bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sequencing and Transcriptional Analysis of the Biosynthesis Gene Cluster of Putrescine-Producing Lactococcus lactis
Authors: Ladero Victor , Rattray Fergal P. , Mayo Baltasar , Martín María Cruz , Fernández María , Alvarez Miguel A. 
Published: July 2011

ABSTRACT

Lactococcus lactis
is a prokaryotic microorganism with great importance as a culture starter and has become the model species among the lactic acid bacteria. The long and safe history of use of
L. lactis
in dairy fermentations has resulted in the classification of this species as GRAS (General Regarded As Safe) or QPS (Qualified Presumption of Safety). However, our group has identified several strains of
L. lactis
subsp.
lactis
and
L. lactis
subsp.
cremoris
that are able to produce putrescine from agmatine via the agmatine deiminase (AGDI) pathway. Putrescine is a biogenic amine that confers undesirable flavor characteristics and may even have toxic effects. The AGDI cluster of
L. lactis
is composed of a putative regulatory gene,
aguR
, followed by the genes (
aguB
,
aguD
,
aguA
, and
aguC
) encoding the catabolic enzymes. These genes are transcribed as an operon that is induced in the presence of agmatine. In some strains, an insertion (IS) element interrupts the transcription of the cluster, which results in a non-putrescine-producing phenotype. Based on this knowledge, a PCR-based test was developed in order to differentiate nonproducing
L. lactis
strains from those with a functional AGDI cluster. The analysis of the AGDI cluster and their flanking regions revealed that the capacity to produce putrescine via the AGDI pathway could be a specific characteristic that was lost during the adaptation to the milk environment by a process of reductive genome evolution.
Monitoring of Microbial Metabolites and Bacterial Diversity in Beef Stored under Different Packaging Conditions
Authors: Ercolini Danilo , Ferrocino Ilario , Nasi Antonella , Ndagijimana Maurice , Vernocchi Pamela , La Storia Antonietta , Laghi Luca , Mauriello Gianluigi , Guerzoni M. Elisabetta , Villani Francesco 
Published: July 2011

ABSTRACT

Beef chops were stored at 4°C under different conditions: in air (A), modified-atmosphere packaging (MAP), vacuum packaging (V), or bacteriocin-activated antimicrobial packaging (AV). After 0 to 45 days of storage, analyses were performed to determine loads of spoilage microorganisms, microbial metabolites (by solid-phase microextraction [SPME]-gas chromatography [GC]-mass spectrometry [MS] and proton nuclear magnetic resonance [
1
H NMR]), and microbial diversity (by PCR–denaturing gradient gel electrophoresis [DGGE] and pyrosequencing). The microbiological shelf life of meat increased with increasing selectivity of storage conditions. Culture-independent analysis by pyrosequencing of DNA extracted directly from meat showed that
Brochothrix thermosphacta
dominated during the early stages of storage in A and MAP, while
Pseudomonas
spp. took over during further storage in A. Many different bacteria, several of which are usually associated with soil rather than meat, were identified in V and AV; however, lactic acid bacteria (LAB) dominated during the late phases of storage, and
Carnobacterium divergens
was the most frequent microorganism in AV. Among the volatile metabolites, butanoic acid was associated with the growth of LAB under V and AV storage conditions, while acetoin was related to the other spoilage microbial groups and storage conditions.
1
H NMR analysis showed that storage in air was associated with decreases in lactate, glycogen, IMP, and ADP levels and with selective increases in levels of 3-methylindole, betaine, creatine, and other amino acids. The meat microbiota is significantly affected by storage conditions, and its changes during storage determine complex shifts in the metabolites produced, with a potential impact on meat quality.
Development of a Method To Produce Hemoglobin in a Bioreactor Culture of Escherichia coli BL21(DE3) Transformed with a Plasmid Containing Plesiomonas shigelloides Heme Transport Genes and Modified Human Hemoglobin Genes
Authors: Smith B. J. Z. , Gutierrez P. , Guerrero E. , Brewer C. J. , Henderson D. P. 
Published: July 2011

