bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization of Three
mnp
Genes of
Fomitiporia mediterranea
and Report of Additional Class II Peroxidases in the Order Hymenochaetales
Authors: Morgenstern Ingo , Robertson Deborah L. , Hibbett David S. 
Published: July 2010

ABSTRACT

We report the sequence-based characterization and expression patterns of three manganese peroxidase genes from the white rot fungus and grape vine pathogen
Fomitiporia mediterranea
(Agaricomycotina, Hymenochaetales), termed
Fmmnp1
,
Fmmnp2
, and
Fmmnp3
. The predicted open reading frames (ORFs) are 1,516-, 1,351-, and 1,345-bp long and are interrupted by seven, four, and four introns, respectively. The deduced amino acid sequences encode manganese peroxidases (EC 1.11.1.13) containing 371, 369, and 371 residues, respectively, and are similar to the manganese peroxidases of the model white rot organism
Phanerochaete chrysosporium
. The expression of the genes is most likely differentially regulated, as revealed by real-time PCR analysis. Phylogenetic analysis reveals that other members of the order Hymenochaetales harbor
mnp
genes encoding proteins that are related only distantly to those of
F. mediterranea
. Furthermore, multiple partial
lip
- and
mnp
-like sequences obtained for
Pycnoporus cinnabarinus
(Agaricomycotina, Polyporales) suggest that lignin degradation by white rot taxa relies heavily on ligninolytic peroxidases and is not efficiently achieved by laccases only.
Construction of Self-Transmissible Green Fluorescent Protein-Based Biosensor Plasmids and Their Use for Identification ofN-Acyl Homoserine-Producing Bacteria in Lake Sediments
Authors: Lumjiaktase Putthapoom , Aguilar Claudio , Battin Tom , Riedel Kathrin , Eberl Leo 
Published: July 2010

ABSTRACTMany bacteria utilize quorum sensing (QS) systems to communicate with each other by means of the production, release, and response to signal molecules.N-Acyl homoserine lactone (AHL)-based QS systems are particularly widespread among theProteobacteria, in which they regulate various functions. It has become evident that AHLs can also serve as signals for interspecies communication. However, knowledge on the impact of AHLs for the ecology of bacteria in their natural habitat is scarce, due mainly to the lack of tools that allow the study of QS in bacterial communitiesin situ. Here, we describe the construction of self-mobilizable green fluorescent protein (GFP)-based AHL sensors that utilize the conjugation and replication properties of the broad-host-range plasmid RP4. We show that these novel AHL sensor plasmids can be easily transferred to different bacterial species by biparental mating and that they give rise to green fluorescent cells in case the recipient is an AHL producer. We also demonstrate that these sensor plasmids are capable of self-spreading within mixed biofilms and are a suitable tool for the identification of AHL-producing bacteria in lake sediment.
Phenotypic Characterization of
Shewanella oneidensis
MR-1 under Aerobic and Anaerobic Growth Conditions by Using Fourier Transform Infrared Spectroscopy and High-Performance Liquid Chromatography Analyses
Authors: Wang Hui , Hollywood Katherine , Jarvis Roger M. , Lloyd Jonathan R. , Goodacre Royston 
Published: July 2010

ABSTRACT

Shewanella oneidensis
is able to conserve energy for growth by reducing a wide variety of terminal electron acceptors during anaerobic respiration, including several environmentally hazardous pollutants. This bacterium employs various electron transfer mechanisms for anaerobic respiration, including cell-bound reductases and secreted redox mediators. The aim of this study was to develop rapid tools for profiling the key metabolic changes associated with these different growth regimes and physiological responses. Initial experiments focused on comparing cells grown under aerobic and anaerobic conditions. Fourier transform infrared (FT-IR) spectroscopy with cluster analysis showed that there were significant changes in the metabolic fingerprints of the cells grown under these two culture conditions. FT-IR spectroscopy clearly differentiated cells of
S. oneidensis
MR-1 cultured at various growth points and cells grown using different electron acceptors, resulting in different phenotypic trajectories in the cluster analysis. This growth-related trajectory analysis is applied successfully for the first time, here with FT-IR spectroscopy, to investigate the phenotypic changes in contrasting
S. oneidensis
cells. High-performance liquid chromatography (HPLC) was also used to quantify the concentrations of flavin compounds, which have been identified recently as extracellular redox mediators released by a range of
Shewanella
species. The partial least-squares regression (PLSR) multivariate statistical technique was combined with FT-IR spectroscopy to predict the concentrations of the flavins secreted by cells of
S. oneidensis
MR-1, suggesting that this combination could be used as a rapid alternative to conventional chromatographic methods for analysis of flavins in cell cultures. Furthermore, coupling of the FT-IR spectroscopy and HPLC techniques appears to offer a potentially useful tool for rapid characterization of the
Shewanella
cell metabolome in various process environments.
Adaptation of Methanogenic Communities to the Cofermentation of Cattle Excreta and Olive Mill Wastes at 37°C and 55°C
Authors: Goberna Marta , Gadermaier Maria , García Carlos , Wett Bernhard , Insam Heribert 
Published: July 2010

