bluuebunny/crossref_metadata_2025_split huggingface dataset:
Gene expression of cell surface antigens in the early phase of murine influenza pneumonia determined by a cDNA expression array technique
Authors: Sakai Shinya , Mantani Naoki , Kogure Toshiaki , Ochiai Hiroshi , Shimada Yutaka , Terasawa Katsutoshi 
Published: February 2003

Background: Influenza virus is a worldwide health problem with significant economic consequences. To study the gene expression pattern induced by influenza virus infection, it is useful to reveal the pathogenesis of influenza virus infection; but this has not been well examined, especially in vivo study.Aims: To assess the influence of influenza virus infection on gene expression in mice, mRNA levels in the lung and tracheal tissue 48 h after infection were investigated by cDNA array analysis.Methods: Four‐week‐old outbred, specific pathogen free strain, ICR female mice were infected by intra‐nasal inoculation of a virus solution under ether anesthesia. The mice were sacrificed 48 h after infection and the tracheas and lungs were removed. To determine gene expression, the membrane‐based microtechnique with an Atlas cDNA expression array (mouse 1.2 array II) was performed in accordance with the manual provided.Results and conclusions: We focused on the expression of 46 mRNAs for cell surface antigens. Of these 46 mRNAs that we examined, four (CD1d2 antigen, CD39 antigen‐like 1, CD39 antigen‐like 3, CD68 antigen) were up‐regulated and one (CD36 antigen) was down‐regulated. Although further studies are required, these data suggest that these molecules play an important role in influenza virus infection, especially the phase before specific immunity.
Anarchism, Socialism, and Social Reform
Authors: Clark John Bates 
Published: February 2003

[First page missing from the archive] … embitterment against society; but anarchism assimilates to itself that hostility to law which naturally results from evil doing. The antipathy of the rogue for the halter reinforces this movement. It gathers to itself disreputable elements, and by its outbreaks moves society to resistance. It is a self-terminating movement.Socialism appeals to better classes and has far more strength. Attack the state and you excite feelings of loyalty even among the disaffected classes; but attack the industrial system and appeal to the state, and you may have loyalty in your favor.
The conserved Glu‐60 residue in Thermoanaerobacter brockii alcohol dehydrogenase is not essential for catalysis
Authors: Kleifeld Oded , Shi Shu Ping , Zarivach Raz , Eisenstein Miriam , Sagi Irit 
Published: February 2003

AbstractGlu‐60 of the zinc‐dependent Thermoanaerobacter brockii alcohol dehydrogenase (TbADH) is a strictly conserved residue in all members of the alcohol dehydrogenase (ADH) family. Unlike most other ADHs, the crystal structures of TbADH and its analogs, ADH from Clostridium beijerinckii (CbADH), exhibit a unique zinc coordination environment in which this conserved residue is directly coordinated to the catalytic zinc ion in the native form of the enzymes. To explore the role of Glu‐60 in TbADH catalysis, we have replaced it by alanine (E60A‐TbADH) and aspartate (E60D‐TbADH). Steady‐state kinetic measurements show that the catalytic efficiency of these mutants is only four‐ and eightfold, respectively, lower than that of wild‐type TbADH. We applied X‐ray absorption fine‐structure (EXAFS) and near‐UV circular dichroism to characterize the local environment around the catalytic zinc ion in the variant enzymes in their native, cofactor‐bound, and inhibited forms. We show that the catalytic zinc site in the studied complexes of the variant enzymes exhibits minor changes relative to the analogous complexes of wild‐type TbADH. These moderate changes in the kinetic parameters and in the zinc ion environment imply that the Glu‐60 in TbADH does not remain bound to the catalytic zinc ion during catalysis. Furthermore, our results suggest that a water molecule replaces this residue during substrate turnover.
Structural basis for sequential displacement of Ca2+ by Yb3+ in a protozoan EF‐hand calcium binding protein
Authors: Atreya Hanudatta S. , Mukherjee Sulakshana , Chary Kandala V.R. , Lee Yong‐Min , Luchinat Claudio 
Published: February 2003

