bluuebunny/crossref_metadata_2025_split huggingface dataset:
Organization and Regulation of Pentachlorophenol-Degrading Genes in
Sphingobium chlorophenolicum
ATCC 39723
Authors: Cai Mian , Xun Luying 
Published: October 2002

ABSTRACT

The first three enzymes of the pentachlorophenol (PCP) degradation pathway in
Sphingobium chlorophenolicum
(formerly
Sphingomonas chlorophenolica
) ATCC 39723 have been characterized, and the corresponding genes,
pcpA
,
pcpB
, and
pcpC
, have been individually cloned and sequenced. To search for new genes involved in PCP degradation and map the physical locations of the
pcp
genes, a 24-kb fragment containing
pcpA
and
pcpC
was completely sequenced. A putative LysR-type transcriptional regulator gene,
pcpM
, and a maleylacetate reductase gene,
pcpE
, were identified upstream of
pcpA. pcpE
was found to play a role in PCP degradation.
pcpB
was not found on the 24-kb fragment. The four gene products PcpB, PcpC, PcpA, and PcpE were responsible for the metabolism of PCP to 3-oxoadipate in ATCC 39723, and inactivational mutation of each gene disrupted the degradation pathway. The organization of the
pcp
genes is unusual because the four PCP-degrading genes,
pcpA
,
pcpB
,
pcpC
, and
pcpE
, were found to be located at four discrete locations. Two hypothetical LysR-type regulator genes,
pcpM
and
pcpR,
have been identified;
pcpM
was not required, but
pcpR
was essential for the induction of
pcpB
,
pcpA
, and
pcpE
. The coinducers of PcpR were PCP and other polychlorinated phenols. The expression of
pcpC
was constitutive. Thus, the organization and regulation of the genes involved in PCP degradation to 3-oxoadipate were documented.
Pseudomonas aeruginosa
Synthesizes Phosphatidylcholine by Use of the Phosphatidylcholine Synthase Pathway
Authors: Wilderman Paula J. , Vasil Adriana I. , Martin Wesley E. , Murphy Robert C. , Vasil Michael L. 
Published: October 2002

ABSTRACT

Phosphatidylcholine (PC) is a ubiquitous membrane lipid in eukaryotes but has been found in only a limited number of prokaryotes. Both eukaryotes and prokaryotes synthesize PC by methylating phosphatidylethanolamine (PE) by use of a phospholipid methyltransferase (Pmt). Eukaryotes can synthesize PC by the activation of choline to form choline phosphate and then CDP-choline. The CDP-choline then condenses with diacylglycerol (DAG) to form PC. In contrast, prokaryotes condense choline directly with CDP-DAG by use of the enzyme PC synthase (Pcs). PmtA was the first enzyme identified in prokaryotes that catalyzes the synthesis of PC, and Pcs in
Sinorhizobium meliloti
was characterized. The completed release of the
Pseudomonas aeruginosa
PAO1 genomic sequence contains on open reading frame predicted to encode a protein that is highly homologous (35% identity, 54% similarity) to PmtA from
Rhodobacter sphaeroides
. Moreover, the
P. aeruginosa
PAO1 genome encodes a protein with significant homology (39% amino acid identity) to Pcs of
S. meliloti
. Both the
pcs
and
pmtA
homologues were cloned from PAO1, and homologous sequences were found in almost all of the
P. aeruginosa
strains examined. Although the pathway for synthesizing PC by use of Pcs is functional in
P. aeruginosa
, it does not appear that this organism uses the PmtA pathway for PC synthesis. We demonstrate that the PC synthesized by
P. aeruginosa
PAO1 localized to both the inner and outer membranes, where it is readily accessible to its periplasmic, PC-specific phospholipase D.
LasR, a Transcriptional Activator of
Pseudomonas aeruginosa
Virulence Genes, Functions as a Multimer
Authors: Kiratisin Pattarachai , Tucker Kenneth D. , Passador Luciano 
Published: October 2002