ABSTRACT

The acclimatization of methanogens to two-phase olive mill wastes (TPOMW) was investigated in pilot fermenters started up with cattle excreta (37°C) and after changing their feed to excreta plus TPOMW (37°C or 55°C) or TPOMW alone (37°C) until a steady state was reached (28 days). Methanogenic diversity was screened using a phylogenetic microarray (AnaeroChip), and positive targets were quantified by real-time PCR. Results revealed high phylogenetic richness, with representatives of three out of the four taxonomic orders found in digesters.
Methanosarcina
dominated in the starting excreta (>96% of total 16S rRNA gene copies; over 45 times more abundant than any other methanogen) at high acetate (0.21 g liter
−1
) and ammonia N concentrations (1.3 g liter
−1
). Codigestion at 37°C induced a 6-fold increase of
Methanosarcina
numbers, correlated with CH
4
production (
r
Pearson
= 0.94;
P
= 0.02). At 55°C, the rise in temperature and H
2
partial pressure induced a burst of
Methanobacterium
,
Methanoculleus
,
Methanothermobacter
, and a group of uncultured archaea. The digestion of excreta alone resulted in low but constant biogas production despite certain oscillations in the methanogenic biomass. Unsuccessful digestion of TPOMW alone was attributed to high Cu levels inducing inhibition of methanogenic activity. In conclusion, the versatile
Methanosarcina
immediately adapted to the shift from excreta to excreta plus TPOMW and was responsible for the stimulated CH
4
production at 37°C. Higher temperatures (55°C) fostered methanogenic diversity by promoting some H
2
scavengers while yielding the highest CH
4
production. Further testing is needed to find out whether there is a link between increased methanogenic diversity and reactor productivity.
Restoration of Growth Phenotypes of
Escherichia coli
DH5α in Minimal Media through Reversal of a Point Mutation in
purB
Authors: Jung Suk-Chae , Smith Chris L. , Lee Ki-Sung , Hong Min-Eui , Kweon Dae-Hyuk , Stephanopoulos Gregory , Jin Yong-Su 
Published: July 2010

ABSTRACT

A point mutation (E115K) resulting in slower growth of
E
scherichia
coli
DH5α and XL1-Blue in minimal media was identified in the
purB
gene, coding for adenylosuccinate lyase (ASL), through complementation with an
E. coli
K-12 genomic library and serial subcultures. Chromosomal modification reversing the mutation to the wild type restored growth phenotypes in minimal media.
Functional Characterization of Three Leishmania Poly(A) Binding Protein Homologues with Distinct Binding Properties to RNA and Protein Partners
Authors: da Costa Lima Tamara D. , Moura Danielle M. N. , Reis Christian R. S. , Vasconcelos J. Ronnie C. , Ellis Louise , Carrington Mark , Figueiredo Regina C. B. Q. , de Melo Neto Osvaldo P. 
Published: July 2010

