bluuebunny/crossref_metadata_2025_split huggingface dataset:
Altered Oligomerization Properties of N316 Mutants ofEscherichia coliTyrR
Authors: Koyanagi Takashi , Katayama Takane , Suzuki Hideyuki , Kumagai Hidehiko 
Published: October 2008

ABSTRACTThe transcriptional regulator TyrR is known to undergo a dimer-to-hexamer conformational change in response to aromatic amino acids, through which it controls gene expression. In this study, we identified N316D as the second-site suppressor ofEscherichia coliTyrRE274Q, a mutant protein deficient in hexamer formation. N316 variants exhibited altered in vivo regulatory properties, and the most drastic changes were observed for TyrRN316Dand TyrRN316Rmutants. Gel filtration analyses revealed that the ligand-mediated oligomer formation was enhanced and diminished for TyrRN316Dand TyrRN316R, respectively, compared with the wild-type TyrR. ADP was substituted for ATP in the oligomer formation of TyrRN316D.
Induction of the Galactose Enzymes in
Escherichia coli
Is Independent of the C-1-Hydroxyl Optical Configuration of the Inducer
d
-Galactose
Authors: Lee Sang Jun , Lewis Dale E. A. , Adhya Sankar 
Published: October 2008

ABSTRACT

The two optical forms of aldohexose galactose differing at the C-1 position, α-
d
-galactose and β-
d
-galactose, are widespread in nature. The two anomers also occur in di- and polysaccharides, as well as in glycoconjugates. The anomeric form of
d
-galactose, when present in complex carbohydrates, e.g., cell wall, glycoproteins, and glycolipids, is specific. Their interconversion occurs as monomers and is effected by the enzyme mutarotase (aldose-1-epimerase). Mutarotase and other
d
-galactose-metabolizing enzymes are coded by genes that constitute an operon in
Escherichia coli
. The operon is repressed by the repressor GalR and induced by
d
-galactose. Since, depending on the carbon source during growth, the cell can make only one of the two anomers of
d
-galactose, the cell must also convert one anomer to the other for use in specific biosynthetic pathways. Thus, it is imperative that induction of the
gal
operon, specifically the mutarotase, be achievable by either anomer of
d
-galactose. Here we report in vivo and in vitro experiments showing that both α-
d
-galactose and β-
d
-galactose are capable of inducing transcription of the
gal
operon with equal efficiency and kinetics. Whereas all substitutions at the C-1 position in the α configuration inactivate the induction capacity of the sugar, the effect of substitutions in the β configuration varies depending upon the nature of the substitution; methyl and phenyl derivatives induce weakly, but the glucosyl derivative does not.
Identification of
Bdellovibrio bacteriovorus
HD100 Bd0714 as a Nudix dGTPase
Authors: Steyert Susan R. , Messing Simon A. J. , Amzel L. Mario , Gabelli Sandra B. , Piñeiro Silvia A. 
Published: October 2008

ABSTRACT

Bdellovibrio bacteriovorus
bacteria are predatory organisms that attack other gram-negative bacteria. Here, we report that Bd0714 is a Nudix dGTPase from
B. bacteriovoru
s HD100 with a substrate specificity similar to that of
Escherichia coli
MutT and complements an
E. coli mutT
-deficient strain. We observed different transcription levels of the gene throughout the predator life cycle.
Phosphorus Deprivation Responses and Phosphonate Utilization in a Thermophilic
Synechococcus
sp. from Microbial Mats
Authors: Adams Melissa M. , Gómez-García María R. , Grossman Arthur R. , Bhaya Devaki 
Published: October 2008

