bluuebunny/crossref_metadata_2025_split huggingface dataset:
Changes in Biochemical and Phenotypic Properties of
Streptococcus mutans
during Growth with Aeration
Authors: Ahn Sug-Joon , Ahn Sang-Joon , Browngardt Christopher M. , Burne Robert A. 
Published: February 2009

ABSTRACT

Oxygen has a potent influence on the expression of genes and the activity of physiological and biochemical pathways in bacteria. We have found that oxygen significantly altered virulence-related phenotypic properties of
Streptococcus mutans
, the primary etiological agent of human dental caries. Transport of glucose, fructose, or mannose by the sugar:phosphotransferase system was significantly enhanced by growth under aerobic conditions, whereas aeration caused an extended lag phase and slower growth of
S. mutans
in medium containing glucose, fructose, or mannose as the carbohydrate source. Aeration resulted in a decrease in the glycolytic rate and enhanced the production of intracellular storage polysaccharides. Although aeration decreased the acid tolerance of
S. mutans
, aerobically grown cells had higher F-ATPase activity. Aeration altered biofilm architecture but did not change the ability of
S. mutans
to interact with salivary agglutinin. Growth in air resulted in enhanced cell-associated glucosyltransferase (Gtf) activity at the expense of cell-free Gtf activity. These results demonstrate that
S. mutans
can dramatically alter its pathogenic potential in response to exposure to oxygen, suggesting that the phenotype of the organism may be highly variable in the human oral cavity depending on the maturity of the dental plaque biofilm.
Heterologous Production of Methionine-γ-Lyase from
Brevibacterium linens
in
Lactococcus lactis
and Formation of Volatile Sulfur Compounds
Authors: Hanniffy Sean B. , Philo Mark , Peláez Carmen , Gasson Michael J. , Requena Teresa , Martínez-Cuesta M. C. 
Published: February 2009

ABSTRACT

The conversion of methionine to volatile sulfur compounds (VSCs) is of great importance in flavor formation during cheese ripening and is the focus of biotechnological approaches toward flavor improvement. A synthetic
mgl
gene encoding methionine-γ-lyase (MGL) from
Brevibacterium linens
BL2 was cloned into a
Lactococcus lactis
expression plasmid under the control of the nisin-inducible promoter P
nisA
. When expressed in
L. lactis
and purified as a recombinant protein, MGL was shown to degrade
l
-methionine as well as other sulfur-containing compounds such as
l
-cysteine,
l
-cystathionine, and
l
-cystine. Overproduction of MGL in recombinant
L. lactis
also resulted in an increase in the degradation of these compounds compared to the wild-type strain. Importantly, gas chromatography-mass spectrometry analysis identified considerably higher formation of methanethiol (and its oxidized derivatives dimethyl disulfide and dimethyl trisulfide) in reactions containing either purified protein, whole cells, or cell extracts from the heterologous
L. lactis
strain. This is the first report of production of MGL from
B. linens
in
L. lactis
. Given their significance in cheese flavor development, the use of lactic acid bacteria with enhanced VSC-producing abilities could be an efficient way to enhance cheese flavor development.
Temperature-Dependent Phage Resistance of
Listeria monocytogenes
Epidemic Clone II
Authors: Kim Jae-Won , Kathariou Sophia 
Published: February 2009

ABSTRACT

Listeria monocytogenes
epidemic clone II (ECII) has been responsible for two multistate outbreaks in the United States in 1998-1999 and in 2002, in which contaminated ready-to-eat meat products (hot dogs and turkey deli meats, respectively) were implicated. However, ecological adaptations of ECII strains in the food-processing plant environment remain unidentified. In this study, we found that broad-host-range phages, including phages isolated from the processing plant environment, produced plaques on ECII strains grown at 37°C but not when the bacteria were grown at lower temperatures (30°C or below). ECII strains grown at lower temperatures were resistant to phage regardless of the temperature during infection and subsequent incubation. In contrast, the phage susceptibility of all other tested strains of serotype 4b (including epidemic clone I) and of strains of other serotypes and
Listeria
species was independent of the growth temperature of the bacteria. This temperature-dependent phage susceptibility of ECII bacteria was consistently observed with all surveyed ECII strains from outbreaks or from processing plants, regardless of the presence or absence of cadmium resistance plasmids. Phages adsorbed similarly on ECII bacteria grown at 25°C and at 37°C, suggesting that resistance of ECII strains grown at 25°C was not due to failure of the phage to adsorb. Even though the underlying mechanisms remain to be elucidated, temperature-dependent phage resistance may represent an important ecological adaptation of
L. monocytogenes
ECII in processed, cold-stored foods and in the processing plant environment, where relatively low temperatures prevail.
Cloning, Expression, Characterization, and Biocatalytic Investigation of the 4-Hydroxyacetophenone Monooxygenase from
Pseudomonas putida
JD1
Authors: Rehdorf Jessica , Zimmer Christian L. , Bornscheuer Uwe T. 
Published: February 2009