ABSTRACT

The
Pseudomonas aeruginosa
LasR protein functions in concert with
N
-3-oxo-dodecanoyl-
l
-homoserine lactone (3O-C
12
-HSL) to coordinate the expression of target genes, including many genes that encode virulence factors, with cell density. We used a LexA-based protein interaction assay to demonstrate that LasR forms multimers only when 3O-C
12
-HSL is present. A series of LasR molecules containing internal deletions or substitutions in single, conserved amino acid residues indicated that the N-terminal portion of LasR is required for multimerization. Studies performed with these mutant versions of LasR demonstrated that the ability of LasR to multimerize correlates with its ability to function as a transcriptional activator of
lasI
, a gene known to be tightly regulated by the LasR-3O-C
12
-HSL regulatory system. A LasR molecule that carries a C-terminal deletion can function as a dominant-negative mutant in
P. aeruginosa
, as shown by its ability to decrease expression of
lasB,
another LasR-3O-C
12
-HSL target gene. Taken together, our data strongly support the hypothesis that LasR functions as a multimer in vivo.
The
metD
d
-Methionine Transporter Locus of
Escherichia coli
Is an ABC Transporter Gene Cluster
Authors: Gál József , Szvetnik Attila , Schnell Róbert , Kálmán Miklós 
Published: October 2002

ABSTRACT

The
metD
d
-methionine transporter locus of
Escherichia coli
was identified as the
abc
-
yaeE-yaeC
cluster (now renamed
metNIQ
genes). The
abc
open reading frame is preceded by tandem MET boxes bracketed by the −10 and −35 boxes of a promoter. The expression driven by this promoter is controlled by the MetJ repressor and the level of methionine.
The Spectrum of Spontaneous Rifampin Resistance Mutations in the
rpoB
Gene of
Bacillus
subtilis
168 Spores Differs from That of Vegetative Cells and Resembles That of
Mycobacterium tuberculosis
Authors: Nicholson Wayne L. , Maughan Heather 
Published: October 2002

ABSTRACT

Mutations causing rifampin resistance in vegetative cells of
Bacillus subtilis
168 have thus far been mapped to a rather restricted set of alterations at either Q469 or H482 within cluster I of the
rpoB
gene encoding the β subunit of RNA polymerase. In this study, we demonstrated that spores of
B. subtilis
168 exhibit a spectrum of spontaneous rifampin resistance mutations distinct from that of vegetative cells. In addition to the
rpoB
mutations Q469K, Q469R, and H482Y previously characterized in vegetative cells, we isolated a new mutation of
rpoB
, H482R, from vegetative cells. Additional new rifampin resistance mutations arising from spores were detected at A478N and most frequently at S487L. The S487L change is the predominant change found in
rpoB
mutations sequenced from rifampin-resistant clinical isolates of
Mycobacterium tuberculosis.
The observations are discussed in terms of the underlying differences of the DNA environment within dormant cells and vegetatively growing cells.
Synergistic Effects on Crystalline Cellulose Degradation between Cellulosomal Cellulases fromClostridium cellulovorans
Authors: Murashima Koichiro , Kosugi Akihiko , Doi Roy H. 
Published: October 2002

ABSTRACTClostridium cellulovoransproduces a multienzyme cellulose-degrading complex called the cellulosome. In this study, we determined the synergistic effects on crystalline cellulose degradation by three different recombinant cellulosomes containing either endoglucanase EngE, endoglucanase EngH, or exoglucanase ExgS bound to mini-CbpA, a part of scaffolding protein CbpA. EngE, EngH, and ExgS are classified into the glycosyl hydrolase families 5, 9, and 48, respectively. The assembly of ExgS and EngH with mini-CbpA increased the activity against insoluble cellulose 1.5- to 3-fold, although no effects on activity against soluble cellulose were observed. These results indicated that mini-CbpA could help cellulase components degrade insoluble cellulose but not soluble cellulose. The mixture of the cellulosomes containing ExgS and EngH showed higher activity and synergy degrees than the other cellulosome mixtures, indicating the synergistic effect between EngH and ExgS was the most dominant effect among the three mixtures for crystalline cellulose degradation. Reactions were also performed by adding different cellulosomes in a sequential manner. When ExgS was used for the initial reaction followed by EngE and EngH, almost no synergistic effect was observed. On the other hand, when EngE or EngH was used for the first reaction followed by ExgS, synergistic effects were observed. These results indicated that the initial reactions by EngH and/or EngE promoted cellulose degradation by ExgS.
ExbB and ExbD Do Not Function Independently in TonB-Dependent Energy Transduction
Authors: Held Kiara G. , Postle Kathleen 
Published: October 2002

