lacking TraM were fully induced by the quormone at concentrations as low as 100 pM. TraM raised the concentration of signal required for maximum activity to 1 nM. Donors grown in batch culture retained conjugative competence following signal removal, even when in stationary phase. However, donors kept in balanced growth rapidly lost transfer ability following signal removal. Loss of transfer was mirrored by a decrease in levels of active TraR. Decreases in TraR activity and conjugative competence could be accounted for by dilution associated with cell division, suggesting that while induction of Ti plasmid conjugation is an active process, the cells lack a mechanism for disassembling the conjugative apparatus when signals become limiting.
The UP Element Is Necessary but Not Sufficient for Growth Rate-Dependent Control of the
Escherichia coli guaB
Promoter
Authors: Husnain Seyyed I. , Thomas Mark S. 
Published: January 2008

ABSTRACT

The
Escherichia coli guaB
promoter (P

guaB

) regulates the transcription of two genes,
guaB
and
guaA
, that are required for de novo synthesis of GMP, a precursor for the synthesis of guanine nucleoside triphosphates. The activity of P

guaB

is subject to growth rate-dependent control (GRDC). Here we show that the A+T-rich sequence located between positions −59 and −38 relative to the
guaB
transcription start site stimulates transcription from P

guaB

∼8- to 10-fold and, in common with other UP elements, requires the C-terminal domain of the RNA polymerase α subunit for activity. Like the
rrnB
P1 UP element, the P

guaB

UP element contains two independently acting subsites located at positions −59 to −47 and −46 to −38 and can stimulate transcription when placed upstream of the
lacP1
promoter. We reveal a novel role for the P

guaB

UP element by demonstrating that it is required for GRDC. The involvement of the UP element in GRDC also requires the participation of sequences located at least 100 bp upstream of the
guaB
transcription start site. These sequences are required for down-regulation of P

guaB

activity at lower growth rates.
Widespread Distribution in Pathogenic Bacteria of Di-Iron Proteins That Repair Oxidative and Nitrosative Damage to Iron-Sulfur Centers
Authors: Overton Tim W. , Justino Marta C. , Li Ying , Baptista Joana M. , Melo Ana M. P. , Cole Jeffrey A. , Saraiva Lígia M. 
Published: January 2008

ABSTRACT

Expression of two genes of unknown function,
Staphylococcus aureus scdA
and
Neisseria gonorrhoeae dnrN
, is induced by exposure to oxidative or nitrosative stress. We show that DnrN and ScdA are di-iron proteins that protect their hosts from damage caused by exposure to nitric oxide and to hydrogen peroxide. Loss of FNR-dependent activation of
aniA
expression and NsrR-dependent repression of
norB
and
dnrN
expression on exposure to NO was restored in the gonococcal parent strain but not in a
dnrN
mutant, suggesting that DnrN is necessary for the repair of NO damage to the gonococcal transcription factors, FNR and NsrR. Restoration of aconitase activity destroyed by exposure of
S. aureus
to NO or H
2
O
2
required a functional
scdA
gene. Electron paramagnetic resonance spectra of recombinant ScdA purified from
Escherichia coli
confirmed the presence of a di-iron center. The recombinant
scdA
plasmid, but not recombinant plasmids encoding the complete
Escherichia coli sufABCDSE
or
iscRSUAhscBAfdx
operons, complemented repair defects of an
E. coli ytfE
mutant. Analysis of the protein sequence database revealed the importance of the two proteins based on the widespread distribution of highly conserved homologues in both gram-positive and gram-negative bacteria that are human pathogens. We provide in vivo and in vitro evidence that Fe-S clusters damaged by exposure to NO and H
2
O
2
can be repaired by this new protein family, for which we propose the name
r
epair of
i
ron
c
enters, or RIC, proteins.
Functional Analysis of Predicted Coiled-Coil Regions in the
Escherichia coli
K-12 O-Antigen Polysaccharide Chain Length Determinant Wzz
Authors: Marolda Cristina L. , Haggerty Emily R. , Lung Michael , Valvano Miguel A. 
Published: January 2008