ABSTRACT

The genomes of two closely related thermophilic cyanobacterial isolates, designated
Synechococcus
isolate OS-A and
Synechococcus
isolate OS-B′, from the microbial mats of Octopus Spring (Yellowstone National Park) have been sequenced. An extensive suite of genes that are controlled by phosphate levels constitute the putative Pho regulon in these cyanobacteria. We examined physiological responses of an axenic OS-B′ isolate as well as transcript abundances of Pho regulon genes as the cells acclimated to phosphorus-limiting conditions. Upon imposition of phosphorus deprivation, OS-B′ stopped dividing after three to four doublings, and absorbance spectra measurements indicated that the cells had lost most of their phycobiliproteins and chlorophyll
a
. Alkaline phosphatase activity peaked and remained high after 48 h of phosphorus starvation, and there was an accumulation of transcripts from putative Pho regulon genes. Interestingly, the genome of
Synechococcus
isolate OS-B′ harbors a cluster of
phn
genes that are not present in OS-A isolates. The proteins encoded by the
phn
genes function in the transport and metabolism of phosphonates, which could serve as an alternative phosphorus source when exogenous phosphate is low. The
phn
genes were upregulated within a day of eliminating the source of phosphate from the medium. However, the ability of OS-B′ to utilize methylphosphonate as a sole phosphorus source occurred only after an extensive period of exposure to the substrate. Once acclimated, the cells grew rapidly in fresh medium with methylphosphonate as the only source of phosphorus. The possible implications of these results are discussed with respect to the ecophysiology of the microbial mats.
Sequence and Analysis of a Plasmid-Encoded Mercury Resistance Operon from
Mycobacterium marinum
Identifies MerH, a New Mercuric Ion Transporter
Authors: Schué Mathieu , Dover Lynn G. , Besra Gurdyal. S. , Parkhill Julian , Brown Nigel L. 
Published: October 2008

ABSTRACT

In this study, we report the DNA sequence and biological analysis of a mycobacterial mercury resistance operon encoding a novel Hg
2+
transporter. MerH was found to transport mercuric ions in
Escherichia coli
via a pair of essential cysteine residues but only when coexpressed with the mercuric reductase.
Phosphatidylethanolamine Synthesis Is Required for Optimal Virulence of
Brucella abortus
Authors: Bukata Lucas , Altabe Silvia , de Mendoza Diego , Ugalde Rodolfo A. , Comerci Diego J. 
Published: October 2008

ABSTRACT

The
Brucella
cell envelope contains the zwitterionic phospholipids phosphatidylcholine (PC) and phosphatidylethanolamine (PE). Synthesis of PC occurs exclusively via the PC synthase pathway, implying that the pathogen depends on the choline synthesized by the host cell to form PC. Notably, PC is necessary to sustain a chronic infection process, which suggests that the membrane lipid content is relevant for
Brucella
virulence. In this study we investigated the first step of PE biosynthesis in
B. abortus
, which is catalyzed by phosphatidylserine synthase (PssA). Disruption of
pssA
abrogated the synthesis of PE without affecting the growth in rich complex medium. In minimal medium, however, the mutant required choline supplementation for growth, suggesting that at least PE or PC is necessary for
Brucella
viability. The absence of PE altered cell surface properties, but most importantly, it impaired several virulence traits of
B. abortus
, such as intracellular survival in both macrophages and HeLa cells, the maturation of the replicative
Brucella
-containing vacuole, and mouse colonization. These results suggest that membrane phospholipid composition is critical for the interaction of
B. abortus
with the host cell.
Amino Acid Substitutions in Transmembrane Domains 9 and 10 of GerVB That Affect the Germination Properties of
Bacillus megaterium
Spores
Authors: Christie Graham , Lowe Christopher R. 
Published: October 2008

ABSTRACT

The molecular basis for differences in germinant recognition of
Bacillus megaterium
QM B1551 spores containing the GerVB and/or GerUB receptor proteins has been examined by site-directed mutagenesis and the construction of cross-homologue chimeras. Focusing on nonconserved residues predicted to reside in transmembrane domains 9 and 10, we demonstrate that GerVB residues Ser319 and Leu345 are of particular importance in defining the specificity and apparent affinity of the receptor for germinants. Kinetic analyses of mutants with different amino acid substitutions at these positions indicate that Ser319 and Leu345 are not involved directly in the binding of germinants, but probably reside in regions of the receptor where structural perturbations can affect the conformation of, or access to, germinant binding sites. Position 345 is also shown to be of importance in GerUB, where the F345A mutation severely impairs receptor function. Functionality is restored in the GerUB Ala345 background by substituting putative outer-loop residues adjacent to TM10 for the corresponding residues in GerVB, indicating that a degree of structural coordination between these regions is important to receptor function.
Macroamphiphilic Components of Thermophilic Actinomycetes: Identification of Lipoteichoic Acid in
Thermobifida fusca
Authors: Rahman Obaidur , Pfitzenmaier Markus , Pester Oxana , Morath Siegfried , Cummings Stephen P. , Hartung Thomas , Sutcliffe Iain C. 
Published: October 2008

