bluuebunny/crossref_metadata_2025_split huggingface dataset:
Differences in Resolution of
mwr
-Containing Plasmid Dimers Mediated by the
Klebsiella pneumoniae
and
Escherichia coli
XerC Recombinases: Potential Implications in Dissemination of Antibiotic Resistance Genes
Authors: Bui Duyen , Ramiscal Judianne , Trigueros Sonia , Newmark Jason S. , Do Albert , Sherratt David J. , Tolmasky Marcelo E. 
Published: April 2006

ABSTRACT

Xer-mediated dimer resolution at the
mwr
site of the multiresistance plasmid pJHCMW1 is osmoregulated in
Escherichia coli
containing either the
Escherichia coli
Xer recombination machinery or Xer recombination elements from
K. pneumoniae
. In the presence of
K. pneumoniae
XerC (XerC
Kp
), the efficiency of recombination is lower than that in the presence of the
E. coli
XerC (XerC
Ec
) and the level of dimer resolution is insufficient to stabilize the plasmid, even at low osmolarity. This lower efficiency of recombination at
mwr
is observed in the presence of
E. coli
or
K. pneumoniae
XerD proteins. Mutagenesis experiments identified a region near the N terminus of XerC
Kp
responsible for the lower level of recombination catalyzed by XerC
Kp
at
mwr
. This region encompasses the second half of the predicted α-helix B and the beginning of the predicted α-helix C. The efficiencies of recombination at other sites such as
dif
or
cer
in the presence of XerC
Kp
or XerC
Ec
are comparable. Therefore, XerC
Kp
is an active recombinase whose action is impaired on the
mwr
recombination site. This characteristic may result in restriction of the host range of plasmids carrying this site, a phenomenon that may have important implications in the dissemination of antibiotic resistance genes.
Morphology of Isolated Gli349, a Leg Protein Responsible for
Mycoplasma mobile
Gliding via Glass Binding, Revealed by Rotary Shadowing Electron Microscopy
Authors: Adan-Kubo Jun , Uenoyama Atsuko , Arata Toshiaki , Miyata Makoto 
Published: April 2006

ABSTRACT

Several species of mycoplasmas rely on an unknown mechanism to glide across solid surfaces in the direction of a membrane protrusion at the cell pole. Our recent studies on the fastest species,
Mycoplasma mobile
, suggested that a 349-kDa protein, Gli349, localized at the base of the membrane protrusion called the neck, forms legs that stick out from the neck and propel the cell by repeatedly binding to and releasing from a solid surface, based on the energy of ATP hydrolysis. Here, the Gli349 protein was isolated from mycoplasma cells and its structure was analyzed. Gel filtration analysis showed that the isolated Gli349 protein is monomeric. Rotary shadowing electron microscopy revealed that the molecular structure resembles the symbol for an eighth note in music. It contains an oval foot 14 nm long in axis. From this foot extend three rods in tandem of 43, 20, and 20 nm, in that order. The hinge connecting the first and second rods is flexible, while the next hinge has a distinct preference in its angle, near 90 degrees. Molecular images revealed that a monoclonal antibody that can bind to the position at one-third of the total peptide length from the N terminus bound to a position two-thirds from the foot end, suggesting that the foot corresponds to the C-terminal region. The amino acid sequence was assigned to the molecular image, and the topology of the molecule in the gliding machinery is discussed.
A Novel FtsZ-Like Protein Is Involved in Replication of the Anthrax Toxin-Encoding pXO1 Plasmid inBacillus anthracis
Authors: Tinsley Eowyn , Khan Saleem A. 
Published: April 2006

ABSTRACTPlasmid pXO1 encodes the tripartite anthrax toxin, which is the major virulence factor ofBacillus anthracis. In spite of the important role of pXO1 in anthrax pathogenesis, very little is known about its replication and maintenance inB. anthracis. We cloned a 5-kb region of the pXO1 plasmid into anEscherichia colivector and showed that this plasmid can replicate when introduced intoB. anthracis. Mutational analysis showed that open reading frame 45 (repX) of pXO1 was required for the replication of the miniplasmid inB. anthracis. Interestingly,repXshowed limited homology to bacterial FtsZ proteins that are involved in cell division. A mutation in the predicted GTP binding domain of RepX abolished its replication activity. Genes almost identical torepXare contained on several megaplasmids in members of theBacillus cereusgroup, including aB. cereusstrain that causes an anthrax-like disease. Our results identify a novel group of FtsZ-related initiator proteins that are required for the replication of virulence plasmids inB. anthracisand possibly in related organisms. Such replication proteins may provide novel drug targets for the elimination of plasmids encoding the anthrax toxin and other virulence factors.
Identification of Lactaldehyde Dehydrogenase in
Methanocaldococcus jannaschii
and Its Involvement in Production of Lactate for F
420
Biosynthesis
Authors: Grochowski Laura L. , Xu Huimin , White Robert H. 
Published: April 2006

