bluuebunny/crossref_metadata_2025_split huggingface dataset:
Intracellular Accumulation of High Levels of γ-Aminobutyrate by
Listeria monocytogenes
10403S in Response to Low pH: Uncoupling of γ-Aminobutyrate Synthesis from Efflux in a Chemically Defined Medium
Authors: Karatzas Kimon-Andreas G. , Brennan Orla , Heavin Sinéad , Morrissey John , O'Byrne Conor P. 
Published: April 2010

ABSTRACT

It is well established that the glutamate decarboxylase (GAD) system is central to the survival of
Listeria monocytogenes
at low pH, both in acidic foods and within the mammalian stomach. The accepted model proposes that under acidic conditions extracellular glutamate is transported into the cell in exchange for an intracellular γ-aminobutyrate (GABA
i
). The glutamate is then decarboxylated to GABA
i
, a reaction that consumes a proton, thereby helping to prevent acidification of the cytoplasm. In this study, we show that glutamate supplementation had no influence on either growth rate at pH 5.0 or survival at pH 2.5 when
L. monocytogenes
10403S was grown in a chemically defined medium (DM). In response to acidification, cells grown in DM failed to efflux GABA, even when glutamate was added to the medium. In contrast, in brain heart infusion (BHI), the same strain produced significant extracellular GABA (GABA
e
) in response to acidification. In addition, high levels of GABA
i
(>80 mM) were found in the cytoplasm in response to low pH in both growth media. Medium-swap and medium-mixing experiments revealed that the GABA efflux apparatus was nonfunctional in DM, even when glutamate was present. It was also found that the GadT2D2 antiporter/decarboxylase system was transcribed poorly in DM-grown cultures while overexpression of
gadD1T1
and
gadD3
occurred in response to pH 3.5. Interestingly, BHI-grown cells did not respond with upregulation of any of the GAD system genes when challenged at pH 3.5. The accumulation of GABA
i
in cells grown in DM in the absence of extracellular glutamate indicates that intracellular glutamate is the source of the GABA
i
. These results demonstrate that GABA production can be uncoupled from GABA efflux, a finding that alters the way we should view the operation of bacterial GAD systems.
Cellulosilyticum ruminicola
, a Newly Described Rumen Bacterium That Possesses Redundant Fibrolytic-Protein-Encoding Genes and Degrades Lignocellulose with Multiple Carbohydrate- Borne Fibrolytic Enzymes
Authors: Cai Shichun , Li Jiabao , Hu Fen Ze , Zhang Kegui , Luo Yuanming , Janto Benjamin , Boissy Robert , Ehrlich Garth , Dong Xiuzhu 
Published: April 2010

ABSTRACT

Cellulosilyticum ruminicola
H1 is a newly described bacterium isolated from yak (
Bos grunniens
) rumen and is characterized by its ability to grow on a variety of hemicelluloses and degrade cellulosic materials. In this study, we performed the whole-genome sequencing of
C. ruminicola
H1 and observed a comprehensive set of genes encoding the enzymes essential for hydrolyzing plant cell wall. The corresponding enzymatic activities were also determined in strain H1; these included endoglucanases, cellobiohydrolases, xylanases, mannanase, pectinases, and feruloyl esterases and acetyl esterases to break the interbridge cross-link, as well as the enzymes that degrade the glycosidic bonds. This bacterium appears to produce polymer hydrolases that act on both soluble and crystal celluloses. Approximately half of the cellulytic activities, including cellobiohydrolase (50%), feruloyl esterase (45%), and one third of xylanase (31%) and endoglucanase (36%) activities were bound to cellulosic fibers. However, only a minority of mannase (6.78%) and pectinase (1.76%) activities were fiber associated. Strain H1 seems to degrade the plant-derived polysaccharides by producing individual fibrolytic enzymes, whereas the majority of polysaccharide hydrolases contain carbohydrate-binding module. Cellulosome or cellulosomelike protein complex was never isolated from this bacterium. Thus, the fibrolytic enzyme production of strain H1 may represent a different strategy in cellulase organization used by most of other ruminal microbes, but it applies the fungal mode of cellulose production.
Birefringence and DNA Condensation of Liquid Crystalline Chromosomes
Authors: Chow Man H. , Yan Kosmo T. H. , Bennett Michael J. , Wong Joseph T. Y. 
Published: April 2010