ABSTRACT

The cell envelopes of gram-positive bacteria contain structurally diverse membrane-anchored macroamphiphiles (lipoteichoic acids and lipoglycans) whose functions are poorly understood. Since regulation of membrane composition is an important feature of adaptation to life at higher temperatures, we have examined the nature of the macroamphiphiles present in the thermophilic actinomycetes
Thermobifida fusca
and
Rubrobacter xylanophilus
. Following hot-phenol-water extraction and purification by hydrophobic interaction chromatography, Western blotting with a monoclonal antibody against lipoteichoic acid strongly suggested the presence of a polyglycerophosphate lipoteichoic acid in
T. fusca
. This structure was confirmed by chemical and nuclear magnetic resonance analyses, which confirmed that the lipoteichoic acid is substituted with β-glucosyl residues, in common with the teichoic acid of this organism. In contrast, several extraction methods failed to recover significant macroamphiphilic carbohydrate- or phosphate-containing material from
R. xylanophilus
, suggesting that this actinomycete most likely lacks a membrane-anchored macroamphiphile. The finding of a polyglycerophosphate lipoteichoic acid in
T. fusca
suggests that lipoteichoic acids may be more widely present in the cell envelopes of actinomycetes than was previously assumed. However, the apparent absence of macroamphiphiles in the cell envelope of
R. xylanophilus
is highly unusual and suggests that macroamphiphiles may not always be essential for cell envelope homeostasis in gram-positive bacteria.
Proteasomal Components Required for Cell Growth and Stress Responses in the Haloarchaeon
Haloferax volcanii
Authors: Zhou Guangyin , Kowalczyk David , Humbard Matthew A. , Rohatgi Sunil , Maupin-Furlow Julie A. 
Published: October 2008

ABSTRACT

Little is known regarding the biological roles of archaeal proteases. The haloarchaeon
Haloferax volcanii
is an ideal model for understanding these enzymes, as it is one of few archaea with an established genetic system. In this report, a series of
H. volcanii
mutant strains with markerless and/or conditional knockouts in each known proteasome gene was systematically generated and characterized. This included single and double knockouts of genes encoding the 20S core α1 (
psmA
), β (
psmB
), and α2 (
psmC
) subunits as well as genes (
panA
and
panB
) encoding proteasome-activating nucleotidase (PAN) proteins closely related to the regulatory particle triple-A ATPases (Rpt) of eukaryotic 26S proteasomes. Our results demonstrate that 20S proteasomes are required for growth. Although synthesis of 20S proteasomes containing either α1 or α2 could be separately abolished via gene knockout with little to no impact on growth, conditional depletion of either β alone or α1 and α2 together rendered the cells inviable. In contrast, the PAN proteins were not essential based on the robust growth of the
panA panB
double knockout strain. Deletion of genes encoding either α1 or PanA did, however, render cells more sensitive to growth on organic versus inorganic nitrogen sources and hypo-osmotic stress and limited growth in the presence of
l
-canavanine. Abolishment of α1 synthesis also had a severe impact on the ability of cells to withstand thermal stress. This contrasted with what was seen for
panA
knockouts, which displayed enhanced thermotolerance. Together, these results provide new and important insight into the biological role of proteasomes in archaea.
Loss of a Biofilm-Inhibiting Glycosyl Hydrolase during the Emergence of
Yersinia pestis
Authors: Erickson David L. , Jarrett Clayton O. , Callison Julie A. , Fischer Elizabeth R. , Hinnebusch B. Joseph 
Published: October 2008

ABSTRACT

Yersinia pestis
, the bacterial agent of plague, forms a biofilm in the foregut of its flea vector to produce a transmissible infection. The closely related
Yersinia pseudotuberculosis
, from which
Y. pestis
recently evolved, can colonize the flea midgut but does not form a biofilm in the foregut.
Y. pestis
biofilm in the flea and in vitro is dependent on an extracellular matrix synthesized by products of the
hms
genes; identical genes are present in
Y. pseudotuberculosis
. The
Yersinia
Hms proteins contain functional domains present in
Escherichia coli
and
Staphylococcus
proteins known to synthesize a poly-β-1,6-
N
-acetyl-
d
-glucosamine biofilm matrix. In this study, we show that the extracellular matrices (ECM) of
Y. pestis
and staphylococcal biofilms are antigenically related, indicating a similar biochemical structure. We also characterized a glycosyl hydrolase (NghA) of
Y. pseudotuberculosis
that cleaved β-linked
N
-acetylglucosamine residues and reduced biofilm formation by staphylococci and
Y. pestis
in vitro. The
Y. pestis nghA
ortholog is a pseudogene, and overexpression of functional
nghA
reduced ECM surface accumulation and inhibited the ability of
Y. pestis
to produce biofilm in the flea foregut. Mutational loss of this glycosidase activity in
Y. pestis
may have contributed to the recent evolution of flea-borne transmission.
Defining the Topology of the N-Glycosylation Pathway in the Halophilic Archaeon
Haloferax volcanii
Authors: Plavner Noa , Eichler Jerry 
Published: October 2008