ABSTRACT

One of the early steps in the biosynthesis of coenzyme F
420
in
Methanocaldococcus jannaschii
requires generation of 2-phospho-
l
-lactate, which is formed by the phosphorylation of
l
-lactate. Preliminary studies had shown that
l
-lactate in
M. jannaschii
is not derived from pyruvate, and thus an alternate pathway(s) for its formation was examined. Here we report that
l
-lactate is formed by the NAD
+
-dependent oxidation of
l
-lactaldehyde by the MJ1411 gene product. The lactaldehyde, in turn, was found to be generated either by the NAD(P)H reduction of methylglyoxal or by the aldol cleavage of fuculose-1-phosphate by fuculose-1-phosphate aldolase, the MJ1418 gene product.
Transcriptional Heat Shock Response in the Smallest Known Self-Replicating Cell,Mycoplasma genitalium
Authors: Musatovova Oxana , Dhandayuthapani Subramanian , Baseman Joel B. 
Published: April 2006

ABSTRACTMycoplasma genitaliumis a human bacterial pathogen linked to urethritis and other sexually transmitted diseases as well as respiratory and joint pathologies. Though its complete genome sequence is available, little is understood about the regulation of gene expression in this smallest known, self-replicating cell, as its genome lacks orthologues for most of the conventional bacterial regulators. Still, the transcriptional repressor HrcA (heatregulation atCIRCE [controllinginvertedrepeat ofchaperoneexpression]) is predicted in theM. genitaliumgenome as well as three copies of its corresponding regulatory sequence CIRCE. We investigated the transcriptional response ofM. genitaliumto elevated temperatures and detected the differential induction of fourhspgenes. Three of the up-regulated genes, which encode DnaK, ClpB, and Lon, possess CIRCE within their promoter regions, suggesting that the HrcA-CIRCE regulatory mechanism is functional. Additionally, one of three DnaJ-encoding genes was up-regulated, even though no known regulatory sequences were found in the promoter region. Transcript levels returned to control values after 1 h of incubation at 37°C, reinforcing the transient nature of the heat shock transcriptional response. Interestingly, neither of thegroESLoperon genes, which encode the GroEL chaperone and its cochaperone GroES, responded to heat shock. These data suggest thatM. genitaliumselectively regulates a limited number of genes in response to heat shock.
Genetic Analysis of the Mode of Interplay between an ATPase Subunit and Membrane Subunits of the Lipoprotein-Releasing ATP-Binding Cassette Transporter LolCDE
Authors: Ito Yasuko , Matsuzawa Hitomi , Matsuyama Shin-ichi , Narita Shin-ichiro , Tokuda Hajime 
Published: April 2006

ABSTRACT

The LolCDE complex, an ATP-binding cassette (ABC) transporter, releases lipoproteins from the inner membrane, thereby initiating lipoprotein sorting to the outer membrane of
Escherichia coli
. The LolCDE complex is composed of two copies of an ATPase subunit, LolD, and one copy each of integral membrane subunits LolC and LolE. LolD hydrolyzes ATP on the cytoplasmic side of the inner membrane, while LolC and/or LolE recognize and release lipoproteins anchored to the periplasmic leaflet of the inner membrane. Thus, functional interaction between LolD and LolC/E is critically important for coupling of ATP hydrolysis to the lipoprotein release reaction. LolD contains a characteristic sequence called the LolD motif, which is highly conserved among LolD homologs but not other ABC transporters of
E. coli
. The LolD motif is suggested to be a region in contact with LolC/E, judging from the crystal structures of other ABC transporters. To determine the functions of the LolD motif, we mutagenized each of the 32 residues of the LolD motif and isolated 26 dominant-negative mutants, whose overexpression arrested growth despite the chromosomal
lolD
+
background. We then selected suppressor mutations of the
lolC
and
lolE
genes that correct the growth defect caused by the LolD mutations. Mutations of the
lolC
suppressors were mainly located in the periplasmic loop, whereas ones of
lolE
suppressors were mainly located in the cytoplasmic loop, suggesting that the mode of interaction with LolD differs between LolC and LolE. Moreover, the LolD motif was found to be critical for functional interplay with LolC/E, since some LolD mutations lowered the ATPase activity of LolCDE without affecting that of LolD.
Comparison of OG1RF and an IsogenicfsrBDeletion Mutant by Transcriptional Analysis: the Fsr System ofEnterococcus faecalisIs More than the Activator of Gelatinase and Serine Protease
Authors: Bourgogne Agathe , Hilsenbeck Susan G. , Dunny Gary M. , Murray Barbara E. 
Published: April 2006