ABSTRACT

Trypanosomatid protozoans are reliant on posttranscriptional processes to control gene expression. Regulation occurs at the levels of mRNA processing, stability, and translation, events that may require the participation of the poly(A) binding protein (PABP). Here, we have undertaken a functional study of the three distinct
Leishmania major
PABP (
Lm
PABP) homologues: the previously described
Lm
PABP1;
Lm
PABP2, orthologous to the PABP described from
Trypanosoma
species; and
Lm
PABP3, unique to
Leishmania
. Sequence identity between the three PABPs is no greater than 40%. In assays measuring binding to A-rich sequences,
Lm
PABP1 binding was poly(A) sensitive but heparin insensitive;
Lm
PABP2 binding was heparin sensitive and less sensitive to poly(A), compatible with unique substitutions observed in residues implicated in poly(A) binding; and
Lm
PABP3 displayed intermediate properties. All three homologues are simultaneously expressed as abundant cytoplasmic proteins in
L. major
promastigotes, but only
Lm
PABP1 is present as multiple isoforms. Upon transcription inhibition,
Lm
PABP2 and -3 migrated to the nucleus, while
Lm
PABP1 remained predominantly cytoplasmic. Immunoprecipitation assays showed an association between
Lm
PABP2 and -3. Although the three proteins bound to a
Leishmania
homologue of the translation initiation factor eukaryotic initiation factor 4G (eIF4G) (
Lm
EIF4G3)
in vitro
,
Lm
PABP1 was the only one to copurify with native
Lm
EIF4G3 from cytoplasmic extracts. Functionality was tested using RNA interference (RNAi) in
Trypanosoma brucei
, where both orthologues to
Lm
PABP1 and -2 are required for cellular viability. Our results indicate that these homologues have evolved divergent functions, some of which may be unique to the trypanosomatids, and reinforces a role for
Lm
PABP1 in translation through its interaction with the eIF4G homologue.
Transcription Factors in Light and Circadian Clock Signaling Networks Revealed by Genomewide Mapping of Direct Targets for Neurospora White Collar Complex
Authors: Smith Kristina M. , Sancar Gencer , Dekhang Rigzin , Sullivan Christopher M. , Li Shaojie , Tag Andrew G. , Sancar Cigdem , Bredeweg Erin L. , Priest Henry D. , McCormick Ryan F. , Thomas Terry L. , Carrington James C. , Stajich Jason E. , Bell-Pedersen Deborah , Brunner Michael , Freitag Michael 
Published: July 2010

ABSTRACT

Light signaling pathways and circadian clocks are inextricably linked and have profound effects on behavior in most organisms. Here, we used chromatin immunoprecipitation (ChIP) sequencing to uncover direct targets of the
Neurospora crassa
circadian regulator White Collar Complex (WCC). The WCC is a blue-light receptor and the key transcription factor of the circadian oscillator. It controls a transcriptional network that regulates ∼20% of all genes, generating daily rhythms and responses to light. We found that in response to light, WCC binds to hundreds of genomic regions, including the promoters of previously identified clock- and light-regulated genes. We show that WCC directly controls the expression of 24 transcription factor genes, including the clock-controlled
adv-1
gene, which controls a circadian output pathway required for daily rhythms in development. Our findings provide links between the key circadian activator and effectors in downstream regulatory pathways.
Identification of
GIG1
, a GlcNAc-Induced Gene in Candida albicans Needed for Normal Sensitivity to the Chitin Synthase Inhibitor Nikkomycin Z
Authors: Gunasekera Angelo , Alvarez Francisco J. , Douglas Lois M. , Wang Hong X. , Rosebrock Adam P. , Konopka James B. 
Published: July 2010

ABSTRACT

The amino sugar
N
-acetylglucosamine (GlcNAc) is known to be an important structural component of cells from bacteria to humans, but its roles in cell signaling are less well understood. GlcNAc induces two pathways in the human fungal pathogen
Candida albicans
. One activates cyclic AMP (cAMP) signaling, which stimulates the formation of hyphal cells and the expression of virulence genes, and the other pathway induces genes needed to catabolize GlcNAc. Microarray analysis of gene expression was carried out under four different conditions in order to characterize the transcriptional changes induced by GlcNAc. The most highly induced genes include those that encode a GlcNAc transporter (
NGT1
) and the GlcNAc catabolic enzymes (
HXK1
,
DAC1
, and
NAG1
). GlcNAc also activated most of the genes whose expression is increased when cells are triggered with other stimuli to form hyphae. Surprisingly, GlcNAc also induced a subset of genes that are regulated by galactose (
GAL1
,
GAL7
, and
GAL10
), which may be due to cross talk between signaling pathways. A novel GlcNAc-induced gene,
GIG1
, which is not essential for GlcNAc catabolism or the induction of hyphae, was identified. However, a Gig1-green fluorescent protein (GFP) fusion protein was specifically induced by GlcNAc, and not by other sugars. Gig1-GFP localized to the cytoplasm, where GlcNAc metabolism occurs. Significantly, a
gig1
Δ mutant displayed increased resistance to nikkomycin Z, which inhibits chitin synthase from converting UDP-GlcNAc into cell wall chitin. Gig1 is highly conserved in fungi, especially those that contain GlcNAc catabolic genes. These results implicate Gig1 in GlcNAc metabolism.
The Twin Arginine Transport System Appears To Be Essential for Viability in
Sinorhizobium meliloti
Authors: Pickering Brad S. , Oresnik Ivan J. 
Published: July 2010