ABSTRACT

In
Eukarya
, N glycosylation involves the actions of enzymes working on both faces of the endoplasmic reticulum membrane. The steps of bacterial N glycosylation, in contrast, transpire essentially on the cytoplasmic side of the plasma membrane, with only transfer of the assembled glycan to the target protein occurring on the external surface of the cell. For
Archaea
, virtually nothing is known about the topology of enzymes involved in assembling those glycans that are subsequently N linked to target proteins on the external surface of the cell. To remedy this situation, subcellular localization and topology predictive algorithms, protease accessibility, and immunoblotting, together with cysteine modification following site-directed mutagenesis, were enlisted to define the topology of
Haloferax volcanii
proteins experimentally proven to participate in the N-glycosylation process. AglJ and AglD, involved in the earliest and latest stages, respectively, of assembly of the pentasaccharide decorating the
H. volcanii
S-layer glycoprotein, were shown to present their soluble N-terminal domain, likely containing the putative catalytic site of each enzyme, to the cytosol. The same holds true for Alg5-B, Dpm1-A, and Mpg1-D, proteins putatively involved in this posttranslational event. The results thus point to the assembly of the pentasaccharide linked to certain Asn residues of the
H. volcanii
S-layer glycoprotein as occurring within the cell.
A Novel Class of Modular Transporters for Vitamins in Prokaryotes
Authors: Rodionov Dmitry A. , Hebbeln Peter , Eudes Aymerick , ter Beek Josy , Rodionova Irina A. , Erkens Guus B. , Slotboom Dirk J. , Gelfand Mikhail S. , Osterman Andrei L. , Hanson Andrew D. , Eitinger Thomas 
Published: October 2008

ABSTRACT
The specific and tightly controlled transport of numerous nutrients and metabolites across cellular membranes is crucial to all forms of life. However, many of the transporter proteins involved have yet to be identified, including the vitamin transporters in various human pathogens, whose growth depends strictly on vitamin uptake. Comparative analysis of the ever-growing collection of microbial genomes coupled with experimental validation enables the discovery of such transporters. Here, we used this approach to discover an abundant class of vitamin transporters in prokaryotes with an unprecedented architecture. These transporters have energy-coupling modules comprised of a conserved transmembrane protein and two nucleotide binding proteins similar to those of ATP binding cassette (ABC) transporters, but unlike ABC transporters, they use small integral membrane proteins to capture specific substrates. We identified 21 families of these substrate capture proteins, each with a different specificity predicted by genome context analyses. Roughly half of the substrate capture proteins (335 cases) have a dedicated energizing module, but in 459 cases distributed among almost 100 gram-positive bacteria, including numerous human pathogens, different and unrelated substrate capture proteins share the same energy-coupling module. The shared use of energy-coupling modules was experimentally confirmed for folate, thiamine, and riboflavin transporters. We propose the name energy-coupling factor transporters for the new class of membrane transporters.
An Essential DnaB Helicase ofBacillus anthracis: Identification, Characterization, and Mechanism of Action
Authors: Biswas Esther E. , Barnes Marjorie H. , Moir Donald T. , Biswas Subhasis B. 
Published: October 2008

ABSTRACTWe have described a novel essential replicative DNA helicase fromBacillus anthracis, the identification of its gene, and the elucidation of its enzymatic characteristics. Anthrax DnaB helicase (DnaBBA) is a 453-amino-acid, 50-kDa polypeptide with ATPase and DNA helicase activities. DnaBBAdisplayed distinct enzymatic and kinetic properties. DnaBBAhas low single-stranded DNA (ssDNA)-dependent ATPase activity but possesses a strong 5′→3′ DNA helicase activity. The stimulation of ATPase activity appeared to be a function of the length of the ssDNA template rather than of ssDNA binding alone. The highest specific activity was observed with M13mp19 ssDNA. The results presented here indicated that the ATPase activity of DnaBBAwas coupled to its migration on an ssDNA template rather than to DNA binding alone. It did not require nucleotide to bind ssDNA. DnaBBAdemonstrated a strong DNA helicase activity that required ATP or dATP. Therefore, DnaBBAhas an attenuated ATPase activity and a highly active DNA helicase activity. Based on the ratio of DNA helicase and ATPase activities, DnaBBAis highly efficient in DNA unwinding and its coupling to ATP consumption.
TROP2 expression as prognostic marker for gastric carcinoma
Authors: Mühlmann G , Spizzo G , Gostner J , Zitt M , Maier H , Moser P , Gastl G , Zitt M , Müller H M , Margreiter R , Öfner D , Fong D 
Published: October 2008