ABSTRACT

While the number of available recombinant Baeyer-Villiger monooxygenases (BVMOs) has grown significantly over the last few years, there is still the demand for other BVMOs to expand the biocatalytic diversity. Most BVMOs that have been described are dedicated to convert efficiently cyclohexanone and related cyclic aliphatic ketones. To cover a broader range of substrate types and enantio- and/or regioselectivities, new BVMOs have to be discovered. The gene encoding a BVMO identified in
Pseudomonas putida
JD1 converting aromatic ketones (HAPMO; 4-hydroxyacetophenone monooxygenase) was amplified from genomic DNA using SiteFinding-PCR, cloned, and functionally expressed in
Escherichia coli
. Furthermore, four other open reading frames could be identified clustered around this HAPMO. It has been suggested that these proteins, including the HAPMO, might be involved in the degradation of 4-hydroxyacetophenone. Substrate specificity studies revealed that a large variety of other arylaliphatic ketones are also converted via Baeyer-Villiger oxidation into the corresponding esters, with preferences for
para
-substitutions at the aromatic ring. In addition, oxidation of aldehydes and some heteroaromatic compounds was observed. Cycloketones and open-chain ketones were not or poorly accepted, respectively. It was also found that this enzyme oxidizes aromatic ketones such as 3-phenyl-2-butanone with excellent enantioselectivity (E ≫100).
Antibody-Mediated Immobilization of
Cryptococcus neoformans
Promotes Biofilm Formation
Authors: Robertson Emma J. , Casadevall Arturo 
Published: February 2009

ABSTRACT

Most microbes, including the fungal pathogen
Cryptococcus neoformans
, can grow as biofilms. Biofilms confer upon microbes a range of characteristics, including an ability to colonize materials such as shunts and catheters and increased resistance to antibiotics. Here, we provide evidence that coating surfaces with a monoclonal antibody to glucuronoxylomannan, the major component of the fungal capsular polysaccharide, immobilizes cryptococcal cells to a surface support and, subsequently, promotes biofilm formation. We used time-lapse microscopy to visualize the growth of cryptococcal biofilms, generating the first movies of fungal biofilm growth. We show that when fungal cells are immobilized using surface-attached specific antibody to the capsule, the initial stages of biofilm formation are significantly faster than those on surfaces with no antibody coating or surfaces coated with unspecific monoclonal antibody. Time-lapse microscopy revealed that biofilm growth was a dynamic process in which cells shuffled position during budding and was accompanied by emergence of planktonic variant cells that left the attached biofilm community. The planktonic variant cells exhibited mobility, presumably by Brownian motion. Our results indicate that microbial immobilization by antibody capture hastens biofilm formation and suggest that antibody coating of medical devices with immunoglobulins must exclude binding to common pathogenic microbes and the possibility that this effect could be exploited in industrial microbiology.
Four-Antigen Mixture Containing V-Cyclin for Serological Screening of Human Herpesvirus 8 Infection
Authors: Burbelo Peter D. , Leahy Hannah P. , Groot Sandra , Bishop Lisa R. , Miley Wendell , Iadarola Michael J. , Whitby Denise , Kovacs Joseph A. 
Published: March 2009

ABSTRACT

Improved diagnostic reagents and testing are currently needed for the serological detection of human herpesvirus 8 (HHV-8) infections. We evaluated the luciferase immunoprecipitation systems (LIPS) for profiling antibody responses to a panel of HHV-8 proteins for diagnosis of Kaposi sarcoma (KS)-infected individuals. Using a pilot serum set, LIPS detected robust antibody responses to several known antigens, and a screen of 14 additional HHV-8 proteins identified v-cyclin as a potentially new diagnostic antigen. In evaluating a training-serum set, a four-antigen panel (K8.1, v-cyclin, ORF65, and a LANA fragment) was found to provide sufficient information for diagnosis. Analysis of a validation serum set using the combined results from these four separate antigen tests showed 100% sensitivity and 100% specificity. Furthermore, a LIPS format using a mixture of the four antigens, which simplifies data collection and analysis, closely matched the diagnostic performance of the combined separate tests (
R
= 0.95). This four-antigen mixture format analyzed with the validation serum set also showed 100% sensitivity and 100% specificity but was not statistically different from two separate enzyme-linked immunosorbent assays (94% sensitivity and 100% specificity) using baculovirus-produced LANA and bacterially produced K8.1. Heat map analysis of KS patient antibody titers revealed marked heterogeneity in humoral responses to this four-antigen panel. Overall, the LIPS assay showed 97% sensitivity, and positive anti-v-cyclin antibodies were detected in approximately 75% of the KS sera. These results suggest that LIPS screening using an antigen mixture is a sensitive and high-throughput method for serological screening of HHV-8 infection in individuals with KS.
Iron Activates In Vivo DNA Binding of
Schizosaccharomyces pombe
Transcription Factor Fep1 through Its Amino-Terminal Region
Authors: Jbel Mehdi , Mercier Alexandre , Pelletier Benoit , Beaudoin Jude , Labbé Simon 
Published: February 2009