AbstractWe have studied the displacement of Ca2+by the trivalent lanthanide ions (Yb3+) in a protozoan (Entamoeba histolytica) Ca2+‐binding protein (EhCaBP), by NMR and thermodynamics. We have demonstrated, for the first time, how one can use in a combined fashion the utility of NMR and thermodynamics to have an insight to the relative binding specificities/affinity between Ca2+ and Yb3+. As revealed by the titration experiments, Yb3+ displaces Ca2+ from the four metal binding sites present in EhCaBP in a sequential manner. The study provides a structural origin for such a sequential Ca2+ displacement by Yb3+ in EhCaBP.
Interplay between hydrophobic cluster and loop propensity in β‐hairpin formation: A mechanistic study
Authors: Colombo Giorgio , De Mori Giacomo M.S. , Roccatano Danilo 
Published: February 2003

AbstractWe investigated the structural determinants of the stability of a designed β‐hairpin containing a natural hydrophobic cluster from the protein GB1 and a D‐Pro‐Gly turn forming sequence. The results of our simulations shed light on the factors leading to an ordered secondary structure in a model peptide: in particular, the importance of the so‐called diagonal interactions in forming a stable hydrophobic nucleus in the β‐hairpin, together with the more obvious lateral interactions, is examined. With the use of long timescale MD simulations in explicit water, we show the role of diagonal interactions in driving the peptide to the correct folded structure (formation of the hydrophobic core with Trp 2, Tyr 4, and Phe 9 in the first stages of refolding) and in keeping it in the ensemble of folded conformations. The combination of the stabilizing effects of the D‐Pro‐Gly turn sequence and of the hydrophobic nucleus formation thus favors the attainment of an ordered secondary structure compatible with the one determined experimentally. Moreover, our data underline the importance of the juxtapositions of the side chains of amino acids not directly facing each other in the three‐dimensional structure. The combination of these interactions forces the peptide to sample a nonrandom portion of the conformational space, as can be seen in the rapid collapse to an ordered structure in the refolding simulation, and shows that the unfolded state can be closely correlated to the folded ensemble of structures, at least in the case of small model peptides.
Probing the structure of falcipain‐3, a cysteine protease from Plasmodium falciparum: Comparative protein modeling and docking studies
Authors: Sabnis Yogesh A. , Desai Prashant V. , Rosenthal Philip J. , Avery Mitchell A. 
Published: February 2003

AbstractIncreasing resistance of malaria parasites to conventional antimalarial drugs is an important factor contributing to the persistence of the disease as a major health threat. The ongoing search for novel targets has resulted in identification and expression of several enzymes including cysteine proteases that are implicated in hemoglobin degradation. Falcipain‐2 and falcipain‐3 are considered to be the two principal cysteine proteases in this degradation, and hence, are potential drug targets. A homology model of falcipain‐3 was built and validated by various structure/geometry verification tools as well as docking studies of known substrates. The correlation coefficient of 0.975 between interaction energies and Km values of these substrates provided additional support for the model. On comparison with the previously reported falcipain‐2 homology model, the currently constructed falcipain‐3 structure showed important differences between the S2 pockets that might explain the variations in the Km values of various substrates for these enzymes. Further, docking studies also provided insight into possible binding modes and interactions of ligands with falcipain‐3. Results of the current study could be employed in de novo drug design leading to development of new antimalarial agents.
Asparagine 285 plays a key role in transition state stabilization in rabbit muscle creatine kinase
Authors: Borders Charles L. , MacGregor Katherine M. , Edmiston Paul L. , Gbeddy Elikem R.K. , Thomenius Michael J. , Mulligan Guy B. , Snider Mark J. 
Published: February 2003

AbstractTo explore the possibility that asparagine 285 plays a key role in transition state stabilization in phosphagen kinase catalysis, the N285Q, N285D, and N285A site‐directed mutants of recombinant rabbit muscle creatine kinase (rmCK) were prepared and characterized. Kinetic analysis of phosphocreatine formation showed that the catalytic efficiency of each N285 mutant was reduced by approximately four orders of magnitude, with the major cause of activity loss being a reduction in kcat in comparison to the recombinant native CK. The data for N285Q still fit a random‐order, rapid‐equilibrium mechanism, with either MgATP or creatine binding first with affinities very nearly equal to those for native CK. However, the affinity for the binding of the second substrate is reduced approximately 10‐fold, suggesting that addition of a single methylene group at position 285 disrupts the symphony of substrate binding. The data for the N285A mutant only fit an ordered binding mechanism, with MgATP binding first. Isosteric replacement to form the N285D mutant has almost no effect on the KM values for either creatine or MgATP, thus the decrease in activity is due almost entirely to a 5000‐fold reduction in kcat. Using the quenching of the intrinsic CK tryptophan fluorescence by added MgADP (Borders et al. 2002), it was found that, unlike native CK, none of the mutants have the ability to form a quaternary TSAC. We use these data to propose that asparagine 285 indeed plays a key role in transition state stabilization in the reaction catalyzed by creatine kinase and other phosphagen kinases.
X‐ray structure of the AAC(6′)‐Ii antibiotic resistance enzyme at 1.8 Å resolution; examination of oligomeric arrangements in GNAT superfamily members
Authors: Burk David L. , Ghuman Navleen , Wybenga‐Groot Leanne E. , Berghuis Albert M. 
Published: February 2003