ABSTRACT

We describe a method for production of recombinant human hemoglobin by
Escherichia coli
grown in a bioreactor.
E. coli
BL21(DE3) transformed with a plasmid containing hemoglobin genes and
Plesiomonas shigelloides
heme transport genes reached a cell dry weight of 83.64 g/liter and produced 11.92 g/liter of hemoglobin in clarified lysates.
Inactivation of Escherichia coli O157:H7 on Radish Seeds by Sequential Treatments with Chlorine Dioxide, Drying, and Dry Heat without Loss of Seed Viability
Authors: Bang Jihyun , Kim Haeyoung , Kim Hoikyung , Beuchat Larry R. , Ryu Jee-Hoon 
Published: July 2011

ABSTRACT

We developed and validated a treatment to inactivate
Escherichia coli
O157:H7 on radish seeds without decreasing seed viability. Treatments with aqueous ClO
2
followed by drying and dry-heat treatments were evaluated for efficacy to inactivate the pathogen. Conditions to dry radish seeds after treatment with water (control) or ClO
2
were established. When treated seeds with high water activity (a
w
) (>0.99) were stored at 45°C and 23% relative humidity (RH), the a
w
decreased to <0.30 within 24 h. Drying high-a
w
seeds before exposing them to dry-heat treatment (≥60°C) was essential to preserve seed viability. The germination rate of radish seeds which had been immersed in water for 5 min, dried at 45°C and 23% RH for 24 h, and heated at 70°C for 48 h or at 80°C for 24 h was not significantly decreased (
P
≤ 0.05) compared to that of untreated radish seeds. Sequential treatments with ClO
2
(500 μg/ml, 5 min), drying (45°C, 23% RH, 24 h), and dry heating (70°C, 23% RH, 48 h) eliminated
E. coli
O157:H7 (5.9 log CFU/g) on radish seeds and, consequently, sprouts produced from them without decreasing the germination rate. These sequential treatments are recommended for application to radish seeds intended for sprout production.
Necessity of OxyR for the Hydrogen Peroxide Stress Response and Full Virulence in Ralstonia solanacearum
Authors: Flores-Cruz Zomary , Allen Caitilyn 
Published: July 2011

ABSTRACT

The plant pathogen
Ralstonia solanacearum
, which causes bacterial wilt disease, is exposed to reactive oxygen species (ROS) during tomato infection and expresses diverse oxidative stress response (OSR) genes during midstage disease on tomato. The
R. solanacearum
genome predicts that the bacterium produces multiple and redundant ROS-scavenging enzymes but only one known oxidative stress response regulator, OxyR. An
R. solanacearum
oxyR
mutant had no detectable catalase activity, did not grow in the presence of 250 μM hydrogen peroxide, and grew poorly in the oxidative environment of solid rich media. This phenotype was rescued by the addition of exogenous catalase, suggesting that
oxyR
is essential for the hydrogen peroxide stress response. Unexpectedly, the
oxyR
mutant strain grew better than the wild type in the presence of the superoxide generator paraquat. Gene expression studies indicated that
katE
,
kaG
,
ahpC1
,
grxC
, and
oxyR
itself were each differentially expressed in the
oxyR
mutant background and in response to hydrogen peroxide, suggesting that
oxyR
is necessary for hydrogen peroxide-inducible gene expression. Additional OSR genes were differentially regulated in response to hydrogen peroxide alone. The virulence of the
oxyR
mutant strain was significantly reduced in both tomato and tobacco host plants, demonstrating that
R. solanacearum
is exposed to inhibitory concentrations of ROS
in planta
and that OxyR-mediated responses to ROS during plant pathogenesis are important for
R. solanacearum
host adaptation and virulence.
Recombinant Protein Production by
In Vivo
Polymer Inclusion Display
Authors: Grage Katrin , Peters Verena , Rehm Bernd H. A. 
Published: July 2011

ABSTRACT
A novel approach to produce purified recombinant proteins was established. The target protein is produced as polyhydroxyalkanoate (PHA) synthase fusion protein, which mediates intracellular formation of PHA inclusions displaying the target protein. After isolation of the PHA inclusions, the pure target protein was released by simple enterokinase digestion.
The Chlamydomonas reinhardtii Molybdenum Cofactor Enzyme crARC Has a Zn-Dependent Activity and Protein Partners Similar to Those of Its Human Homologue
Authors: Chamizo-Ampudia Alejandro , Galvan Aurora , Fernandez Emilio , Llamas Angel 
Published: July 2011