ABSTRACT

The twin arginine transport (Tat) system is responsible for transporting prefolded proteins to the periplasmic space. The Tat pathway has been implicated in many bacterial cellular functions, including motility, biofilm formation, and pathogenesis and symbiosis. Since the annotation of
Sinorhizobium meliloti
Rm1021 genome suggests that there may be up to 94 putative Tat substrates, we hypothesized that characterizing the twin arginine transport system in this organism might yield unique data that could help in the understanding of twin arginine transport. To initiate this work we attempted a targeted mutagenesis of the
tat
locus. Despite repeated attempts using a number of different types of media, the attempts at mutation construction were unsuccessful unless the experiment was carried out in a strain that was merodiploid for
tatABC
. In addition, it was shown that a plasmid carrying
tatABC
was stable in the absence of antibiotic selection in a
tat
deletion background. Finally, fluorescence microscopy and live/dead assays of these cultures show a high proportion of dead and irregularly shaped cells, suggesting that the loss of
tatABC
is inversely correlated with viability. Taken together, the results of this work provide evidence that the twin arginine transport system of
S. meliloti
appears to be essential for viability under all the conditions that we had tested.
Characterization of NADH Oxidase/NADPH Polysulfide Oxidoreductase and Its Unexpected Participation in Oxygen Sensitivity in an Anaerobic Hyperthermophilic Archaeon
Authors: Kobori Hiroki , Ogino Masayuki , Orita Izumi , Nakamura Satoshi , Imanaka Tadayuki , Fukui Toshiaki 
Published: July 2010

ABSTRACT

Many genomes of anaerobic hyperthermophiles encode multiple homologs of NAD(P)H oxidase that are thought to function in response to oxidative stress. We investigated one of the seven NAD(P)H oxidase homologs (TK1481) in the sulfur-reducing hyperthermophilic archaeon
Thermococcus kodakarensis
, focusing on the catalytic properties and roles in oxidative-stress defense and sulfur-dependent energy conservation. The recombinant form of TK1481 exhibited both NAD(P)H oxidase and NAD(P)H:polysulfide oxidoreductase activities. The enzyme also possessed low NAD(P)H peroxidase and NAD(P)H:elemental sulfur oxidoreductase activities under anaerobic conditions. A mutant form of the enzyme, in which the putative redox-active residue Cys43 was replaced by Ala, still showed NADH-dependent flavin adenine dinucleotide (FAD) reduction activity. Although it also retained successive oxidase and anaerobic peroxidase activities, the ability to reduce polysulfide and sulfur was completely lost, suggesting the specific reactivity of the Cys43 residue for sulfur. To evaluate the physiological function of TK1481, we constructed a gene deletant, ΔTK1481, and mutant KUTK1481C43A, into which two base mutations altering Cys43 of TK1481 to Ala were introduced. ΔTK1481 exhibited growth properties nearly identical to those of the parent strain, KU216, in sulfur-containing media. Interestingly, in the absence of elemental sulfur, the growth of ΔTK1481 was not affected by dissolved oxygen, whereas the growth of KU216 and KUTK1481C43A was significantly impaired. These results indicate that although TK1481 does not play a critical role in either sulfur reduction or the response to oxidative stress, the NAD(P)H oxidase activity of TK1481 unexpectedly participates in the oxygen sensitivity of the hyperthermophilic archaeon
T. kodakarensis
in the absence of sulfur.
Pyruvate Is Synthesized by Two Pathways in Pea Bacteroids with Different Efficiencies for Nitrogen Fixation
Authors: Mulley Geraldine , Lopez-Gomez Miguel , Zhang Ye , Terpolilli Jason , Prell Jurgen , Finan Turlough , Poole Philip 
Published: July 2010