ABSTRACTExbB and ExbD proteins are part of the TonB-dependent energy transduction system and are encoded by theexboperon inEscherichia coli. TonB, the energy transducer, appears to go through a cycle during energy transduction, with the absence of both ExbB and ExbD creating blocks at two points: (i) in the inability of TonB to respond to the cytoplasmic membrane proton motive force and (ii) in the conversion of TonB from a high-affinity outer membrane association to a high-affinity cytoplasmic membrane association. The recent observation that ExbB exists in 3.5-fold molar excess relative to the molarity of ExbD inE. colisuggests the possibility of two types of complexes, those containing both ExbB and ExbD and those containing only ExbB. Such distinct complexes might individually manifest one of the two activities described above. In the present study this hypothesis was tested and rejected. Specifically, both ExbB and ExbD were found to be required for TonB to conformationally respond to proton motive force. Both ExbB and ExbD were also required for association of TonB with the cytoplasmic membrane. Together, these results support an alternative model where all of the ExbB in the cell occurs in complex with all of the ExbD in the cell. Based on recently determined cellular ratios of TonB system proteins, these results suggest the existence of a cytoplasmic membrane complex that may be as large as 520 kDa.
Global Expression Profile of
Bacillus subtilis
Grown in the Presence of Sulfate or Methionine
Authors: Auger Sandrine , Danchin Antoine , Martin-Verstraete Isabelle 
Published: October 2002

ABSTRACT

DNA arrays were used to investigate the global transcriptional profile of
Bacillus subtilis
grown in the presence of sulfate or methionine as the sole sulfur source. The expression of at least 56 genes differed significantly under the two growth conditions. The expression of several genes belonging to the S-box regulon was repressed in the presence of methionine probably in response to
S
-adenosylmethionine availability. The expression of genes encoding transporters (
yhcL
,
ytmJKLMN
, and
yxeMO
) was high when the sulfur source was methionine or taurine and reduced when it was sulfate.
Amplification of the Tetracycline Resistance Determinant of pAMα1 in
Enterococcus faecalis
Requires a Site-Specific Recombination Event Involving Relaxase
Authors: Francia M. Victoria , Clewell Don B. 
Published: October 2002

ABSTRACT

The small multicopy plasmid pAMα1 (9.75 kb) encoding tetracycline resistance in
Enterococcus faecalis
is known to generate tandem repeats of a 4.1-kb segment carrying
tet
(L) when cells are grown extensively in the presence of tetracycline. Here we show that the initial (rate-limiting) step involves a site-specific recombination event involving plasmid-encoded relaxase activity acting at two recombination sequences (RS1 and RS2) that flank the
tet
determinant. We also present the complete nucleotide sequence of pAMα1.
Rapid Detection of Methicillin-Resistant Staphylococci from Blood Culture Bottles by Using a Multiplex PCR Assay
Authors: Louie L. , Goodfellow J. , Mathieu P. , Glatt A. , Louie M. , Simor A. E. 
Published: October 2002

ABSTRACT

Rapid detection and accurate identification of methicillin-resistant staphylococci are critical for the effective management of infections caused by these organisms. We describe a multiplex PCR-based assay for the direct detection of methicillin-resistant staphylococci from blood culture bottles (BacT/Alert; Organon-Teknika, Durham, N.C.). A simple lysis method followed by a multiplex PCR assay designed to detect the
nuc
,
mecA
, and bacterial 16S rRNA genes was performed. A total of 306 blood culture specimens were collected over a period of 10 months from June 1998 to April 1999, consisting of 236 blood cultures growing staphylococci (including 124 methicillin-resistant
Staphylococcus
spp.), 50 positive blood cultures which grew organisms other than staphylococci, and 20 blood cultures that were negative for bacterial and fungal pathogens after 5 days of incubation and terminal subculture. DNA extraction, PCR, and detection could be completed in 2.5 h. Of the positive blood cultures with staphylococci, the multiplex PCR assay had a sensitivity and specificity of 99.2% and 100%, respectively. Our results show that rapid, direct detection of methicillin-resistant staphylococci is possible, allowing clinicians to make prompt and effective decisions for the management of patients with staphylococcal bacteremia.
Diagnosis of Community-Acquired Pertussis Infection: Comparison of Both Culture and Fluorescent-Antibody Assays with PCR Detection Using Electrophoresis or Dot Blot Hybridization
Authors: Lingappa Jairam R. , Lawrence William , West-Keefe Sheyla , Gautom Romesh , Cookson Brad T. 
Published: October 2002

