bluuebunny/crossref_metadata_2025_split huggingface dataset:
High-Affinity Vanadate Transport System in the CyanobacteriumAnabaena variabilisATCC 29413
Authors: Pratte Brenda S. , Thiel Teresa 
Published: December 2005

ABSTRACTHigh-affinity vanadate transport systems have not heretofore been identified in any organism.Anabaena variabilis, which can fix nitrogen by using an alternative V-dependent nitrogenase, transported vanadate well. The concentration of vanadate giving half-maximum V-nitrogenase activity when added to V-starved cells was about 3 × 10−9M. The genes for an ABC-type vanadate transport system,vupABC, were found inA. variabilisabout 5 kb from the major cluster of genes encoding the V-nitrogenase, and like those genes, thevupABCgenes were repressed by molybdate; however, unlike the V-nitrogenase genes the vanadate transport genes were expressed in vegetative cells. AvupBmutant failed to grow by using V-nitrogenase unless high levels of vanadate were provided, suggesting that there was also a low-affinity vanadate transport system that functioned in thevupBmutant. ThevupABCgenes belong to a family of putative metal transport genes that include only one other characterized transport system, the tungstate transport genes ofEubacterium acidaminophilum. Similar genes are not present in the complete genomes of other bacterial strains that have a V-nitrogenase, includingAzotobacter vinelandii,Rhodopseudomonas palustris, andMethanosarcina barkeri.
Biochemical and Structural Characterization of an Essential Acyl Coenzyme A Carboxylase from
Mycobacterium tuberculosis
Authors: Gago Gabriela , Kurth Daniel , Diacovich Lautaro , Tsai Shiou-Chuan , Gramajo Hugo 
Published: December 2005

ABSTRACT

Pathogenic mycobacteria contain a variety of unique fatty acids that have methyl branches at an even-numbered position at the carboxyl end and a long
n
-aliphatic chain. One such group of acids, called mycocerosic acids, is found uniquely in the cell wall of pathogenic mycobacteria, and their biosynthesis is essential for growth and pathogenesis. Therefore, the biosynthetic pathway of the unique precursor of such lipids, methylmalonyl coenzyme A (CoA), represents an attractive target for developing new antituberculous drugs. Heterologous protein expression and purification of the individual subunits allowed the successful reconstitution of an essential acyl-CoA carboxylase from
Mycobacterium tuberculosis
, whose main role appears to be the synthesis of methylmalonyl-CoA. The enzyme complex was reconstituted from the α biotinylated subunit AccA3, the carboxyltransferase β subunit AccD5, and the ε subunit AccE5 (Rv3281). The kinetic properties of this enzyme showed a clear substrate preference for propionyl-CoA compared with acetyl-CoA (specificity constant fivefold higher), indicating that the main physiological role of this enzyme complex is to generate methylmalonyl-CoA for the biosynthesis of branched-chain fatty acids. The α and β subunits are capable of forming a stable α6-β6 subcomplex but with very low specific activity. The addition of the ε subunit, which binds tightly to the α-β subcomplex, is essential for gaining maximal enzyme activity.
Rap Phosphatase of Virulence Plasmid pXO1 InhibitsBacillus anthracisSporulation
Authors: Bongiorni Cristina , Stoessel Ricarda , Shoemaker Dorinda , Perego Marta 
Published: December 2005

ABSTRACTThis study shows that theBacillus anthracispXO1 virulence plasmid carries a Rap-Phr system, BXA0205, which regulates sporulation initiation in this organism. The BXA0205Rap protein was shown to dephosphorylate the Spo0F response regulator intermediate of the phosphorelay signal transduction system that regulates the initiation of the developmental pathway in response to environmental, metabolic, and cell cycle signals. The activity of the Rap protein was shown to be inhibited by the carboxy-terminal pentapeptide generated through an export-import processing pathway from the associated BXA0205Phr protein. Deregulation of the Rap activity by either overexpression or lack of the Phr pentapeptide resulted in severe inhibition of sporulation. Five additional Rap-Phr encoding systems were identified on the chromosome ofB. anthracis, one of which, BA3790-3791, also affected sporulation initiation. The results suggest that the plasmid-borne Rap-Phr system may provide a selective advantage to the virulence ofB. anthracis.
Stringent and Relaxed Recognition of
oriT
by Related Systems for Plasmid Mobilization: Implications for Horizontal Gene Transfer
Authors: Jandle Sarah , Meyer Richard 
Published: December 2005