ABSTRACT

Nitrogen fixation in legume bacteroids is energized by the metabolism of dicarboxylic acids, which requires their oxidation to both oxaloacetate and pyruvate. In alfalfa bacteroids, production of pyruvate requires NAD
+
malic enzyme (Dme) but not NADP
+
malic enzyme (Tme). However, we show that
Rhizobium leguminosarum
has two pathways for pyruvate formation from dicarboxylates catalyzed by Dme and by the combined activities of phosphoenolpyruvate (PEP) carboxykinase (PckA) and pyruvate kinase (PykA). Both pathways enable N
2
fixation, but the PckA/PykA pathway supports N
2
fixation at only 60% of that for Dme. Double mutants of
dme
and
pckA/pykA
did not fix N
2
. Furthermore,
dme pykA
double mutants did not grow on dicarboxylates, showing that they are the only pathways for the production of pyruvate from dicarboxylates normally expressed. PckA is not expressed in alfalfa bacteroids, resulting in an obligate requirement for Dme for pyruvate formation and N
2
fixation. When PckA was expressed from a constitutive
nptII
promoter in alfalfa
dme
bacteroids, acetylene was reduced at 30% of the wild-type rate, although this level was insufficient to prevent nitrogen starvation. Dme has N-terminal, malic enzyme (Me), and C-terminal phosphotransacetylase (Pta) domains. Deleting the Pta domain increased the peak acetylene reduction rate in 4-week-old pea plants to 140 to 150% of the wild-type rate, and this was accompanied by increased nodule mass. Plants infected with Pta deletion mutants did not have increased dry weight, demonstrating that there is not a sustained change in nitrogen fixation throughout growth. This indicates a complex relationship between pyruvate synthesis in bacteroids, nitrogen fixation, and plant growth.
Unexpected Abundance of Coenzyme F
420
-Dependent Enzymes in
Mycobacterium tuberculosis
and Other Actinobacteria
Authors: Selengut Jeremy D. , Haft Daniel H. 
Published: July 2010

ABSTRACT

Regimens targeting
Mycobacterium tuberculosis
, the causative agent of tuberculosis (TB), require long courses of treatment and a combination of three or more drugs. An increase in drug-resistant strains of
M. tuberculosis
demonstrates the need for additional TB-specific drugs. A notable feature of
M. tuberculosis
is coenzyme F
420
, which is distributed sporadically and sparsely among prokaryotes. This distribution allows for comparative genomics-based investigations. Phylogenetic profiling (comparison of differential gene content) based on F
420
biosynthesis nominated many actinobacterial proteins as candidate F
420
-dependent enzymes. Three such families dominated the results: the luciferase-like monooxygenase (LLM), pyridoxamine 5′-phosphate oxidase (PPOX), and deazaflavin-dependent nitroreductase (DDN) families. The DDN family was determined to be limited to F
420
-producing species. The LLM and PPOX families were observed in F
420
-producing species as well as species lacking F
420
but were particularly numerous in many actinobacterial species, including
M. tuberculosis
. Partitioning the LLM and PPOX families based on an organism's ability to make F
420
allowed the application of the SIMBAL (sites inferred by metabolic background assertion labeling) profiling method to identify F
420
-correlated subsequences. These regions were found to correspond to flavonoid cofactor binding sites. Significantly, these results showed that
M. tuberculosis
carries at least 28 separate F
420
-dependent enzymes, most of unknown function, and a paucity of flavin mononucleotide (FMN)-dependent proteins in these families. While prevalent in mycobacteria, markers of F
420
biosynthesis appeared to be absent from the normal human gut flora. These findings suggest that
M. tuberculosis
relies heavily on coenzyme F
420
for its redox reactions. This dependence and the cofactor's rarity may make F
420
-related proteins promising drug targets.
Transcriptional Profiling of XdrA, a New Regulator ofspaTranscription inStaphylococcus aureus
Authors: McCallum N. , Hinds J. , Ender M. , Berger-Bächi B. , Stutzmann Meier P. 
Published: July 2010

ABSTRACTTranscription ofspa, encoding the virulence factor protein A inStaphylococcus aureus, is tightly controlled by a complex regulatory network, ensuring its temporal expression over growth and at appropriate stages of the infection process. Transcriptomic profiling of XdrA, a DNA-binding protein that is conserved in allS. aureusgenomes and shares similarity with the XRE family of helix-turn-helix, antitoxin-like proteins, revealed it to be a previously unidentified activator ofspatranscription. To assess how XdrA fits into the complex web ofsparegulation, a series of regulatory mutants were constructed; consisting of single, double, triple, and quadruple mutants lacking XdrA and/or the three key regulators previously shown to influencespatranscription directly (SarS, SarA, and RNAIII). A series oflacZreporter gene fusions containing nested deletions of thespapromoter identified regions influenced by XdrA and the other three regulators. XdrA had almost as strong an activating effect onspaas SarS and acted on the samespaoperator regions as SarS, or closely overlapping regions. All data from microarrays, Northern and Western blot analyses, and reporter gene fusion experiments indicated that XdrA is a major activator ofspaexpression that appears to act directly on thespapromoter and not through previously characterized regulators.