ABSTRACT

The ARC (amidoxime reducing component) proteins are molybdenum cofactor (Moco) enzymes named hmARC1 and hmARC2 (human ARCs [hmARCs]) in humans and YcbX in
Escherichia coli.
They catalyze the reduction of a broad range of N-hydroxylated compounds (NHC) using reducing power supplied by other proteins. Some NHC are prodrugs or toxic compounds. YcbX contains a ferredoxin (Fd) domain and requires the NADPH flavin reductase CysJ to reduce NHC. In contrast, hmARCs lack the Fd domain and require a human cytochrome
b5
(hCyt
b5
) and a human NADH Cyt
b5
reductase (hCyt
b5-
R) to reduce NHC. The ARC proteins in the plant kingdom are uncharacterized. We demonstrate that
Chlamydomonas reinhardtii
mutants defective in Moco biosynthesis genes are sensitive to the NHC
N
6
-hydroxylaminopurine (HAP). The
Chlamydomonas reinhardtii
ARC protein crARC has been purified and characterized. The six
Chlamydomonas
Fds were isolated, but none of them are required by crARC to reduce HAP. We have also purified and characterized five
C. reinhardtii
Cyt
b5
(crCyt
b5
) and two flavin reductases, one that is NADPH dependent (crCysJ) and one that is NADH dependent (crCyt
b5
-R). The data show that crARC uses crCyt
b5
-
1
and crCyt
b5
-R to reduce HAP. The crARC has a Zn-dependent activity, and the presence of Zn increases its
V
max
more than 14-fold. In addition, all five cysteines of crARC were substituted by alanine, and we demonstrate that the fully conserved cysteine 252 is essential for both Moco binding and catalysis. Therefore, it is proposed that crARC belongs to the sulfite oxidase family of Moco enzymes.
CCR4-Associated Factor 1 Coordinates the Expression of Plasmodium falciparum Egress and Invasion Proteins
Authors: Balu Bharath , Maher Steven P. , Pance Alena , Chauhan Chitra , Naumov Anatoli V. , Andrews Robert M. , Ellis Peter D. , Khan Shahid M. , Lin Jing-wen , Janse Chris J. , Rayner Julian C. , Adams John H. 
Published: July 2011

ABSTRACT

Coordinated regulation of gene expression is a hallmark of the
Plasmodium falciparum
asexual blood-stage development cycle. We report that carbon catabolite repressor protein 4 (CCR4)-associated factor 1 (CAF1) is critical in regulating more than 1,000 genes during malaria parasites' intraerythrocytic stages, especially egress and invasion proteins. CAF1 knockout results in mistimed expression, aberrant accumulation and localization of proteins involved in parasite egress, and invasion of new host cells, leading to premature release of predominantly half-finished merozoites, drastically reducing the intraerythrocytic growth rate of the parasite. This study demonstrates that CAF1 of the CCR4-Not complex is a significant gene regulatory mechanism needed for
Plasmodium
development within the human host.
RNA-Mediated Silencing in Algae: Biological Roles and Tools for Analysis of Gene Function
Authors: Cerutti Heriberto , Ma Xinrong , Msanne Joseph , Repas Timothy 
Published: July 2011

ABSTRACT
Algae are a large group of aquatic, typically photosynthetic, eukaryotes that include species from very diverse phylogenetic lineages, from those similar to land plants to those related to protist parasites. The recent sequencing of several algal genomes has provided insights into the great complexity of these organisms. Genomic information has also emphasized our lack of knowledge of the functions of many predicted genes, as well as the gene regulatory mechanisms in algae. Core components of the machinery for RNA-mediated silencing show widespread distribution among algal lineages, but they also seem to have been lost entirely from several species with relatively small nuclear genomes. Complex sets of endogenous small RNAs, including candidate microRNAs and small interfering RNAs, have now been identified by high-throughput sequencing in green, red, and brown algae. However, the natural roles of RNA-mediated silencing in algal biology remain poorly understood. Limited evidence suggests that small RNAs may function, in different algae, in defense mechanisms against transposon mobilization, in responses to nutrient deprivation and, possibly, in the regulation of recently evolved developmental processes. From a practical perspective, RNA interference (RNAi) is becoming a promising tool for assessing gene function by sequence-specific knockdown. Transient gene silencing, triggered with exogenously synthesized nucleic acids, and/or stable gene repression, involving genome-integrated transgenes, have been achieved in green algae, diatoms, yellow-green algae, and euglenoids. The development of RNAi technology in conjunction with system level “omics” approaches may provide the tools needed to advance our understanding of algal physiological and metabolic processes.
The Iron-Responsive Regulator Irr Is Required for Wild-Type Expression of the Gene Encoding the Heme Transporter BhuA in Brucella abortus 2308
Authors: Anderson Eric S. , Paulley James T. , Martinson David A. , Gaines Jennifer M. , Steele Kendra H. , Roop R. Martin 
Published: July 2011