ABSTRACT

In
Schizosaccharomyces pombe
, the iron sensor Fep1 mediates the transcriptional repression of iron transport genes in response to high concentrations of iron. On the other hand,
fep1

+

expression is downregulated under conditions of iron starvation by the CCAAT-binding factor Php4. In this study, we created a
fep1
Δ
php4
Δ double mutant strain where expression of
fep1

+

was disengaged from its iron limitation-dependent repression by Php4 to examine the effects of iron on constitutively expressed functional
fep1

+

-
GFP
and
TAP-fep1

+

alleles and their gene products. In these cells, Fep1-green fluorescent protein was invariably localized in the nucleus under both iron-limiting and iron-replete conditions. Using chromatin immunoprecipitation assays, we found that Fep1 is associated with iron-responsive promoters in vivo. Chromatin binding was iron dependent, with a loss of binding observed in the presence of low iron. Functional dissection of the protein revealed that the N-terminal 241-residue segment that includes two consensus Cys
2
/Cys
2
-type zinc finger motifs and a Cys-rich region is required for optimal promoter occupancy by Fep1. Within this segment, a minimal module encompassing amino acids 60 to 241 is sufficient for iron-dependent chromatin binding. Using yeast one-hybrid analysis, we showed that the replacement of the repression domain of Fep1 by fusing the activation domain of VP16 to the chromatin-binding fragment of amino acids 1 to 241 of Fep1 converts the protein from an iron-dependent repressor into an iron-dependent transcriptional activator. Thus, the repression function of Fep1 can be replaced with that of a transcriptional activation function without the loss of its iron-dependent DNA-binding activity.
Yap1-Regulated Glutathione Redox System Curtails Accumulation of Formaldehyde and Reactive Oxygen Species in Methanol Metabolism of
Pichia pastoris
Authors: Yano Taisuke , Takigami Emiko , Yurimoto Hiroya , Sakai Yasuyoshi 
Published: February 2009

ABSTRACT

The glutathione redox system, including the glutathione biosynthesis and glutathione regeneration reaction, has been found to play a critical role in the yeast
Pichia pastoris
during growth on methanol, and this regulation was at least partly executed by the transcription factor PpYap1. During adaptation to methanol medium, PpYap1 transiently localized to the nucleus and activated the expression of the glutathione redox system and upregulated glutathione reductase 1 (Glr1). Glr1 activates the regeneration of the reduced form of glutathione (GSH). Depletion of Glr1 caused a severe growth defect on methanol and hypersensitivity to formaldehyde (HCHO), which could be complemented by addition of GSH to the medium. Disruption of the genes for the HCHO-oxidizing enzymes PpFld1 and PpFgh1 caused a comparable phenotype, but disruption of the downstream gene Pp
FDH1
did not, demonstrating the importance of maintaining intracellular GSH levels. Absence of the peroxisomal glutathione peroxidase Pmp20 also triggered nuclear localization of PpYap1, and although cells were not sensitive to HCHO, growth on methanol was again severely impaired due to oxidative stress. Thus, the PpYap1-regulated glutathione redox system has two important roles, i.e., HCHO metabolism and detoxification of reactive oxygen species.
Candida albicans RFX2
Encodes a DNA Binding Protein Involved in DNA Damage Responses, Morphogenesis, and Virulence
Authors: Hao Binghua , Clancy Cornelius J. , Cheng Shaoji , Raman Suresh B. , Iczkowski Kenneth A. , Nguyen M. Hong 
Published: February 2009