AbstractThe rise of antibiotic resistance as a public health concern has led to increased interest in studying the ways in which bacteria avoid the effects of antibiotics. Enzymatic inactivation by several families of enzymes has been observed to be the predominant mechanism of resistance to aminoglycoside antibiotics such as kanamycin and gentamicin. Despite the importance of acetyltransferases in bacterial resistance to aminoglycoside antibiotics, relatively little is known about their structure and mechanism. Here we report the three‐dimensional atomic structure of the aminoglycoside acetyltransferase AAC(6′)‐Ii in complex with coenzyme A (CoA). This structure unambiguously identifies the physiologically relevant AAC(6′)‐Ii dimer species, and reveals that the enzyme structure is similar in the AcCoA and CoA bound forms. AAC(6′)‐Ii is a member of the GCN5‐related N‐acetyltransferase (GNAT) superfamily of acetyltransferases, a diverse group of enzymes that possess a conserved structural motif, despite low sequence homology. AAC(6′)‐Ii is also a member of a subset of enzymes in the GNAT superfamily that form multimeric complexes. The dimer arrangements within the multimeric GNAT superfamily members are compared, revealing that AAC(6′)‐Ii forms a dimer assembly that is different from that observed in the other multimeric GNAT superfamily members. This different assembly may provide insight into the evolutionary processes governing dimer formation.
Identification and Characterization of a
Streptococcus pyogenes
Operon Involved in Binding of Hemoproteins and Acquisition of Iron
Authors: Bates Christopher S. , Montañez Griselle E. , Woods Charles R. , Vincent Rebecca M. , Eichenbaum Zehava 
Published: February 2003

ABSTRACT

The hemolytic
Streptococcus pyogenes
can use a variety of heme compounds as an iron source. In this study, we investigate hemoprotein utilization by
S. pyogenes
. We demonstrate that surface proteins contribute to the binding of hemoproteins to
S. pyogenes
. We identify an ABC transporter from the iron complex family named
sia
for streptococcal iron acquisition, which consists of a lipoprotein (
siaA
), membrane permease (
siaB
), and ATPase (
siaC
). The
sia
transporter is part of a highly conserved, iron regulated, 10-gene operon. SiaA, which was localized to the cell membrane, could specifically bind hemoglobin. The operon's first gene encodes a novel bacterial protein that bound hemoglobin, myoglobin, heme-albumin, and hemoglobin-haptoglobin (but not apo-haptoglobin) and therefore was named Shr, for streptococcal hemoprotein receptor. PhoZ fusion and Western blot analysis showed that Shr has a leader peptide and is found in both membrane-bound and soluble forms. An M1 SF370 strain with a polar mutation in
shr
was more resistant to streptonigrin and hydrogen peroxide, suggesting decreased iron uptake. The addition of hemoglobin to the culture medium increased cell resistance to hydrogen peroxide in SF370 but not in the mutant, implying the
sia
operon may be involved in hemoglobin-dependent resistance to oxidative stress. The
shr
mutant demonstrated reduced hemoglobin binding, though cell growth in iron-depleted medium supplemented with hemoglobin, whole blood, or ferric citrate was not affected, suggesting additional systems are involved in hemoglobin utilization. SiaA and Shr are the first hemoprotein receptors identified in
S. pyogenes
; their possible role in iron capture is discussed.
The Afa/Dr Adhesins of Diffusely AdheringEscherichia coliStimulate Interleukin-8 Secretion, Activate Mitogen-Activated Protein Kinases, and Promote Polymorphonuclear Transepithelial Migration in T84 Polarized Epithelial Cells
Authors: Bétis Fréderic , Brest Patrick , Hofman Véronique , Guignot Julie , Bernet-Camard Marie-Françoise , Rossi Bernard , Servin Alain , Hofman Paul 
Published: February 2003