ABSTRACT

DNA can self-assemble
in vitro
into several liquid crystalline phases at high concentrations. The largest known genomes are encoded by the cholesteric liquid crystalline chromosomes (LCCs) of the dinoflagellates, a diverse group of protists related to the malarial parasites. Very little is known about how the liquid crystalline packaging strategy is employed to organize these genomes, the largest among living eukaryotes—up to 80 times the size of the human genome. Comparative measurements using a semiautomatic polarizing microscope demonstrated that there is a large variation in the birefringence, an optical property of anisotropic materials, of the chromosomes from different dinoflagellate species, despite their apparently similar ultrastructural patterns of bands and arches. There is a large variation in the chromosomal arrangements in the nuclei and individual karyotypes. Our data suggest that both macroscopic and ultrastructural arrangements affect the apparent birefringence of the liquid crystalline chromosomes. Positive correlations are demonstrated for the first time between the level of absolute retardance and both the DNA content and the observed helical pitch measured from transmission electron microscopy (TEM) photomicrographs. Experiments that induced disassembly of the chromosomes revealed multiple orders of organization in the dinoflagellate chromosomes. With the low protein-to-DNA ratio, we propose that a highly regulated use of entropy-driven force must be involved in the assembly of these LCCs. Knowledge of the mechanism of packaging and arranging these largest known DNAs into different shapes and different formats in the nuclei would be of great value in the use of DNA as nanostructural material.
Isocitrate Dehydrogenase Is Important for Nitrosative Stress Resistance in Cryptococcus neoformans, but Oxidative Stress Resistance Is Not Dependent on Glucose-6-Phosphate Dehydrogenase
Authors: Brown Sarah M. , Upadhya Rajendra , Shoemaker James D. , Lodge Jennifer K. 
Published: April 2010

ABSTRACTThe opportunistic intracellular fungal pathogenCryptococcus neoformansdepends on many antioxidant and denitrosylating proteins and pathways for virulence in the immunocompromised host. These include the glutathione and thioredoxin pathways, thiol peroxidase, cytochromecperoxidase, and flavohemoglobin denitrosylase. All of these ultimately depend on NADPH for either catalytic activity or maintenance of a reduced, functional form. The need for NADPH during oxidative stress is well established in many systems, but a role in resistance to nitrosative stress has not been as well characterized. In this study we investigated the roles of two sources of NADPH, glucose-6-phosphate dehydrogenase (Zwf1) and NADP+-dependent isocitrate dehydrogenase (Idp1), in production of NADPH and resistance to oxidative and nitrosative stress. Deletion ofZWF1inC. neoformansdid not result in an oxidative stress sensitivity phenotype or changes in the amount of NADPH produced during oxidative stress compared to those for the wild type. Deletion ofIDP1resulted in greater sensitivity to nitrosative stress than to oxidative stress. The amount of NADPH increased 2-fold over that in the wild type during nitrosative stress, and yet theidp1Δ strain accumulated more mitochondrial damage than the wild type during nitrosative stress. This is the first report of the importance of Idp1 and NADPH for nitrosative stress resistance.
Naegleria gruberi
De Novo
Basal Body Assembly Occurs via Stepwise Incorporation of Conserved Proteins
Authors: Fritz-Laylin Lillian K. , Assaf Zoe June , Chen Sean , Cande W. Zacheus 
Published: April 2010

ABSTRACT

Centrioles and basal bodies are discrete structures composed of a cylinder of nine microtubule triplets and associated proteins. Metazoan centrioles can be found at mitotic spindle poles and are called basal bodies when used to organize microtubules to form the core structure of flagella.
Naegleria gruberi
, a unicellular eukaryote, grows as an amoeba that lacks a cytoplasmic microtubule cytoskeleton. When stressed,
Naegleria
rapidly (and synchronously) differentiates into a flagellate, forming a complete cytoplasmic cytoskeleton
de novo
, including two basal bodies and flagella. Here, we show that
Naegleria
has genes encoding conserved centriole proteins. Using novel antibodies, we describe the localization of three centrosomal protein homologs (SAS-6, γ-tubulin, and centrin-1) during the assembly of the flagellate microtubule cytoskeleton. We also used these antibodies to show that
Naegleria
expresses the proteins in the same order as their incorporation into basal bodies, with SAS-6 localizing first, followed by centrin and finally γ-tubulin. The similarities between basal body assembly in
Naegleria
and centriole assembly in animals indicate that mechanisms of assembly, as well as structure, have been conserved throughout eukaryotic evolution.
A Role for the Class A Penicillin-Binding Protein PonA2 in the Survival ofMycobacterium smegmatisunder Conditions of Nonreplication
Authors: Patru Maria-Magdalena , Pavelka Martin S. 
Published: April 2010