ABSTRACT

We previously showed that
Candida albicans
orf19.4590, which we have renamed
RFX2
, expresses a protein that is reactive with antibodies in persons with candidiasis. In this study, we demonstrate that
C. albicans RFX2
shares some functional redundancy with
Saccharomyces cerevisiae RFX1
. Complementation of an
S. cerevisiae rfx1
mutant with
C. albicans RFX2
partially restored UV susceptibility and the repression of DNA damage response genes. DNA damage- and UV-induced genes
RAD6
and
DDR48
were derepressed in a
C. albicans rfx2
null mutant strain under basal conditions, and the mutant was significantly more resistant to UV irradiation, heat shock, and ethanol than wild-type strain SC5314. The
rfx2
mutant was hyperfilamentous on solid media and constitutively expressed hypha-specific genes
HWP1
,
ALS3
,
HYR1
,
ECE1
, and
CEK1
. The mutant also demonstrated increased invasion of solid agar and significantly increased adherence to human buccal epithelial cells. During hematogenously disseminated candidiasis, mice infected with the mutant had a significantly delayed time to death compared to the wild type. During oropharyngeal candidiasis, mice infected with the mutant had significantly lower tissue burdens in the oral cavity and esophagus at 7 days and they were less likely to develop disseminated infections because of mucosal translocation. The data demonstrate that
C. albicans
Rfx2p regulates DNA damage responses, morphogenesis, and virulence.
Aspergillus fumigatus
Calcipressin CbpA Is Involved in Hyphal Growth and Calcium Homeostasis
Authors: Pinchai Nadthanan , Perfect B. Zachary , Juvvadi Praveen Rao , Fortwendel Jarrod R. , Cramer Robert A. , Asfaw Yohannes G. , Heitman Joseph , Perfect John R. , Steinbach William J. 
Published: February 2009

ABSTRACT

Calcineurin is a conserved protein phosphatase that plays a critical role in Ca
2+
signaling and stress responses. Previously, a new class of conserved calcineurin-binding proteins, the calcipressins, was identified. However, the role of these proteins remains controversial, and both inhibitory and stimulatory effects on calcineurin were observed. In this study, we investigate the role of CbpA, the
Aspergillus fumigatus
member of the calcipressin family, and report that deletion of the
cbpA
gene resulted in reduced hyphal growth and limited attenuated virulence. Interestingly, under high-calcium-level conditions, the Δ
cbpA
strain displayed improved Ca
2+
tolerance compared to the wild-type strain and revealed increased expression of
vcxA
,
chsA
, and
cnaA
, which encode the vacuolar Ca
2+
/H
+
exchanger VcxA, chitin synthase A, and the calcineurin catalytic subunit CnaA, respectively. The increased transcript levels of these three genes were reversed in the presence of the calcineurin inhibitor FK506, indicating a calcineurin-dependent mechanism. Overexpression of
cbpA
resulted in decreased transcription of
vcxA
,
chsA
, and
cnaA
, associated with wild-type sensitivity to Ca
2+
. Taken together, our study highlights the importance of CbpA in the regulation of hyphal growth and calcium adaptation of
A. fumigatus
and provides evidence that CbpA may serve as a feedback inhibitor in some aspects of calcineurin functions.
Deregulation of
DSE1
Gene Expression Results in Aberrant Budding within the Birth Scar and Cell Wall Integrity Pathway Activation in
Saccharomyces cerevisiae
Authors: Frýdlová Ivana , Malcová Ivana , Vašicová Pavla , Hašek Jiří 
Published: February 2009

ABSTRACT

Strains of
Saccharomyces cerevisiae
lacking Isw2, the catalytic subunit of the Isw2 chromatin remodeling complex, show the mating type-independent activation of the cell wall integrity (CWI) signaling pathway. Since the CWI pathway activation usually reflects cell wall defects, we searched for the cell wall-related genes changed in expression. The genes
DSE1
,
CTS1
, and
CHS1
were upregulated as a result of the absence of Isw2, according to previously published gene expression profiles (I. Frydlova, M. Basler, P. Vasicova, I. Malcova, and J. Hasek, Curr. Genet. 52:87-95, 2007). Western blot analyses of double deletion mutants, however, did not indicate the contribution of the chitin metabolism-related genes
CTS1
and
CHS1
to the CWI pathway activation. Nevertheless, the deletion of the
DSE1
gene encoding a daughter cell-specific protein with unknown function suppressed CWI pathway activation in
isw2
Δ cells. In addition, the deletion of
DSE1
also abolished the budding-within-the-birth-scar phenotype of
isw2
Δ cells. The plasmid-driven overexpression proved that the deregulation of Dse1 synthesis was also responsible for CWI pathway activation and manifestation of the budding-within-the-birth-scar phenotype in wild-type cells. The overproduced Dse1-green fluorescent protein localized to both sides of the septum and persisted in unbudded cells. Although the exact cellular role of this daughter cell-specific protein has to be elucidated, our data point to the involvement of Dse1 in bud site selection in haploid cells.
The Stationary-Phase Sigma Factor σ
S
Is Responsible for the Resistance of
Escherichia coli
Stationary-Phase Cells to
mazEF
-Mediated Cell Death
Authors: Kolodkin-Gal Ilana , Engelberg-Kulka Hanna 
Published: February 2009