ABSTRACTThe FsrABC system ofEnterococcus faecaliscontrols the expression of gelatinase and a serine protease via a quorum-sensing mechanism, and recent studies suggest that the Fsr system may also regulate other genes important for virulence. To investigate the possibility that Fsr influences the expression of additional genes, we used transcriptional profiling, with microarrays based on theE. faecalisstrain V583 sequence, to compare theE. faecalisstrain OG1RF with its isogenic mutant, TX5266, anfsrBdeletion mutant. We found that the presence of an intactfsrBinfluences expression of numerous genes throughout the growth phases tested, namely, late log to early stationary phase. In addition, the Fsr regulon is independent of the activity of the proteases, GelE and SprE, whose expression was confirmed to be activated at all three time points tested. While expression of some genes (i.e., ef1097 and ef0750 to -757, encoding hypothetical proteins) was activated in late log phase in OG1RF versus thefsrBdeletion mutant, expression of ef1617 to -1634 (eut-pduorthologues) was highly repressed by the presence of an intact Fsr at entry into stationary phase. This is the first time that Fsr has been characterized as a negative regulator. The newly recognized Fsr-regulated targets include other factors, besides gelatinase, described as important for biofilms (BopD), and genes predicted to encode the surface proteins EF0750 to -0757 and EF1097, along with proteins implicated in several metabolic pathways, indicating that the FsrABC system may be an important regulator in strain OG1RF, with both positive and negative effects.
Functional Analysis ofluxSinStaphylococcus aureusReveals a Role in Metabolism but Not Quorum Sensing
Authors: Doherty Neil , Holden Matthew T. G. , Qazi Saara N. , Williams Paul , Winzer Klaus 
Published: April 2006

ABSTRACTThe function of AI-2 in many bacteria and the physiological role of LuxS, the enzyme responsible for its production, remain matters of debate. Here, we show that inStaphylococcus aureustheluxSgene forms a monocistronic transcriptional unit under the control of a σ70-dependent promoter. The gene was transcribed throughout growth under a variety of conditions, including intracellular growth in MAC-T cells. AI-2 was produced in rich media under aerobic and anaerobic conditions, peaking during the transition to stationary phase, but was hardly detectable in a sulfur-limited defined medium. In the presence of glucose or under anaerobic conditions, cultures retained considerable AI-2 activity after entry into stationary phase. Inactivation ofluxSin variousS. aureusstrains did not affect virulence-associated traits, such as production of hemolysins and extracellular proteases, biofilm formation, and theagrsignaling system. Conversely, AI-2 production remained unchanged in anagrmutant. However,luxSmutants grown in a sulfur-limited defined medium exhibited a growth defect. When grown together with the wild type in mixed culture,luxSmutants of variousS. aureusstrains showed reduced ability to compete for growth under these conditions. In contrast, a complementedluxSmutant grew as well as the parent strain, suggesting that the observed growth defect was of an intracellular nature and had not been caused by either second-site mutations or the lack of a diffusible factor. However, the LuxS/AI-2 system does not appear to contribute to the overall fitness ofS. aureusRN6390B during intracellular growth in epithelial cells: the wild type and aluxSmutant showed very similar growth patterns after their internalization by MAC-T cells.
TheppuI-rsaL-ppuRQuorum-Sensing System Regulates Biofilm Formation ofPseudomonas putidaPCL1445 by Controlling Biosynthesis of the Cyclic Lipopeptides Putisolvins I and II
Authors: Dubern Jean-Frédéric , Lugtenberg Ben J. J. , Bloemberg Guido V. 
Published: April 2006