ABSTRACT

Irr and RirA, rather than Fur, serve as the major iron-responsive regulators in the alphaproteobacteria. With only a few exceptions, however, the relative contributions of these transcriptional regulators to the differential expression of specific iron metabolism genes in
Brucella
strains are unclear. The gene encoding the outer membrane heme transporter BhuA exhibits maximum expression in
Brucella abortus
2308 during growth under iron-deprived conditions, and mutational studies indicate that this pattern of
bhuA
expression is mediated by the iron-responsive regulator Irr. Specifically, a
bhuA-lacZ
transcriptional fusion does not produce elevated levels of β-galactosidase in response to iron deprivation in the isogenic
irr
mutant BEA5, and, unlike the parental strain,
B. abortus
BEA5 cannot utilize heme as an iron source
in vitro
and is attenuated in mice. A derivative of the
bhuA-lacZ
transcriptional fusion lacking the predicted Irr binding site upstream of the
bhuA
promoter does not produce elevated levels of β-galactosidase in response to iron deprivation in the parental
B. abortus
2308 strain, and a direct and specific interaction between a recombinant version of the
Brucella
Irr and the
bhuA
promoter region was observed in an electrophoretic mobility shift assay. Despite the fact that it lacks the heme regulatory element linked to the iron-responsive degradation of its counterpart in
Bradyrhizobium japonicum
, readily detectable levels of Irr were found only in
B. abortus
2308 cells by Western blot analysis following growth under iron-deprived conditions.
Protection and Attachment of Vibrio cholerae Mediated by the Toxin-Coregulated Pilus in the Infant Mouse Model
Authors: Krebs Shelly J. , Taylor Ronald K. 
Published: July 2011

ABSTRACT

Colonization of the human small intestine by
Vibrio cholerae
is an essential step in pathogenesis that requires the type IV toxin-coregulated pilus (TCP). To date, three functions of TCP have been characterized: it serves as the CTXΦ receptor, secretes the colonization factor TcpF, and functions in microcolony formation by mediating bacterium-bacterium interactions. Although type IV pili in other pathogenic bacteria have been characterized as playing a major role in attachment to epithelial cells, there are very few studies to suggest that TCP acts as an attachment factor. Taking this into consideration, we investigated the function of TCP in attachment to Caco-2 cells and found that mutants lacking TCP were defective in attachment compared to the wild type. Overexpression of ToxT, the activator of TCP, significantly increased attachment of wild-type
V. cholerae
to Caco-2 cells. Using field-emission scanning electron microscopy (FESEM), we also observed TCP-mediated attachment to the small intestines of infected infant mice by using antibodies specific to TCP and
V. cholerae
. Remarkably, we also visualized matrices comprised of TCP appearing to engulf
V. cholerae
during infection, and we demonstrated that these matrices protected the bacteria from a component of bile, disclosing a possible new role of this pilus in protection of the bacterial cells from antimicrobial agents. This study provides new insights into TCP's function in
V. cholerae
colonization of the small intestine, describing additional roles in mediating attachment and protection of
V. cholerae
bacterial cells.
O-Linked Glycosylation of the PilA Pilin Protein of Francisella tularensis: Identification of the Endogenous Protein-Targeting Oligosaccharyltransferase and Characterization of the Native Oligosaccharide
Authors: Egge-Jacobsen Wolfgang , Salomonsson Emelie Näslund , Aas Finn Erik , Forslund Anna-Lena , Winther-Larsen Hanne C. , Maier Josef , Macellaro Anna , Kuoppa Kerstin , Oyston Petra C. F. , Titball Richard W. , Thomas Rebecca M. , Forsberg Åke , Prior Joann L. , Koomey Michael 
Published: July 2011