ABSTRACT

Wzz is a membrane protein that determines the chain length distribution of the O-antigen lipopolysaccharide by an unknown mechanism. Wzz proteins consist of two transmembrane helices separated by a large periplasmic loop. The periplasmic loop of
Escherichia coli
K-12 Wzz (244 amino acids from K65 to A308) was purified and found to be a monomer with an extended conformation, as determined by gel filtration chromatography and analytical ultracentrifugation. Circular dichroism showed that the loop has a 60% helical content. The Wzz periplasmic loop also contains three regions with predicted coiled coils. To probe the function of the predicted coiled coils, we constructed amino acid replacement mutants of the
E. coli
K-12 Wzz protein, which were designed so that the coiled coils could be separate without compromising the helicity of the individual molecules. Mutations in one of the regions, spanning amino acids 108 to 130 (region I), were associated with a partial defect in O-antigen chain length distribution, while mutants with mutations in the region spanning amino acids 209 to 223 (region III) did not have an apparent functional defect. In contrast, mutations in the region spanning amino acids 153 to 173 (region II) eliminated the Wzz function. This phenotype was associated with protein instability, most likely due to conformational changes caused by the amino acid replacements, which was confirmed by limited trypsin proteolysis. Additional mutagenesis based on a three-dimensional model of region I demonstrated that the amino acids implicated in function are all located at the same face of a predicted α-helix, suggesting that a coiled coil actually does not exist in this region. Together, our results suggest that the regions predicted to be coiled coils are important for Wzz function because they maintain the native conformation of the protein, although the existence of coiled coils could not be demonstrated experimentally.
Utilization of Lactoferrin-Bound and Transferrin-Bound Iron by
Campylobacter jejuni
Authors: Miller Claire E. , Rock Jonathan D. , Ridley Kristian A. , Williams Peter H. , Ketley Julian M. 
Published: January 2008

ABSTRACT

Campylobacter jejuni
NCTC 11168 was capable of growth to levels comparable with FeSO
4
in defined iron-limited medium (minimal essential medium alpha [MEMα]) containing ferrilactoferrin, ferritransferrin, or ferri-ovotransferrin. Iron was internalized in a contact-dependent manner, with 94% of cell-associated radioactivity from either
55
Fe-loaded transferrin or lactoferrin associated with the soluble cell fraction. Partitioning the iron source away from bacteria significantly decreased cellular growth. Excess cold transferrin or lactoferrin in cultures containing
55
Fe-loaded transferrin or lactoferrin resulted in reduced levels of
55
Fe uptake. Growth of
C. jejuni
in the presence of ferri- and an excess of apoprotein reduced overall levels of growth. Following incubation of cells in the presence of ferrilactoferrin, lactoferrin became associated with the cell surface; binding levels were higher after growth under iron limitation. A strain carrying a mutation in the cj0178 gene from the iron uptake system Cj0173c-Cj0178 demonstrated significantly reduced growth promotion in the presence of ferrilactoferrin in MEMα compared to wild type but was not affected in the presence of heme. Moreover, this mutant acquired less
55
Fe than wild type when incubated with
55
Fe-loaded protein and bound less lactoferrin. Complementation restored the wild-type phenotype when cells were grown with ferrilactoferrin. A mutant in the ABC transporter system permease gene (cj0174c) showed a small but significant growth reduction. The cj0176c-cj0177 intergenic region contains two separate Fur-regulated iron-repressible promoters. This is the first demonstration that
C. jejuni
is capable of acquiring iron from members of the transferrin protein family, and our data indicate a role for Cj0178 in this process.
Nondecarboxylating and Decarboxylating Isocitrate Dehydrogenases: Oxalosuccinate Reductase as an Ancestral Form of Isocitrate Dehydrogenase
Authors: Aoshima Miho , Igarashi Yasuo 
Published: January 2008