ABSTRACTPseudomonas putidastrain PCL1445 produces two cyclic lipopeptides, putisolvin I and putisolvin II, which possess surface tension-reducing abilities and are able to inhibit biofilm formation and to break down existing biofilms of severalPseudomonasspp., includingP. aeruginosa. Putisolvins are secreted in the culture medium during growth at late exponential phase, indicating that production is possibly regulated by quorum sensing. In the present study, we identified a quorum-sensing system in PCL1445 that is composed ofppuI,rsaL, andppuRand shows very high similarity with gene clusters ofP. putidastrains IsoF and WCS358. Strains with mutations inppuIandppuRshowed a severe reduction of putisolvin production. Expression analysis of the putisolvin biosynthetic gene in appuIbackground showed decreased expression, which could be complemented by the addition of synthetic 3-oxo-C10-N-acyl homoserine lactone (3-oxo-C10-AHL) or 3-oxo-C12-AHL to the medium. AnrsaLmutant overproduces AHLs, and production of putisolvins is induced early during growth. Analysis of biofilm formation on polyvinylchloride showed thatppuIandppuRmutants produce a denser biofilm than PCL1445, which correlates with decreased production of putisolvins, whereas anrsaLmutant shows a delay in biofilm production, which correlates with early production of putisolvins. The results demonstrate that quorum-sensing signals induce the production of cyclic lipopeptides putisolvin I and II and consequently control biofilm formation byPseudomonas putida.
The DtxR Regulon ofCorynebacterium glutamicum
Authors: Wennerhold Julia , Bott Michael 
Published: April 2006

ABSTRACTPrevious studies withCorynebacterium diphtheriaeandMycobacteriumspecies revealed that the transcriptional regulator DtxR and its ortholog IdeR play a central role in the control of iron metabolism. In the present work, we used genome-based approaches to determine the DtxR regulon ofCorynebacterium glutamicum, a nonpathogenic relative ofC. diphtheriae. First, global gene expression of adtxRdeletion mutant was compared with that of the wild type using DNA microarrays. Second, we used a computer-based approach to identify 117 putative DtxR binding sites in theC. glutamicumgenome. In the third step, 74 of the corresponding genome regions were amplified by PCR, 51 of which were shifted by the DtxR protein. Finally, we analyzed which of the genes preceded by a functional DtxR binding site showed altered mRNA levels in the transcriptome comparison. Fifty-one genes organized in 27 putative operons displayed an increased mRNA level in theΔdtxRmutant and thus are presumably repressed by DtxR. The majority of these genes are obviously involved in iron acquisition, three encode transcriptional regulators, e.g., the recently identified repressor of iron proteins RipA, and the others encode proteins of diverse or unknown functions. Thirteen genes showed a decreased mRNA level in theΔdtxRmutant and thus might be activated by DtxR. This group included thesufoperon, whose products are involved in the formation and repair of iron-sulfur clusters, and several genes for transcriptional regulators. Our results clearly establish DtxR as the master regulator of iron-dependent gene expression inC. glutamicum.
Aerobic Benzoyl-Coenzyme A (CoA) Catabolic Pathway in
Azoarcus evansii
: Conversion of Ring Cleavage Product by 3,4-Dehydroadipyl-CoA Semialdehyde Dehydrogenase
Authors: Gescher Johannes , Ismail Wael , Ölgeschläger Ellen , Eisenreich Wolfgang , Wörth Jürgen , Fuchs Georg 
Published: April 2006

ABSTRACT

Benzoate, a strategic intermediate in aerobic aromatic metabolism, is metabolized in various bacteria via an unorthodox pathway. The intermediates of this pathway are coenzyme A (CoA) thioesters throughout, and ring cleavage is nonoxygenolytic. The fate of the ring cleavage product 3,4-dehydroadipyl-CoA semialdehyde was studied in the β-proteobacterium
Azoarcus evansii
. Cell extracts contained a benzoate-induced, NADP
+
-specific aldehyde dehydrogenase, which oxidized this intermediate. A postulated putative long-chain aldehyde dehydrogenase gene, which might encode this new enzyme, is located on a cluster of genes encoding enzymes and a transport system required for aerobic benzoate oxidation. The gene was expressed in
Escherichia coli
, and the maltose-binding protein-tagged enzyme was purified and studied. It is a homodimer composed of 54 kDa (without tag) subunits and was confirmed to be the desired 3,4-dehydroadipyl-CoA semialdehyde dehydrogenase. The reaction product was identified by nuclear magnetic resonance spectroscopy as the corresponding acid 3,4-dehydroadipyl-CoA. Hence, the intermediates of aerobic benzoyl-CoA catabolic pathway recognized so far are benzoyl-CoA; 2,3-dihydro-2,3-dihydroxybenzoyl-CoA; 3,4-dehydroadipyl-CoA semialdehyde plus formate; and 3,4-dehydroadipyl-CoA. The further metabolism is thought to lead to 3-oxoadipyl-CoA, the intermediate at which the conventional and the unorthodox pathways merge.
Green Fluorescent Chimeras Indicate Nonpolar Localization of Pullulanase Secreton Components PulL and PulM
Authors: Buddelmeijer Nienke , Francetic Olivera , Pugsley Anthony P. 
Published: April 2006