ABSTRACT

The plasmids R1162 and pSC101 have origins of conjugative transfer (
oriT
s) and corresponding relaxases that are closely related. The
oriT
s are made up of a highly conserved core, where DNA is cleaved by the relaxase prior to transfer, and an inverted repeat that differs in size and sequence. We show that in each case the seven base pairs adjacent to the core and within one arm of the inverted repeat are sufficient to determine specificity. Within this DNA there are three AT base pairs located 4 bp from the core. Mutations in the AT base pairs suggest that the relaxase makes essential contacts at these locations to the minor groove of the DNA. The remaining four bases are different for each
oriT
and are both necessary and sufficient for stringent recognition of
oriT
by the pSC101 mobilization proteins. In contrast, the R1162 mobilization proteins have a much more relaxed requirement for the base sequence of this specificity region. As a result, the R1162 mobilization proteins can initiate transfer from a variety of sites, including those derived from the chromosome. The R1162 mobilization proteins could therefore contribute to the horizontal gene transfer of DNA from diverse sources.
Mutational Analysis of the
Myxococcus xanthus
Ω4406 Promoter Region Reveals an Upstream Negative Regulatory Element That Mediates C-Signal Dependence
Authors: Viswanathan Kartik , Viswanathan Poorna , Kroos Lee 
Published: December 2005

ABSTRACT

C signaling plays a key role in coordinating cell movement and differentiation during the multicellular developmental process of
Myxococcus xanthus
. C signaling regulates expression of genes induced after about 6 h into development, when cells are forming mounds. One gene whose expression depends absolutely on C signaling was identified by insertion of a transposable element at site Ω4406 which generated a transcriptional fusion between
lacZ
and an upstream promoter. We have investigated regulation of the Ω4406 promoter. A 5′ deletion revealed a negative regulatory element located between bp −533 and −100 relative to the transcriptional start site. In the absence of this element, the promoter was still developmentally regulated but about fourfold more active. Also, the truncated promoter region retained normal dependence on two developmental regulators, FruA and DevS, but lost its dependence on the C-signaling protein CsgA. We infer that C signaling partially overcomes the negative effect of the upstream element on activity of the Ω4406 promoter. Deletion of downstream DNA between bp 50 and 140 caused a threefold loss in expression, suggesting that a positive regulatory element lies in this region. Additional positive and negative regulatory elements are present in the region from bp −69 to −49, based on the effects of multiple-base-pair mutations. Within this region, a 5-bp element and a C-box-like sequence resemble sequences found in other developmentally regulated
M. xanthus
promoter regions, but the effects of single-base-pair changes in these sequences suggest that each functions uniquely. We conclude that regulation of the Ω4406 promoter involves multiple positive and negative regulatory elements located upstream and downstream of the region typically bound by RNA polymerase.
Genes for Small, Noncoding RNAs under Sporulation Control in
Bacillus subtilis
Authors: Silvaggi Jessica M. , Perkins John B. , Losick Richard 
Published: December 2005

ABSTRACT

The process of sporulation in the bacterium
Bacillus subtilis
is known to involve the programmed activation of several hundred protein-coding genes. Here we report the discovery of previously unrecognized genes under sporulation control that specify small, non-protein-coding RNAs (sRNAs). Genes for sRNAs were identified by transcriptional profiling with a microarray bearing probes for intergenic regions in the genome and by use of a comparative genomics algorithm that predicts regions of conserved RNA secondary structure. The gene for one such sRNA, SurA, which is located in the region between
yndK
and
yndL
, was induced at the start of development under the indirect control of the master regulator for entry into sporulation, Spo0A. The gene for a second sRNA, SurC, located in the region between
dnaJ
and
dnaK
, was switched on at a late stage of sporulation by the RNA polymerase sigma factor σ
K
, which directs gene transcription in the mother cell compartment of the developing sporangium. Finally, a third intergenic region, that between
polC
and
ylxS
, which specified several sRNAs, including two transcripts produced under the control of the forespore-specific sigma factor σ
G
and a third transcript generated by σ
K
, was identified. Our results indicate that the full repertoire of sporulation-specific gene expression involves the activation of multiple genes for small, noncoding RNAs.
Polar Flagellum Biogenesis inAeromonas hydrophila
Authors: Canals Rocío , Ramirez Silvia , Vilches Silvia , Horsburgh Gavin , Shaw Jonathan G. , Tomás Juan M. , Merino Susana 
Published: December 2005