ABSTRACTDiagnosis ofBordetella pertussisinfection has been difficult due to the low sensitivity of culture. PCR tests have been shown to be more sensitive than culture, but the reported sensitivity of PCR is variable. We evaluated PCR product detection by using either agarose gel electrophoresis (PCR-gel) or dot blot hybridization with32P-labeled oligonucleotide probes, and we compared these methods to both culture and direct fluorescent-antibody (DFA) assays with microscopy for the detection of pertussis. This was done with 225 nasopharyngeal swab specimens collected in community clinic settings. The multiplexed PCR amplified the multiply repeated IS481 B. pertussissequence and a sequence from the human globin gene as a positive control for specimen adequacy. Of 225 specimens, 179 were judged to be adequate for PCR analysis. Among the adequate specimens, 9, 4, and 10 were culture, DFA, and PCR-gel positive, respectively. The sensitivity of PCR-gel versus culture was 89% while the sensitivity of culture versus PCR-gel was 80%. DFA had the lowest sensitivity. Thirty specimens were positive by PCR with dot blot hybridization; no negative control specimens showed a signal above the background. Among the 79 (44%) adequate specimens with clinical data available, the rates of reported cough or persistent cough were similar for persons who were pertussis positive by each assay. The IS481PCR, with either electrophoresis or dot blot hybridization, is a sensitive assay; however, at this time it cannot completely replace culture without an overall loss in sensitivity for the detection of pertussis. Further study is required to understand the clinical significance ofB. pertussisPCR products detected by dot blot hybridization alone.
Cytomegalovirus Assemblin (pUL80a): Cleavage at Internal Site Not Essential for Virus Growth; Proteinase Absent from Virions
Authors: Chan Chee-Kai , Brignole Edward J. , Gibson Wade 
Published: October 2002

ABSTRACT
The human cytomegalovirus (HCMV) maturational proteinase is synthesized as an enzymatically active 74-kDa precursor that cleaves itself at four sites. Two of these, called the maturational (M) and release (R) sites, are conserved in the homologs of all herpesviruses. The other two, called the internal (I) and cryptic (C) sites, have recognized consensus sequences only among cytomegalovirus (CMV) homologs and are located in the 28-kDa proteolytic portion of the precursor, called assemblin. I-site cleavage cuts assemblin in half without detected effect on its enzymatic behavior in vitro. To investigate the requirement for this cleavage during virus infection, we used the CMV-bacterial artificial chromosome system (E. M. Borst, G. Hahn, U. H. Koszinowski, and M. Messerle, J. Virol. 73:8320-8329, 1999) to construct a virus encoding a mutant I site (Ala143 to Val) intended to be blocked for cleavage. Characterizations of the resulting mutant (i) confirmed the presence of the mutation in the viral genome and the inability of the mutant virus to effect I-site cleavage in infected cells; (ii) determined that the mutation has no gross effect on the rate of virus production or on the amounts of extracellular virions, noninfectious enveloped particles (NIEPs), and dense bodies; (iii) established that assemblin and its cleavage products are present in NIEPs but are absent from CMV virions, an apparent difference from what is found for virions of herpes simplex virus; and (iv) showed that the 23-kDa protein product of C-site cleavage is more abundant in mutant virus-than in wild-type virus-infected cells and NIEPs. We conclude that the production of infectious CMV requires neither I-site cleavage of assemblin nor the presence of assemblin in the mature virion.
Hemagglutinin Protein of Wild-Type Measles Virus Activates Toll-Like Receptor 2 Signaling
Authors: Bieback Karen , Lien Egil , Klagge Ingo M. , Avota Elita , Schneider-Schaulies Jürgen , Duprex W. Paul , Wagner Herrmann , Kirschning Carsten J. , ter Meulen Volker , Schneider-Schaulies Sibylle 
Published: October 2002