ABSTRACTTheKlebsiella oxytocapullulanase secreton (type II secretion system) components PulM and PulL were tagged at their N termini with green fluorescent protein (GFP), and their subcellular location was examined by fluorescence microscopy and fractionation. When produced at moderate levels without other secreton components inEscherichia coli, both chimeras were envelope associated, as are the native proteins. Fluorescent GFP-PulM was evenly distributed over the cell envelope, with occasional brighter foci. Under the same conditions, GFP-PulL was barely detectable in the envelope by fluorescence microscopy. When produced together with all other secreton components, GFP-PulL exhibited circumferential fluorescence, with numerous brighter patches. The envelope-associated fluorescence of GFP-PulL was almost completely abolished when native PulL was also produced, suggesting that the chimera cannot compete with PulL for association with other secreton components. The patches of GFP-PulL might represent functional secretons, since GFP-PulM also appeared in similar patches. GFP-PulM and GFP-PulL both appeared in spherical polar foci when made at high levels. InK. oxytoca, GFP-PulM was evenly distributed over the cell envelope, with few patches, whereas GFP-PulL showed only weak envelope-associated fluorescence. These data suggest that, in contrast to theirVibrio choleraeEps secreton counterparts (M. Scott, Z. Dossani, and M. Sandkvist, Proc. Natl. Acad. Sci. USA 98:13978-13983, 2001), PulM and PulL do not localize specifically to the cell poles and that the Pul secreton is distributed over the cell surface.
Transcriptional Control of the MycobacterialembCABOperon by PknH through a Regulatory Protein, EmbR, In Vivo
Authors: Sharma Kirti , Gupta Meetu , Pathak Monika , Gupta Nidhi , Koul Anil , Sarangi Smilona , Baweja Renu , Singh Yogendra 
Published: April 2006

ABSTRACTEmbR, a putative transcriptional regulator fromMycobacterium tuberculosis, is homologous to the OmpR class of transcriptional regulators that possess winged helix-turn-helix DNA binding motifs. In contrast to other OmpR-like response regulators that are usually phosphorylated and controlled by histidine kinases, EmbR was recently shown to be phosphorylated by the cognate mycobacterial serine/threonine kinase PknH. Despite the in vitro evidence of phosphorylation and interaction between the kinase and regulator, the physiological function of the PknH-EmbR pair is still unknown. We identify theembCABoperon encoding arabinosyltransferases inM. tuberculosisas the cellular target of EmbR. Phosphorylation of EmbR enhances its DNA binding activity towards promoter regions ofembCABgenes. In vivo studies involving expression of PknH inMycobacterium smegmatisestablished its positive regulatory effect on transcription of theembCABoperon via phosphorylation of EmbR. Interestingly, increased transcription ofembC, catalyzing arabinosylation of lipomannan (LM) to lipoarabinomannan (LAM), results in a high LAM/LM ratio, which in turn is a crucial factor in mycobacterial virulence. The PknH-mediated increase in the transcription ofembABgenes significantly alters resistance to ethambutol, a frontline antituberculosis drug known to targetembABgenes. These findings and in vivo upregulation of PknH inside the host macrophages suggest a functionally relevant signaling mechanism involving the PknH-EmbR-embCABsystem.
Transcription of the ContiguoussigB,dtxR, andgalEGenes inCorynebacterium diphtheriae: Evidence for Multiple Transcripts and Regulation by Environmental Factors
Authors: Oram Diana Marra , Jacobson Andrew D. , Holmes Randall K. 
Published: April 2006

ABSTRACTThe iron-dependent transcriptional regulator DtxR fromCorynebacterium diphtheriaeis the prototype for a family of metal-dependent regulators found in diverse bacterial species. The structure of DtxR and its action as a repressor have been extensively characterized, but little is known about expression ofdtxR. In the current study, we investigated transcription ofdtxRas well as thesigBandgalEgenes located immediately upstream and downstream fromdtxR, respectively. We identified two promoters that determine transcription ofdtxR.The first, located upstream ofsigB, appears to be controlled by an extracytoplasmic function σ factor. The second, located in the intergenic region betweensigBanddtxR, is similar to promoters used by the primary vegetative σ factors in other actinomycete species. Using quantitative real-time assays, we demonstrated that the number of transcripts initiated upstream fromsigBis affected by several environmental factors. In contrast, the presence of sodium dodecyl sulfate was the only factor tested that conclusively affects the number of transcripts initiated in thesigB-dtxRintergenic region. Additionally, we provided evidence for the existence of transcripts that containsigB,dtxR, andgalE. Our studies provide the first quantitative transcriptional analysis of a gene encoding a DtxR family regulator and give new insights into transcriptional regulation inC. diphtheriae.