ABSTRACTAfa/Dr diffusely adheringEscherichia coli(Afa/Dr DAEC) strains cause symptomatic urinary tract and intestinal infections. The proinflammatory effects of Afa/Dr DAEC strains in vitro have been not investigated to date. In the present study, we used confluent polarized monolayers of intestinal cell line T84 to evaluate the consequences of epithelial infection by Afa/Dr DAEC strains in terms of proinflammatory response. Polymorphonuclear leukocyte (PMNL) migration across the epithelial barrier was induced after incubation of the T84 monolayers with the wild-type Afa/Dr DAEC strain C1845 harboring the fimbrial F1845 adhesin and strain IH11128 harboring the Dr hemagglutinin, and theE. colilaboratory strain HB101 was transformed with the pSSS1 plasmid, producing Afa/Dr F1845 adhesin. PMNL migrations were correlated with a basolateral secretion of interleukin-8 by T84 cells and were abolished after incubation of epithelial cells with an anti-decay accelerating factor (DAF) antibody that recognized the short consensus repeat 3 domain of DAF (monoclonal antibody 1H4). Moreover, Afa/Dr DAEC strains induced tyrosine phosphorylation of several T84 proteins and activated the mitogen-activated protein kinases (ERK1/2 mitogen-activated protein, P38, and Jun-C kinases). These data demonstrated for the first time that, in vitro, Afa/Dr DAEC strains exert a proinflammatory signal in intestinal epithelial cells.
Production of the Type IV Secretion System Differs among
Brucella
Species as Revealed with VirB5- and VirB8-Specific Antisera
Authors: Rouot Bruno , Alvarez-Martinez Maria-Teresa , Marius Carine , Menanteau Pierrette , Guilloteau Laurence , Boigegrain Rose-Anne , Zumbihl Robert , O'Callaghan David , Domke Natalie , Baron Christian 
Published: February 2003

ABSTRACT

Expression of the
virB
operon, encoding the type IV secretion system required for
Brucella suis
virulence, occurred in the acidic phagocytic vacuoles of macrophages and could be induced in minimal medium at acidic pH values. To analyze the production of VirB proteins, polyclonal antisera against
B. suis
VirB5 and VirB8 were generated. Western blot analysis revealed that VirB5 and VirB8 were detected after 3 h in acidic minimal medium and that the amounts increased after prolonged incubation. Unlike what occurs in the related organism
Agrobacterium tumefaciens
, the periplasmic sugar binding protein ChvE did not contribute to VirB protein production, and
B. suis
from which
chvE
was deleted was fully virulent in a mouse model. Comparative analyses of various
Brucella
species revealed that in all of them VirB protein production increased under acidic conditions. However, in rich medium at neutral pH,
Brucella canis
and
B. suis
, as well as the
Brucella abortus-
and
Brucella melitensis
-derived vaccine strains S19, RB51, and Rev.1, produced no VirB proteins or only small amounts of VirB proteins, whereas the parental
B. abortus
and
B. melitensis
strains constitutively produced VirB5 and VirB8. Thus, the vaccine strains were still able to induce
virB
expression under acidic conditions, but the VirB protein production was markedly different from that in the wild-type strains at pH 7. Taken together, the data indicate that VirB protein production and probably expression of the
virB
operon are not uniformly regulated in different
Brucella
species. Since VirB proteins were shown to modulate
Brucella
phagocytosis and intracellular trafficking, the differential regulation of the production of these proteins reported here may provide a clue to explain their role(s) during the infection process.
The Shufflon of
Salmonella enterica
Serovar Typhi Regulates Type IVB Pilus-Mediated Bacterial Self-Association
Authors: Morris Christina , Yip Cecilia M. C. , Tsui Inez S. M. , Wong Danny K.-H. , Hackett Jim 
Published: February 2003