ABSTRACTFindings from a number of studies suggest that the PilA pilin proteins may play an important role in the pathogenesis of disease caused by species within the genusFrancisella. As such, a thorough understanding of PilA structure and chemistry is warranted. Here, we definitively identified the PglA protein-targeting oligosaccharyltransferase by virtue of its necessity for PilA glycosylation inFrancisella tularensisand its sufficiency for PilA glycosylation inEscherichia coli. In addition, we used mass spectrometry to examine PilA affinity purified fromFrancisella tularensissubsp.tularensisandF. tularensissubsp.holarcticaand demonstrated that the protein undergoes multisite,O-linked glycosylation with a pentasaccharide of the structure HexNac-Hex-Hex-HexNac-HexNac. Further analyses revealed microheterogeneity related to forms of the pentasaccharide carrying unusual moieties linked to the distal sugar via a phosphate bridge. Type A and type B strains ofFrancisellasubspecies thus express anO-linked protein glycosylation system utilizing core biosynthetic and assembly pathways conserved in other members of the proteobacteria. As PglA appears to be highly conserved inFrancisellaspecies,O-linked protein glycosylation may be a feature common to members of this genus.
Elucidation of Structural and Antigenic Properties of Pneumococcal Serotype 11A, 11B, 11C, and 11F Polysaccharide Capsules
Authors: Calix Juan J. , Nahm Moon H. , Zartler Edward R. 
Published: July 2011

ABSTRACT

Despite the emerging impact of serogroup 11 serotypes in
Streptococcus pneumoniae
epidemiology, the structures of serogroup 11 capsule types have not been fully elucidated, particularly the locations of
O
-acetyl substitutions. Here, we report the complete structures of the serotype 11B, 11C, and 11F polysaccharides and a revision to the serotype 11A capsular polysaccharide using nuclear magnetic resonance (NMR). All structures shared a linear, tetrasaccharide backbone with a pendant phosphopolyalcohol. Three of four saccharides are conserved in all serotypes. The individual serotype capsules differed in the identity of one saccharide, the pendant phosphopolyalcohol, and the O-acetylation pattern. Though the assigned locations of
O
-acetate substitutions in this study differed from those of previous reports, our findings were corroborated with strong correlations to serology and genetics. We examined the binding of serotyping sera to serogroup 11 polysaccharides by using flow cytometry and an inhibition-type enzyme-linked immunosorbent assay (ELISA) and found that de-O-acetylation of capsular polysaccharides by mild hydrolysis decreases its immunoreactivity, supporting the crucial role of O-acetylation in the antigenicity of these polysaccharides. Due to strong correlations between polysaccharide structures and capsule biosynthesis genes, we were able to assign target substrates for the
O
-acetyltransferases encoded by
wcwC
,
wcwR
,
wcwT
, and
wcjE
. We identified antigenic determinants for serogroup 11 serotyping sera and highlight the idea that conventional serotyping methods are not capable of recognizing all putative variants of
S. pneumoniae
serogroup 11.
Peptide-Regulated Gene Depletion System Developed for Use in Streptococcus pneumoniae
Authors: Berg Kari Helene , Biørnstad Truls Johan , Straume Daniel , Håvarstein Leiv Sigve 
Published: July 2011

ABSTRACTTo facilitate the study of pneumococcal genes that are essential for viability or normal cell growth, we sought to develop a tightly regulated, titratable gene depletion system that interferes minimally with normal cellular functions. A possible candidate for such a system is the recently discovered signal transduction pathway regulating competence for natural transformation inStreptococcus thermophilus. This pathway, which is unrelated to the ComCDE pathway used for competence regulation inStreptococcus pneumoniae, has not been fully elucidated, but it is known to include a short unmodified signaling peptide, ComS*, an oligopeptide transport system, Ami, and a transcriptional activator, ComR. The transcriptional activator is thought to bind to an inverted repeat sequence termed the ECom box. We introduced the ComR protein and the ECom box into the genome ofS. pneumoniaeR6 and demonstrated that addition of synthetic ComS* peptide induced the transcription of a luciferase gene inserted downstream of the ECom box. To determine whether the ComRS system could be used for gene depletion studies, thelicD1gene was inserted behind the chromosomally located ECom box promoter by using the Janus cassette. Then, the native versions oflicD1andlicD2were deleted, and the resulting mutant was recovered in the presence of ComS*. Cultivation of thelicD1 licD2double mutant in the absence of ComS* gradually affected its ability to grow and propagate, demonstrating that the ComRS system functions as intended. In the present study, the ComRS system was developed for use inS. pneumoniae. In principle, however, it should work equally well in many other Gram-positive species.