ABSTRACTPattern recognition via Toll-like receptors (TLR) by antigen-presenting cells is an important element of innate immunity. We report that wild-type measles virus but not vaccine strains activate cells via both human and murine TLR2, and this is a property of the hemagglutinin (H) protein. The ability to activate cells via TLR2 by wild-type MV H protein is abolished by mutation of a single amino acid, asparagine at position 481 to tyrosine, as is found in attenuated strains, which is important for interaction with CD46, the receptor for these strains. TLR2 activation by MV wild-type H protein stimulates induction of proinflammatory cytokines such as interleukin-6 (IL-6) in human monocytic cells and surface expression of CD150, the receptor for all MV strains. Confirming the specificity of this interaction, wild-type H protein did not induce IL-6 release in macrophages from TLR2−/−mice. Thus, the unique property of MV wild-type strains to activate TLR2-dependent signals might essentially contribute not only to immune activation but also to viral spread and pathogenicity by upregulating the MV receptor on monocytes.
Adenovirus Type 37 Uses Sialic Acid as a Cellular Receptor on Chang C Cells
Authors: Arnberg Niklas , Pring-Åkerblom Patricia , Wadell Göran 
Published: October 2002

ABSTRACT

Epidemic keratoconjunctivitis (EKC) is a severe eye infection caused mainly by adenovirus type 8 (Ad8), Ad19, and Ad37. We have shown that the EKC-causing adenoviruses use sialic acid as a cellular receptor on A549 cells instead of the coxsackie-adenovirus receptor, which is used by most adenoviruses. Recently, Wu et al. (Virology 279:78-89, 2001) proposed that Ad37 uses a 50-kDa protein as a receptor on Chang C conjunctival cells and that this interaction is independent of sialic acid. According to the American Type Culture Collection, this cell line carries HeLa cell markers and should be considered to be a genital cell line. This prompted us to investigate the function of sialic acid as a cellular receptor for Ad37 in Chang C cells. In this study, we demonstrate that enzymatic removal or lectin-mediated blocking of cell surface sialic acid inhibits the binding of Ad37 virions to Chang C cells, as does soluble, virion-interacting sialic acid-containing substances. The binding was Ca
2+
or Mg
2+
ion independent and mediated by the knob domain of the trimeric viral fiber polypeptide. Moreover, Ad37 virions infected Chang C cells and two other genital cell lines (HeLa and SiHa) as well as a corneal cell line in a strictly sialic acid-dependent manner. From these results, we conclude that Ad37 uses sialic acid as a major receptor in cell lines derived from both genital and corneal tissues.
Transgenic Expression in Mouse Lung Reveals Distinct Biological Roles for the Adenovirus Type 5 E1A 243- and 289-Amino-Acid Proteins
Authors: Yang Yongping , McKerlie Colin , Borenstein Steven H. , Lu Zhan , Schito Marco , Chamberlain John W. , Buchwald Manuel 
Published: October 2002

ABSTRACTLittle is known about the biological significance of human adenovirus type 5 (Ad5) E1A in vivo. However, Ad5 E1A is well defined in vitro and can be detected frequently in the lungs of patients with pulmonary disease. Transgenic expression of the Ad5E1Agene targeted to the mouse lung reveals distinct biological effects caused by two Ad5 E1A products. Either of two Ad5 E1A proteins was preferentially expressed in vivo in the transgenic lungs. The preferential expression of the Ad5 E1A 243-amino-acid (aa) protein at a moderate level was associated with cellular hyperplasia, nodular lesions of proliferating lymphocyte-like cells, and a low level of p53-dependent apoptosis in the lungs of transgenic mice. In contrast, the preferential expression of the Ad5 E1A 289-aa protein at a moderate level resulted in a proapoptotic injury and an acute pulmonary proinflammation in the lungs of transgenic mice, mediated by multiple apoptotic pathways, as well as an enhancement of the host immune cell response. Expression of the Ad5 E1A 243-aa protein resulted in proliferation-stimulated p53 upregulation, while expression of the Ad5 E1A 289-aa protein led to DNA damage-induced p53 activation. These data suggest that the Ad5 E1A 243- and 289-aa proteins lead to distinct biological roles in vivo.
Decelerating Decay of Latently Infected Cells during Prolonged Therapy for Human Immunodeficiency Virus Type 1 Infection
Authors: Müller Viktor , Vigueras-Gómez Javier Flavio , Bonhoeffer Sebastian 
Published: October 2002

ABSTRACT
Antiviral therapy induces a rapid drop in human immunodeficiency virus type 1 viremia, but the decline of virus levels decelerates over time. Mathematical modeling demonstrates that the source of residual virus production might be a single compartment of latently infected cells with an extended distribution of activation rates.