Background:In gastric cancer the recurrence rate is unacceptably high, even after R0 resection and (neo)adjuvant chemotherapy. Therefore, there is an urgent need for identification of predictive and/or prognostic biomarkers to select high-risk patients who might benefit from additional therapies. Expression of TROP2 has been shown to be associated with tumour aggressiveness and poor prognosis in patients with various epithelial cancers.Aims:To investigate TROP2 expression in gastric cancer and its correlation with clinicopathological features and disease outcome.Methods:Expression of TROP2 was investigated by immunohistochemistry of tumour specimens from 104 patients who underwent resection for gastric cancer. Parameters found to be of prognostic significance in univariate analysis were verified in a multivariate Cox regression model.Results:TROP2 was found to be overexpressed in 58 (56%) tumour samples. Significantly higher expression of TROP2 could be detected in intestinal-type carcinomas (p = 0.03). In intestinal-type gastric cancer, TROP2 overexpression was significantly correlated with shorter disease-free survival (DFS) (p = 0.03). Among the total group, TROP2 overexpression was predictive for poor disease-free (p<0.01) and overall (p = 0.03) survival in lymph node positive patients. Multivariate Cox regression analysis revealed TROP2 overexpression to be an independent prognostic marker for poor DFS in the subgroup of patients with intestinal-type gastric cancer irrespective of lymph node involvement.Conclusion:Results show that TROP2 is an independent prognostic marker for disease recurrence in intestinal type gastric cancer. Due to its wide distribution TROP2 may become an attractive therapeutic target in a subgroup of patients with gastric cancer.
Non-melanocytic mimics of melanoma: Part I: intraepidermal mimics
Authors: Kamil Z S , Tong L C B , Habeeb A Al , Ghazarian D 
Published: October 2008

Melanoma comprises a wide range of cytological and architectural features histopathologically and hence can mimic many benign and malignant lesions of epithelial, mesenchymal and hematopoietic cell lines of differentiation.Therefore, analysis and close clinical, histological, histochemical and immunohistochemical correlation is vital in distinguishing challenging melanoma cases from their mimics. In this review, the different features of the benign, pre-malignant and malignant intraepidermal non-melanocytic tumours and tumour-like lesions that can closely mimic intraepidermal melanoma (melanoma in situ) are emphasised.
Immunohistochemical evaluation of ProEx C in human papillomavirus-induced lesions of the cervix
Authors: Conesa-Zamora P , Doménech-Peris A , Ortiz-Reina S , Orantes-Casado F J , Acosta-Ortega J , García-Solano J , Pérez-Guillermo M 
Published: October 2008

Aims:Considering the sparse information about the clinical utility of the novel immunohistochemical marker ProEx C in histological sections, a decision was taken to study the pattern of ProEx C expression in normal/benign cervical epithelium (N/B), low-grade squamous intraepithelial lesion (LGSIL) and high-grade squamous intraepithelial lesion (HGSIL), as well as the association of ProEx C expression with human papillomavirus (HPV) genotypes.Methods:100 cervical samples, including 21 N/B cervices, 16 LGSILs, 61 HGSILs and two cervical invasive carcinomas, were obtained from conisation and hysterectomy. Surgical specimens were arranged in three tissue microarrays and stained for ProEx C. Ninety-three samples were HPV genotyped. Genotyping was performed by DNA amplification and hybridisation with genotype-specific probes on a low-density DNA array.Results:ProEx C-positive expression in more than the lower third of the epithelium was observed in 14.3% of N/B, 62.5% of LGSIL and 90.2% of HGSIL. Seventy percent of HPV positivity was found in cases with expression in more than the lower third of the epithelium. Of 31 cases that were positive for HPV16, 16.1% showed ProEx C expression restricted to one or two basal layers, and 83.9% showed ProEx C expression in more than the lower third of the epithelium.Conclusions:ProEx C is significantly associated with HPV16 infection and is a useful adjunct in the identification of LGSIL and HGSIL in histological sections when expressed in more than the lower third of the epithelium.