ABSTRACTClass A penicillin-binding proteins (PBPs) are large, bifunctional proteins that are responsible for glycan chain assembly and peptide cross-linking of bacterial peptidoglycan. Bacteria in the genusMycobacteriumhave been reported to have only two class A PBPs, PonA1 and PonA2, that are encoded in their genomes. We report here that the genomes ofMycobacterium smegmatisand other soil mycobacteria contain an additional gene encoding a third class A penicillin-binding protein, PonA3, which is a paralog of PonA2. Both the PonA2 and PonA3 proteins contain apenicillin-binding proteinandserine/threonine protein kinase-associated (PASTA) domain that we propose may be involved in sensing the cell cycle and a C-terminal proline-rich region (PRR) that may have a role in protein-protein or protein-carbohydrate interactions. We show here that anM. smegmatisΔponA2mutant has an unusual antibiotic susceptibility profile, exhibits a spherical morphology and an altered cell surface in stationary phase, and is defective for stationary-phase survival and recovery from anaerobic culture. In contrast, a ΔponA3mutant has no discernible phenotype under laboratory conditions. We demonstrate that PonA2 and PonA3 can bind penicillin and that PonA3 can partially substitute for PonA2 whenponA3is expressed from a constitutive promoter on a multicopy plasmid. Our studies suggest that PonA2 is involved in adaptation to periods of nonreplication in response to starvation or anaerobiosis and that PonA3 may have a similar role. However, the regulation of PonA3 is likely different, suggesting that its importance could be related to stresses encountered in the environmental niches occupied byM. smegmatisand other soil-dwelling mycobacteria.
YneA, an SOS-Induced Inhibitor of Cell Division in
Bacillus subtilis
, Is Regulated Posttranslationally and Requires the Transmembrane Region for Activity
Authors: Mo Allison H. , Burkholder William F. 
Published: April 2010

ABSTRACT

Cell viability depends on the stable transmission of genetic information to each successive generation. Therefore, in the event of intrinsic or extrinsic DNA damage, it is important that cell division be delayed until DNA repair has been completed. In
Bacillus subtilis
, this is accomplished in part by YneA, an inhibitor of division that is induced as part of the SOS response. We sought to gain insight into the mechanism by which YneA blocks cell division and the processes involved in shutting off YneA activity. Our data suggest that YneA is able to inhibit daughter cell separation as well as septum formation. YneA contains a LysM peptidoglycan binding domain and is predicted to be exported. We established that the YneA signal peptide is rapidly cleaved, resulting in secretion of YneA into the medium. Mutations within YneA affect both the rate of signal sequence cleavage and the activity of YneA. YneA does not stably associate with the cell wall and is rapidly degraded by extracellular proteases. Based on these results, we hypothesize that exported YneA is active prior to signal peptide cleavage and that proteolysis contributes to the inactivation of YneA. Finally, we identified mutations in the transmembrane segment of YneA that abolish the ability of YneA to inhibit cell division, while having little or no effect on YneA export or stability. These data suggest that protein-protein interactions mediated by the transmembrane region may be required for YneA activity.
N-Terminal Type III Secretion Signal of Enteropathogenic
Escherichia coli
Translocator Proteins
Authors: Munera Diana , Crepin Valerie F. , Marches Olivier , Frankel Gad 
Published: April 2010

ABSTRACT

We report that the N terminus of the type III secretion system translocator proteins EspB, EspD, and EspA mediate protein secretion and translocation from wild-type enteropathogenic
Escherichia coli
and hypersecretion from
sepL
and
sepD
mutants. EspA containing the translocation signal of Map and Tir containing the secretion signal of EspA are biologically active.
Host-Interactive Genes in AmerindianHelicobacter pyloriDiverge from Their Old World Homologs and Mediate Inflammatory Responses
Authors: Mane S. P. , Dominguez-Bello M. G. , Blaser M. J. , Sobral B. W. , Hontecillas R. , Skoneczka J. , Mohapatra S. K. , Crasta O. R. , Evans C. , Modise T. , Shallom S. , Shukla M. , Varon C. , Mégraud F. , Maldonado-Contreras A. L. , Williams K. P. , Bassaganya-Riera J. 
Published: April 2010

