bluuebunny/crossref_metadata_2025 huggingface dataset:
Dual Role of the PhoP∼P Response Regulator:Bacillus amyloliquefaciensFZB45 Phytase Gene Transcription Is Directed by Positive and Negative Interactions with thephyCPromoter
Authors: Makarewicz Oliwia , Dubrac Sarah , Msadek Tarek , Borriss Rainer
Published: September 2006

SeveralBacillusstrains secrete phytase, an enzyme catalyzing dephosphorylation ofmyo-inositol hexakisphosphate (phytate). We identified thephyC(phytase) gene from environmentalBacillus amyloliquefaciensFZB45 as a member of the phosphate starvation-inducible PhoPR regulon. In vivo and in vitro assays revealed that PhoP∼P is essential forphyC transcription. The transcriptional start site was identified downstream of a σA-like promoter region located 27 bp upstream of the probable translation ATG start codon. Inspection of thephyCpromoter sequence revealed an unusual structure. The− 35 and −10 regions are separated by a window of 21 bp. A pair of tandemly repeated PhoP TT(T/A/C)ACA binding boxes was located within and upstream of the −35 consensus promoter region. A single PhoP box was found within the −10 consensus promoter region. DNase I footprinting experiments performed with isolated PhoP confirmed that PhoP∼P binds at two sites overlapping with thephyC −35 and −10 consensus promoter region. While binding of dimeric PhoP∼P at −35 is essential for activation of thephyCpromoter, binding of PhoP∼P at− 10 suppresses promoter activity. A sixfold enhancement ofphyCgene expression was registered after T:G substitution of nucleotide −13 (mutant MUT13), which eliminates PhoP binding at the single PhoP box without impairing the −10 consensus sequence. Moreover, MUT13 also expressedphyCduring phosphate-replete growth, suggesting that the repressing effect due to binding of PhoP∼P at −10 was abolished. A model is presented in which transcription initiation ofphyCis positively and negatively affected by the actual concentration of the PhoP∼P response regulator.
Modes of Regulation of RpoS by H-NS
Authors: Zhou YanNing , Gottesman Susan
Published: September 2006

Regulated degradation of RpoS requires RssB and ClpXP protease. Mutations in
hns
increase both RpoS synthesis and stability, causing a twofold increase in synthesis and almost complete stabilization of RpoS, independent of effects on synthesis and independent of phosphorylation of RssB. This suggests that H-NS regulates an RssB inhibitor or inhibitors.
The TadV Protein ofActinobacillus actinomycetemcomitansIs a Novel Aspartic Acid Prepilin Peptidase Required for Maturation of the Flp1 Pilin and TadE and TadF Pseudopilins
Authors: Tomich Mladen , Fine Daniel H. , Figurski David H.
Published: September 2006

Thetadlocus ofActinobacillus actinomycetemcomitansencodes genes for the biogenesis of Flp pili, which allow the bacterium to adhere tenaciously to surfaces and form strong biofilms. Althoughtad(tightadherence) loci are widespread among bacterial and archaeal species, very little is known about the functions of the individual components of the Tad secretion apparatus. Here we characterize the mechanism by which the pre-Flp1 prepilin is processed to the mature pilus subunit. We demonstrate that thetadVgene encodes a prepilin peptidase that is both necessary and sufficient for proteolytic maturation of Flp1. TadV was also found to be required for maturation of the TadE and TadF pilin-like proteins, which we term pseudopilins. Using site-directed mutagenesis, we show that processing of pre-Flp1, pre-TadE, and pre-TadF is required for biofilm formation. Mutation of a highly conserved glutamic acid residue at position +5 of Flp1, relative to the cleavage site, resulted in a processed pilin that was blocked in assembly. In contrast, identical mutations in TadE or TadF had no effect on biofilm formation, indicating that the mechanisms by which Flp1 pilin and the pseudopilins function are distinct. We also determined that two conserved aspartic acid residues in TadV are critical for function of the prepilin peptidase. Together, our results indicate that theA. actinomycetemcomitansTadV protein is a member of a novel subclass of nonmethylating aspartic acid prepilin peptidases.
Structural Basis for Regioselectivity and Stereoselectivity of Product Formation by Naphthalene 1,2-Dioxygenase
Authors: Ferraro Daniel J. , Okerlund Adam L. , Mowers Jonathan C. , Ramaswamy S.
Published: September 2006