ABSTRACT

Previously, it was shown that type IVB pili encoded by the
Salmonella enterica
serovar Typhi
pil
operon are used to facilitate bacterial entry into human intestinal epithelial cells in vitro and that such entry is inhibited by purified prepilin (pre-PilS) protein (X.-L. Zhang, I. S. M. Tsui, C. M. C. Yip, A. W. Y. Fung, D. K.-H. Wong, X. Dai, Y. Yang, J. Hackett, and C. Morris, Infect. Immun. 68:3067-3073, 2000). The
pil
operon concludes with a simple shufflon, and a recombinase gene product (Rci) inverts DNA in the C-terminal region of the
pilV
gene to allow synthesis of two distinct PilV proteins, PilV1 and PilV2, which are presumptive minor pilus proteins. We show here that the type IVB pili mediate bacterial self-association, but only when the PilV1 and PilV2 proteins are not expressed. This may be achieved in wild-type serovar Typhi by rapid DNA inversion activity of the shufflon. We show that the inversion activity inhibits the expression of genes inserted between the 19-bp inverted repeats used for Rci-mediated recombination and that the activity of Rci increases when DNA is supercoiled. The data suggest that serovar Typhi self-associates under conditions (such as low oxygen tension in the gut) that favor DNA supercoiling. These results explain (i) the function of the serovar Typhi shufflon and (ii) why there are only two possible shufflon states, in contrast to the many possible states of other shufflon systems. The data further indicate that a very early step in serovar Typhi pathogenesis may be type IVB pilus-mediated self-association of bacteria in the anaerobic human small intestine prior to invasion of the human gut epithelium. The suggested type IVB pilus-dependent step in typhoid fever pathogenesis may partially explain the enhanced invasiveness of serovar Typhi for humans.
Expression and Immunogenicity ofMycoplasma hyopneumoniaeHeat Shock Protein Antigen P42 by DNA Vaccination
Authors: Chen Ya-Lei , Wang Shao-Ning , Yang Wen-Jen , Chen Yi-Jiun , Lin Hsi-Hsun , Shiuan David 
Published: February 2003

ABSTRACTMycoplasma hyopneumoniaeis the etiological agent of swine enzootic pneumonia, a chronic nonfatal disease affecting pigs of all ages. The goal of this study was to design DNA vaccines by constructing plasmid pcDNA3/P42, carrying the heat shock protein gene P42 ofM. hyopneumoniae, and to evaluate the immune responses elicited in BALB/c mice. The expression of P42 was first examined in transfected NIH 3T3 cells by reverse transcription-PCR to ensure that the construct was functional. The humoral and cell-mediated immune responses induced by the plasmid were further evaluated in BALB/c mice through intramuscular injection. Both immunoglobulin G1 (IgG1) and IgG2a levels were 64 times those of the control groups during the first 8 weeks. The levels of interleukin-2 (IL-2), IL-4, and gamma interferon mRNAs in the immunized animals were elevated, and the proliferation of spleen cells was also enhanced in the immunized animals. The results indicate that pcDNA3/P42 DNA immunization induces both Th1 and Th2 immune responses. In addition, antiserum from the immunized animals was found to inhibit the growth ofM. hyopneumoniae. The present study reveals that DNA vaccination could be a new strategy against infection byM. hyopneumoniaeand may have potential for developing vaccines for other infectious diseases as well.
Purification, Gene Cloning, Gene Expression, and Mutants of Dps from the Obligate Anaerobe
Porphyromonas gingivalis
Authors: Ueshima Junichi , Shoji Mikio , Ratnayake Dinath B. , Abe Kihachiro , Yoshida Shinichi , Yamamoto Kenji , Nakayama Koji 
Published: February 2003

ABSTRACT

The periodontopathogen
Porphyromonas gingivalis
is an obligate anaerobe that is devoid of catalase but exhibits a relatively high degree of resistance to peroxide stress. In the present study, we demonstrate that
P. gingivalis
contains a Dps homologue that plays an important role in the protection of cells from peroxide stress. The Dps protein isolated from
P. gingivalis
displayed a ferritin-like spherical polymer consisting of 19-kDa subunits. Molecular cloning and sequencing of the gene encoding this protein revealed that it had a high similarity in nucleotide and amino acid sequences to Dps proteins from other species. The expression of Dps was significantly increased by exposure of
P. gingivalis
to atmospheric oxygen in an OxyR-dependent manner, indicating that it is regulated by the reactive oxygen species-regulating gene
oxyR
. The Dps-deficient mutants, including the
dps
single mutant and the
ftn dps
double mutant, showed no viability loss upon exposure to atmospheric oxygen for 6 h. In contrast to the wild type, however, these mutants exhibited the high susceptibility to hydrogen peroxide, thereby disrupting the viability. On the other hand, no significant difference in sensitivity to mitomycin C and metronidazole was observed between the wild type and the mutants. Furthermore, the
dps
single mutant, compared with the wild type, showed a lower viability in infected human umbilical vein endothelial cells.