ABSTRACTHelicobacter pyloriis the dominant member of the gastric microbiota and has been associated with an increased risk of gastric cancer and peptic ulcers in adults.H. pyloripopulations have migrated and diverged with human populations, and health effects vary. Here, we describe the whole genome of thecag-positive strain V225d, cultured from a Venezuelan Piaroa Amerindian subject. To gain insight into the evolution and host adaptation of this bacterium, we undertook comparativeH. pylorigenomic analyses. A robust multiprotein phylogenetic tree reflects the major human migration out of Africa, across Europe, through Asia, and into the New World, placing AmerindianH. pylorias a particularly close sister group to East AsianH. pylori. In contrast, phylogenetic analysis of the host-interactive genesvacAandcagAshows substantial divergence of Amerindian from Old World forms and indicates new genotypes (e.g., VacA m3) involving these loci. Despite deletions in CagA EPIYA and CRPIA domains, V225d stimulates interleukin-8 secretion and the hummingbird phenotype in AGS cells. However, following a 33-week passage in the mouse stomach, these phenotypes were lost in isolate V225-RE, which had a 15-kb deletion in thecagpathogenicity island that truncated CagA and eliminated some of the type IV secretion system genes. Thus, the unusual V225dcagarchitecture was fully functional via conserved elements, but the natural deletion of 13cagpathogenicity island genes and the truncation of CagA impaired the ability to induce inflammation.
Substrate-Level Phosphorylation Is the Primary Source of Energy Conservation during Anaerobic Respiration of
Shewanella oneidensis
Strain MR-1
Authors: Hunt Kristopher A. , Flynn Jeffrey M. , Naranjo Belén , Shikhare Indraneel D. , Gralnick Jeffrey A. 
Published: April 2010

ABSTRACT

It is well established that respiratory organisms use proton motive force to produce ATP via F-type ATP synthase aerobically and that this process may reverse during anaerobiosis to produce proton motive force. Here, we show that
Shewanella oneidensis
strain MR-1, a nonfermentative, facultative anaerobe known to respire exogenous electron acceptors, generates ATP primarily from substrate-level phosphorylation under anaerobic conditions. Mutant strains lacking
ackA
(SO2915) and
pta
(SO2916), genes required for acetate production and a significant portion of substrate-level ATP produced anaerobically, were tested for growth. These mutant strains were unable to grow anaerobically with lactate and fumarate as the electron acceptor, consistent with substrate-level phosphorylation yielding a significant amount of ATP. Mutant strains lacking
ackA
and
pta
were also shown to grow slowly using
N
-acetylglucosamine as the carbon source and fumarate as the electron acceptor, consistent with some ATP generation deriving from the Entner-Doudoroff pathway with this substrate. A deletion strain lacking the sole F-type ATP synthase (SO4746 to SO4754) demonstrated enhanced growth on
N
-acetylglucosamine and a minor defect with lactate under anaerobic conditions. ATP synthase mutants grown anaerobically on lactate while expressing proteorhodopsin, a light-dependent proton pump, exhibited restored growth when exposed to light, consistent with a proton-pumping role for ATP synthase under anaerobic conditions. Although
S. oneidensis
requires external electron acceptors to balance redox reactions and is not fermentative, we find that substrate-level phosphorylation is its primary anaerobic energy conservation strategy. Phenotypic characterization of an
ackA
deletion in
Shewanella
sp. strain MR-4 and genomic analysis of other sequenced strains suggest that this strategy is a common feature of
Shewanella
.
A Unique Group of Virus-Related, Genome-Integrating Elements Found Solely in the Bacterial Family
Thermaceae
and the Archaeal Family
Halobacteriaceae
Authors: Jalasvuori Matti , Pawlowski Alice , Bamford Jaana K. H. 
Published: April 2010

ABSTRACT

Viruses SH1 and P23-77, infecting archaeal
Haloarcula
species and bacterial
Thermus
species, respectively, were recently designated to form a novel viral lineage. In this study, the lineage is expanded to archaeal
Halomicrobium
and bacterial
Meiothermus
species by analysis of five genome-integrated elements that share the core genes with these viruses.
The Rut Pathway for Pyrimidine Degradation: Novel Chemistry and Toxicity Problems
Authors: Kim Kwang-Seo , Pelton Jeffrey G. , Inwood William B. , Andersen Ulla , Kustu Sydney , Wemmer David E. 
Published: April 2010