ABSTRACT

Escherichia coli mazEF
is a toxin-antitoxin gene module that mediates cell death during exponential-phase cellular growth through either reactive oxygen species (ROS)-dependent or ROS-independent pathways. Here, we found that the stationary-phase sigma factor σ
S
was responsible for the resistance to
mazEF
-mediated cell death during stationary growth phase. Deletion of
rpoS
, the gene encoding σ
S
from the bacterial chromosome, permitted
mazEF
-mediated cell death during stationary growth phase.
The Putative Coupling Protein TcpA Interacts with Other pCW3-Encoded Proteins To Form an Essential Part of the Conjugation Complex
Authors: Steen Jennifer A. , Bannam Trudi L. , Teng Wee Lin , Devenish Rodney J. , Rood Julian I. 
Published: February 2009

ABSTRACT

Conjugative plasmids encode antibiotic resistance determinants or toxin genes in the anaerobic pathogen
Clostridium perfringens
. The paradigm conjugative plasmid in this bacterium is pCW3, a 47-kb tetracycline resistance plasmid that encodes the unique
tcp
transfer locus. The
tcp
locus consists of 11 genes,
intP
and
tcpA
-
tcpJ
, at least three of which,
tcpA
,
tcpF
, and
tcpH
, are essential for the conjugative transfer of pCW3. In this study we examined protein-protein interactions involving TcpA, the putative coupling protein. Use of a bacterial two-hybrid system identified interactions between TcpA and TcpC, TcpG, and TcpH. This analysis also demonstrated TcpA, TcpC, and TcpG self-interactions, which were confirmed by chemical cross-linking studies. Examination of a series of deletion and site-directed derivatives of TcpA identified the domains and motifs required for these interactions. Based on these results, we have constructed a model for this unique conjugative transfer apparatus.
Activation and Repression at the
Escherichia coli ynfEFGHI
Operon Promoter
Authors: Xu Meng , Busby Stephen J. W. , Browning Douglas F. 
Published: February 2009

ABSTRACT

Induction of the
Escherichia coli
K-12
ynfEFGHI
operon in response to anaerobiosis is repressed by nitrate ions. In this study, we show that the global transcription factor FNR is a class II activator at the
ynfEFGHI
promoter and that NarL represses activation by binding to a single target that overlaps the promoter −10 element. Electromobility shift assays show that NarL does not prevent RNA polymerase binding and suggest that repression may involve a quaternary NarL-FNR-RNA polymerase-promoter complex.
Regulation of
Bacillus subtilis aprE
Expression by
glnA
through Inhibition of
scoC
and σ
D
-Dependent
degR
Expression
Authors: Abe Sadanobu , Yasumura Ayako , Tanaka Teruo 
Published: February 2009

ABSTRACT

Expression of the gene for the extracellular alkaline protease (
aprE
) of
Bacillus subtilis
is subject to regulation by many positive and negative regulators. We have found that
aprE
expression was increased by disruption of the glutamine synthetase gene
glnA
. The increase in
aprE
expression was attributed to a decreased in expression of
scoC
, which encodes a negative regulator of
aprE
expression. The
glnA
effect on
scoC
expression was abolished by further disruption of
tnrA
, indicating that
aprE
expression is under global regulation through TnrA. In the
scoC
background, however,
aprE
expression was decreased by
glnA
deletion, and it was shown that the decrease was due to a defect in positive regulation by DegU. Among the genes that affect
aprE
expression through DegU, the expression of
degR
, encoding a protein that stabilizes phosphorylated DegU, was inhibited by
glnA
deletion. It was further shown that the decrease in
degR
expression by
glnA
deletion was caused by inhibition of the expression of
sigD
, encoding the σ
D
factor, which is required for
degR
expression. In accordance with these findings, the expression levels of
aprE-lacZ
in
glnA scoC degR
and
scoC degR
strains were identical. These results led us to conclude that
glnA
deletion brings about two effects on
aprE
expression, i.e., a positive effect through inhibition of
scoC
expression and a negative effect through inhibition of
degR
expression, with the former predominating over the latter.