ABSTRACTMesophilicAeromonasspp. constitutively express a single polar flagellum that helps the bacteria move to more favorable environments and is an important virulence and colonization factor. Certain strains can also produce multiple lateral flagella in semisolid media or over surfaces. We have previously reported 16 genes (flgNtoflgL) that constitute region 1 of theAeromonas hydrophilaAH-3 polar flagellum biogenesis gene clusters. We identified 39 new polar flagellum genes distributed in four noncontiguous chromosome regions (regions 2 to 5). Region 2 contained six genes (flaAtomaf-1), including a modification accessory factor gene (maf-1) that has not been previously reported and is thought to be involved in glycosylation of polar flagellum filament. Region 3 contained 29 genes (fliEtoorf29), most of which are involved in flagellum basal body formation and chemotaxis. Region 4 contained a single gene involved in the motor stator formation (motX), and region 5 contained the three master regulatory genes for theA. hydrophilapolar flagella (flrAtoflrC). Mutations in theflaH,maf-1,fliM,flhA,fliA, andflrCgenes, as well as the double mutantflaA flaB, all caused loss of polar flagella and reduction in adherence and biofilm formation. A defined mutation in thepomBstator gene did not affect polar flagellum motility, in contrast to themotXmutant, which was unable to swim even though it expressed a polar flagellum. Mutations in all of these genes did not affect lateral flagellum synthesis or swarming motility, showing that bothA. hydrophilaflagellum systems are entirely distinct.
Identification ofListeria monocytogenesGenes Contributing to Intracellular Replication by Expression Profiling and Mutant Screening
Authors: Joseph Biju , Przybilla Karin , Stühler Claudia , Schauer Kristina , Slaghuis Jörg , Fuchs Thilo M. , Goebel Werner 
Published: December 2005

ABSTRACTA successful transition ofListeria monocytogenesfrom the extracellular to the intracellular environment requires a precise adaptation response to conditions encountered in the host milieu. Although many key steps in the intracellular lifestyle of this gram-positive pathogen are well characterized, our knowledge about the factors required for cytosolic proliferation is still rather limited. We used DNA microarray and real-time reverse transcriptase PCR analyses to investigate the transcriptional profile of intracellularL. monocytogenesfollowing epithelial cell infection. Approximately 19% of the genes were differentially expressed by at least 1.6-fold relative to their level of transcription when grown in brain heart infusion medium, including genes encoding transporter proteins essential for the uptake of carbon and nitrogen sources, factors involved in anabolic pathways, stress proteins, transcriptional regulators, and proteins of unknown function. To validate the biological relevance of the intracellular gene expression profile, a random mutant library ofL. monocytogeneswas constructed by insertion-duplication mutagenesis and screened for intracellular-growth-deficient strains. By interfacing the results of both approaches, we provide evidence thatL. monocytogenescan use alternative carbon sources like phosphorylated glucose and glycerol and nitrogen sources like ethanolamine during replication in epithelial cells and that the pentose phosphate cycle, but not glycolysis, is the predominant pathway of sugar metabolism in the host environment. Additionally, we show that the synthesis of arginine, isoleucine, leucine, and valine, as well as a species-specific phosphoenolpyruvate-dependent phosphotransferase system, play a major role in the intracellular growth ofL. monocytogenes.
Novel Mechanism of
Escherichia coli
Porin Regulation
Authors: Castillo-Keller Maria , Vuong Phu , Misra Rajeev 
Published: December 2005