ABSTRACTThe Rut pathway is composed of seven proteins, all of which are required byEscherichia coliK-12 to grow on uracil as the sole nitrogen source. The RutA and RutB proteins are central: no spontaneous suppressors arise in strains lacking them. RutA works in conjunction with a flavin reductase (RutF or a substitute) to catalyze a novel reaction. It directly cleaves the uracil ring between N-3 and C-4 to yield ureidoacrylate, as established by both nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry. Although ureidoacrylate appears to arise by hydrolysis, the requirements for the reaction and the incorporation of18O at C-4 from molecular oxygen indicate otherwise. Mass spectrometry revealed the presence of a small amount of product with the mass of ureidoacrylate peracid in reaction mixtures, and we infer that this is the direct product of RutA.In vitroRutB cleaves ureidoacrylate hydrolytically to release 2 mol of ammonium, malonic semialdehyde, and carbon dioxide. Presumably the direct products are aminoacrylate and carbamate, both of which hydrolyze spontaneously. Together with bioinformatic predictions and published crystal structures, genetic and physiological studies allow us to predict functions for RutC, -D, and -E.In vivowe postulate that RutB hydrolyzes the peracid of ureidoacrylate to yield the peracid of aminoacrylate. We speculate that RutC reduces aminoacrylate peracid to aminoacrylate and RutD increases the rate of spontaneous hydrolysis of aminoacrylate. The function of RutE appears to be the same as that of YdfG, which reduces malonic semialdehyde to 3-hydroxypropionic acid. RutG appears to be a uracil transporter.
The Phosphoenolpyruvate Phosphotransferase System Regulates
Vibrio cholerae
Biofilm Formation through Multiple Independent Pathways
Authors: Houot Laetitia , Chang Sarah , Pickering Bradley S. , Absalon Cedric , Watnick Paula I. 
Published: April 2010

ABSTRACT

The bacterial phosphoenolpyruvate phosphotransferase system (PTS) is a highly conserved phosphotransfer cascade that participates in the transport and phosphorylation of selected carbohydrates and modulates many cellular functions in response to carbohydrate availability. It plays a role in the virulence of many bacterial pathogens. Components of the carbohydrate-specific PTS include the general cytoplasmic components enzyme I (EI) and histidine protein (HPr), the sugar-specific cytoplasmic components enzymes IIA (EIIA) and IIB (EIIB), and the sugar-specific membrane-associated multisubunit components enzymes IIC (EIIC) and IID (EIID). Many bacterial genomes also encode a parallel PTS pathway that includes the EI homolog EI
Ntr
, the HPr homolog NPr, and the EIIA homolog EIIA
Ntr
. This pathway is thought to be nitrogen specific because of the proximity of the genes encoding this pathway to the genes encoding the nitrogen-specific σ factor σ
54
. We previously reported that phosphorylation of HPr and FPr by EI represses
Vibrio cholerae
biofilm formation in minimal medium supplemented with glucose or pyruvate. Here we report two additional PTS-based biofilm regulatory pathways that are active in LB broth but not in minimal medium. These pathways involve the glucose-specific enzyme EIIA (EIIA
Glc
) and two nitrogen-specific EIIA homologs, EIIA
Ntr1
and EIIA
Ntr2
. The presence of multiple, independent biofilm regulatory circuits in the PTS supports the hypothesis that the PTS and PTS-dependent substrates have a central role in sensing environments suitable for a surface-associated existence.
Identification of the Quorum-Sensing Target DNA Sequence andN-Acyl Homoserine Lactone Responsiveness of theBrucella abortus virBpromoter
Authors: Arocena Gastón M. , Sieira Rodrigo , Comerci Diego J. , Ugalde Rodolfo A. 
Published: April 2010

ABSTRACTVjbR is a LuxR-type quorum-sensing (QS) regulator that plays an essential role in the virulence of the intracellular facultative pathogenBrucella, the causative agent of brucellosis. It was previously described that VjbR regulates a diverse group of genes, including thevirBoperon. The latter codes for a type IV secretion system (T4SS) that is central for the pathogenesis ofBrucella. Although the regulatory role of VjbR on thevirBpromoter (PvirB) was extensively studied by different groups, the VjbR-binding site had not been identified so far. Here, we identified the target DNA sequence of VjbR in PvirBby DNase I footprinting analyses. Surprisingly, we observed that VjbR specifically recognizes a sequence that is identical to a half-binding site of the QS-related regulator MrtR ofMesorhizobium tianshanense. As shown by DNase I footprinting and electrophoretic mobility shift assays, generation of a palindromic MrtR-like-binding site in PvirBincreased both the affinity and the stability of the VjbR-DNA complex, which confirmed that the QS regulator ofBrucellais highly related to that ofM. tianshanense. The addition ofN-dodecanoyl homoserine lactone dissociated VjbR from the promoter, which confirmed previous reports that indicated a negative effect of this signal on the VjbR-mediated activation of PvirB. Our results provide new molecular evidence for the structure of thevirBpromoter and reveal unusual features of the QS target DNA sequence of the main regulator of virulence inBrucella.