ABSTRACTIsocitrate dehydrogenase (ICDH) fromHydrogenobacter thermophiluscatalyzes the reduction of oxalosuccinate, which corresponds to the second step of the reductive carboxylation of 2-oxoglutarate in the reductive tricarboxylic acid cycle. In this study, the oxidation reaction catalyzed byH. thermophilusICDH was kinetically analyzed. As a result, a rapid equilibrium random-order mechanism was suggested. The affinities of both substrates (isocitrate and NAD+) toward the enzyme were extremely low compared to other known ICDHs. The binding activities of isocitrate and NAD+were not independent; rather, the binding of one substrate considerably promoted the binding of the other. A product inhibition assay demonstrated that NADH is a potent inhibitor, although 2-oxoglutarate did not exhibit an inhibitory effect. Further chromatographic analysis demonstrated that oxalosuccinate, rather than 2-oxoglutarate, is the reaction product. Thus, it was shown thatH. thermophilusICDH is a nondecarboxylating ICDH that catalyzes the conversion between isocitrate and oxalosuccinate by oxidation and reduction. This nondecarboxylating ICDH is distinct from well-known decarboxylating ICDHs and should be categorized as a new enzyme. Oxalosuccinate-reducing enzyme may be the ancestral form of ICDH, which evolved to the extant isocitrate oxidative decarboxylating enzyme by acquiring higher substrate affinities.
Global Responses of
Methanococcus maripaludis
to Specific Nutrient Limitations and Growth Rate
Authors: Hendrickson Erik L. , Liu Yuchen , Rosas-Sandoval Guillermina , Porat Iris , Söll Dieter , Whitman William B. , Leigh John A. 
Published: January 2008

ABSTRACT

Continuous culture, transcriptome arrays, and measurements of cellular amino acid pools and tRNA charging levels were used to determine the response of
Methanococcus maripaludis
to leucine limitation. For comparison, the responses to phosphate and H
2
limitations were measured as well. In addition, the effect of growth rate was determined. Leucine limitation resulted in a broad response. tRNA
Leu
charging decreased, but only small increases in mRNA were seen for amino acid biosynthesis genes. However, the cellular levels of free isoleucine and valine showed significant increases, indicating a coordinate regulation of branched-chain amino acids at a post-mRNA level. Leucine limitation also resulted in increased mRNA abundance for ribosomal protein genes, increased rRNA abundance, and decreased mRNA abundance for genes of methanogenesis. In contrast, phosphate limitation induced a specific response, a marked increase in mRNA levels for a phosphate transporter. Some mRNA levels responded to more than one factor; for example, transcripts for flagellum synthesis genes decreased under conditions of leucine limitation and increased under H
2
limitation. Increased growth rate resulted in increased mRNA levels for ribosomal protein genes, increased rRNA abundance, and increased mRNA for a gene encoding an S-layer protein.
Collegiate rugby union injury patterns in New England: A prospective cohort study
Authors: Kerr Hamish A. , Curtis Christine M. , Micheli Lyle J. , Kocher Mininder S. , Zurakowski David , Brooks John H.M. , Kemp Simon P.T. 
Published: January 2008

Objective:Establish injury profile of collegiate rugby union in the U.S.A.Design/setting:31 men's and 38 women's collegiate rugby union teams prospectively recorded injuries during games and practice during 2005-06. 3 teams withdrew prior to data collection. An injury was defined as;1. Occurring in an organized intercollegiate game or practice; and2. Requiring medical attention during or after the game or practice, or3. Resulting in any restriction of the athletes' participation for ≥1 day(s) beyond the day of injury, or in a dental injury. Main Outcome measurements: A total of 847 injuries (men:447; women:400) during 48,026 practice (men:24,280; women:23,746) and 25,808 game (men:13,943; women:11,865) exposures were recorded.Results:During games, injury rates of 22.5(95 per cent confidence interval: 20.2-25.0) and 22.7(20.2-25.5)/1,000 game athletic exposures (GAE) or 16.9 (15.1-18.9) and 17.1(15.1-19.1)/1,000 player game-hours (PGH) were recorded (men; women). Over half of all match injuries were of major severity (> 7 days absence) (men: 56%; women: 51%) and the tackle was the game event most associated with injury (men: 48%; women: 53%).Conclusions:Collegiate game injury rates for rugby were lower than rates recorded previously in men's professional club and international rugby and lower than reported by the National Collegiate Athletic Association Injury Surveillance System (N.C.A.A. I.S.S.) for American football but similar to rates reported for men's and women's soccer in 2005-06.