ABSTRACT

A novel mechanism of
Escherichia coli
porin regulation was discovered from multicopy suppressors that permitted growth of cells expressing a mutant OmpC protein in the absence of DegP. Analyses of two suppressors showed that both substantially lowered OmpC expression. Suppression activities were confined to a short DNA sequence, which we designated
ipeX
for
i
nhibition of
p
orin
ex
pression, and to DNA containing a 3′-truncated
ompR
gene. The major effect of
ipeX
on
ompC
expression was exerted posttranscriptionally, whereas the truncated OmpR protein reduced
ompC
transcription.
ipeX
was localized within an untranslated region of 247 base pairs between the stop codon of
nmpC
—a remnant porin gene from the cryptic phage
qsr
′ (DLP12) genome—and its predicted Rho-independent transcriptional terminator. Interestingly, another prophage, PA-2, which encodes a porin similar to NmpC, known as Lc, has sequences downstream from
lc
identical to that of
ipeX
. PA-2 lysogenization leads to Lc expression and OmpC inhibition. Our data show that the synthesis of the
lc
transcript, whose 3′ end contains the corresponding
ipeX
sequence, inhibits OmpC expression. Overexpression of
ipeX
RNA inhibited both OmpC and OmpF expression but not that of OmpA.
ompC
-
phoA
chimeric gene constructs revealed a 248-bp untranslated region of
ompC
required for
ipeX
-mediated inhibition. However, no sequence complementarity was found between
ipeX
and this region of
ompC
, indicating that inhibition may not involve simple base pairing between the two RNA molecules. The effect of
ipeX
on
ompC
, but not on
ompF
, was independent of the RNA chaperone Hfq.
YdgG (TqsA) Controls Biofilm Formation inEscherichia coliK-12 through Autoinducer 2 Transport
Authors: Herzberg Moshe , Kaye Ian K. , Peti Wolfgang , Wood Thomas K. 
Published: December 2005

ABSTRACTYdgG is an uncharacterized protein that is induced inEscherichia colibiofilms. Here it is shown that deletion ofydgGdecreased extracellular and increased intracellular concentrations of autoinducer 2 (AI-2); hence, YdgG enhances transport of AI-2. Consistent with this hypothesis, deletion ofydgGresulted in a 7,000-fold increase in biofilm thickness and 574-fold increase in biomass in flow cells. Also consistent with the hypothesis, deletion ofydgGincreased cell motility by increasing transcription of flagellar genes (genes induced by AI-2). By expressingydgGintrans, the wild-type phenotypes for extracellular AI-2 activity, motility, and biofilm formation were restored. YdgG is also predicted to be a membrane-spanning protein that is conserved in many bacteria, and it influences resistance to several antimicrobials, including crystal violet and streptomycin (this phenotype could also be complemented). Deletion ofydgGalso caused 31% of the bacterial chromosome to be differentially expressed in biofilms, as expected, since AI-2 controls hundreds of genes. YdgG was found to negatively modulate expression of flagellum- and motility-related genes, as well as other known products essential for biofilm formation, including operons for type 1 fimbriae, autotransporter protein Ag43, curli production, colanic acid production, and production of polysaccharide adhesin. Eighty genes not previously related to biofilm formation were also identified, including those that encode transport proteins (yihNandyihP), polysialic acid production (gutMandgutQ), CP4-57 prophage functions (yfjRandalpA), methionine biosynthesis (metR), biotin and thiamine biosynthesis (bioFandthiDFH), anaerobic metabolism (focB,hyfACDR,ttdA, andfumB), and proteins with unknown function (ybfG,yceO,yjhQ, andyjbE); 10 of these genes were verified through mutation to decrease biofilm formation by 40% or more (yfjR,bioF,yccW,yjbE,yceO,ttdA,fumB,yjiP,gutQ, andyihR). Hence, it appears YdgG controls the transport of the quorum-sensing signal AI-2, and so we suggest the gene nametqsA.
Dissection of the Bacteriophage Mu Strong Gyrase Site (SGS): Significance of the SGS Right Arm in Mu Biology and DNA Gyrase Mechanism
Authors: Oram Mark , Travers Andrew A. , Howells Alison J. , Maxwell Anthony , Pato Martin L. 
Published: December 2005

ABSTRACT
The bacteriophage Mu strong gyrase site (SGS), required for efficient phage DNA replication, differs from other gyrase sites in the efficiency of gyrase binding coupled with a highly processive supercoiling activity. Genetic studies have implicated the right arm of the SGS as a key structural feature for promoting rapid Mu replication. Here, we show that deletion of the distal portion of the right arm abolishes efficient binding, cleavage, and supercoiling by DNA gyrase in vitro. DNase I footprinting analysis of the intact SGS revealed an adenylyl imidodiphosphate-dependent change in protection in the right arm, indicating that this arm likely forms the T segment that is passed through the cleaved G segment during the supercoiling reaction. Furthermore, in an SGS derivative with an altered right-arm sequence, the left arm showed these changes, suggesting that the selection of a T segment by gyrase is determined primarily by the sequences of the arms. Analysis of the sequences of the SGS and other gyrase sites suggests that the choice of T segment correlates with which arm possesses the more extensive set of phased anisotropic bending signals, with the Mu right arm possessing an unusually extended set of such signals. The implications of these observations for the structure of the gyrase-DNA complex and for the biological function of the Mu SGS are discussed.
Roles of Lsr2 in Colony Morphology and Biofilm Formation ofMycobacterium smegmatis
Authors: Chen Jeffrey M. , German Greg J. , Alexander David C. , Ren Huiping , Tan Tracy , Liu Jun 
Published: December 2005

ABSTRACTThe lipid-rich cell wall is a defining feature ofMycobacteriumspecies. Individual cell wall components affect diverse mycobacterial phenotypes including colony morphology, biofilm formation, antibiotic resistance, and virulence. In this study, we describe a transposon insertion mutant ofMycobacterium smegmatismc2155 that exhibits altered colony morphology and defects in biofilm formation. The mutation was localized to thelsr2gene. First identified as an immunodominant T-cell antigen ofMycobacterium leprae,lsr2orthologs have been identified in all sequenced mycobacterial genomes, and homologs are found in many actinomycetes. Although its precise function remains unknown, localization experiments indicate that Lsr2 is a cytosolic protein, and cross-linking experiments demonstrate that it exists as a dimer. Characterization of cell wall lipid components reveals that theM. smegmatis lsr2mutant lacks two previously unidentified apolar lipids. Characterization by mass spectrometry and thin-layer chromatography indicate that these two apolar lipids are novel mycolate-containing compounds, called mycolyl-diacylglycerols (MDAGs), in which a mycolic acid (α- or α′-mycolate) molecule is esterified to a glycerol. Upon complementation with an intactlsr2gene, the mutant reverts to the parental phenotypes and MDAG production is restored. This study demonstrates that due to its impact on the biosynthesis of the hydrophobic MDAGs, Lsr2 plays an important role in the colony morphology and biofilm formation ofM. smegmatis.
Anaerobic Survival ofPseudomonas aeruginosaby Pyruvate Fermentation Requires an Usp-Type Stress Protein
Authors: Schreiber Kerstin , Boes Nelli , Eschbach Martin , Jaensch Lothar , Wehland Juergen , Bjarnsholt Thomas , Givskov Michael , Hentzer Morten , Schobert Max 
Published: December 2005

ABSTRACTRecently, we identified a pyruvate fermentation pathway inPseudomonas aeruginosasustaining anaerobic survival in the absence of alternative anaerobic respiratory and fermentative energy generation systems (M. Eschbach, K. Schreiber, K. Trunk, J. Buer, D. Jahn, and M. Schobert, J. Bacteriol.186:4596-4604, 2004). Anaerobic long-term survival ofP. aeruginosamight be essential for survival in deeper layers of a biofilm and the persistent infection of anaerobic mucus plaques in the cystic fibrosis lung. Proteome analysis ofP. aeruginosacells during a 7-day period of pyruvate fermentation revealed the induced synthesis of three enzymes involved in arginine fermentation, ArcA, ArcB, and ArcC, and the outer membrane protein OprL. Moreover, formation of two proteins of unknown function, PA3309 and PA4352, increased by factors of 72- and 22-fold, respectively. Both belong to the group of universal stress proteins (Usp). Long-term survival of a PA3309 knockout mutant by pyruvate fermentation was found drastically reduced. The oxygen-sensing regulator Anr controls expression of thePPA3309-lacZreporter gene fusion after a shift to anaerobic conditions and further pyruvate fermentation. PA3309 expression was also found induced during the anaerobic and aerobic stationary phases. This aerobic stationary-phase induction is independent of the regulatory proteins Anr, RpoS, RelA, GacA, RhlR, and LasR, indicating a currently unknown mechanism of stationary-phase-dependent gene activation. PA3309 promoter activity was detected in the deeper layers of aP. aeruginosabiofilm using aPPA3309-gfp(green fluorescent protein gene) fusion and confocal laser-scanning microscopy. This is the first description of an Anr-dependent, anaerobically induced, and functional Usp-like protein in bacteria.