Rieske oxygenase (RO) systems are two- and three-component enzyme systems that catalyze the formation of
cis
-dihydrodiols from aromatic substrates. Degradation of pollutants in contaminated soil and generation of chiral synthons have been the major foci of RO research. Substrate specificity and product regio- and stereoselectivity have been shown to vary between individual ROs. While directed evolution methods for altering RO function have been successful in the past, rational engineering of these enzymes still poses a challenge due to the lack of structural understanding. Here we examine the structural changes induced by mutation of Phe-352 in naphthalene 1,2-dioxygenase from
Pseudomonas
sp. strain NCIB 9816-4 (NDO-O
9816-4
). Structures of the Phe-352-Val mutant in native form and in complex with phenanthrene and anthracene, along with those of wild-type NDO-O
9816-4
in complex with phenanthrene, anthracene, and 3-nitrotoluene, are presented. Phenanthrene was shown to bind in a different orientation in the Phe-352-Val mutant active site from that in the wild type, while anthracene was found to bind in similar positions in both enzymes. Two orientations of 3-nitrotoluene were observed, i.e., a productive and a nonproductive orientation. These orientations help explain why NDO-O
9816-4
forms different products from 3-nitrotoluene than those made from nitrobenzene dioxygenase. Comparison of these structures among themselves and with other known ROs bound to substrates reveals that the orientation of substrate binding at the active site is the primary determinant of product regio- and stereoselectivity.
Analysis of RNase P Protein (
rnpA
) Expression in
Bacillus subtilis
Utilizing Strains with Suppressible
rnpA
Expression
Authors: Gößringer Markus , Kretschmer-Kazemi Far Rosel , Hartmann Roland K.
Published: September 2006

Bacterial RNase P is composed of an RNA subunit and a single protein subunit (encoded by the
rnpB
and
rnpA
genes, respectively). We constructed
Bacillus subtilis
mutant strains that conditionally express the RNase P protein under control of the xylose promoter (P

xyl

). In one strain (d7),
rnpA
expression was efficiently repressed in the absence of the inducer xylose, leading to cell growth arrest. Growth could be restored by a second functional
rnpA
allele. This is the first RNase P protein knockdown strain, providing the first direct proof that the
rnpA
gene is essential in
B. subtilis
and, by inference, in other bacteria. We further show (i) that, in the wild-type context,
rnpA
expression is attenuated by transcriptional polarity and (ii) that translation of
rnpA
mRNA in
B. subtilis
can be initiated at two alternative start codons. His-tagged RNase P protein variants are functional in vivo and permit purification of in vivo-assembled holoenzymes by affinity chromatography. Simultaneous expression of plasmid-encoded RNase P RNA and His-tagged protein increased RNase P holoenzyme yields. Massive overproduction of RNase P protein in strain d7 is compatible with cell viability.
FlhF Is Required for Swimming and Swarming in
Pseudomonas aeruginosa
Authors: Murray Thomas S. , Kazmierczak Barbara I.
Published: September 2006

FlhF is a signal recognition particle-like protein present in monotrichous bacteria. The loss of FlhF in various bacteria results in decreased transcription of class II, III, or IV flagellar genes, leads to diminished or absent motility, and results in the assembly of flagella at nonpolar locations on the cell surface. In this work, we demonstrate that the loss of FlhF results in defective swimming and swarming motility of
Pseudomonas aeruginosa
. The FlhF protein localizes to the flagellar pole; in the absence of FlhF, flagellar assembly occurs but is no longer restricted to the pole. Δ
flhF
bacteria swim at lower velocities than wild-type bacteria in liquid media and can no longer swarm when assayed under standard swarming conditions (0.5% agar). However, Δ
flhF
bacteria regain swarming behavior when plated on 0.3% agar. Δ
flhF
organisms show decreased transcription and expression of flagellin (FliC) both in liquid media and on swarming plates compared to wild-type bacteria. However, changes in flagellin expression do not explain the different motility patterns observed for Δ
flhF
bacteria. Instead, the aberrant placement of flagella in Δ
flhF
bacteria may reduce their ability to move this rod-shaped organism effectively.
Genomic Analysis of
Pseudomonas aeruginosa
Phages LKD16 and LKA1: Establishment of the φKMV Subgroup within the T7 Supergroup
Authors: Ceyssens Pieter-Jan , Lavigne Rob , Mattheus Wesley , Chibeu Andrew , Hertveldt Kirsten , Mast Jan , Robben Johan , Volckaert Guido
Published: September 2006

Lytic
Pseudomonas aeruginosa
phages LKD16 and LKA1 were locally isolated and morphologically classified as
Podoviridae
. While LKD16 adsorbs weakly to its host, LKA1 shows efficient adsorption (

k
a

= 3.9 × 10
−9
ml min
−1
). LKA1, however, displays a narrow host range on clinical
P. aeruginosa
strains compared to LKD16. Genome analysis of LKD16 (43,200 bp) and LKA1 (41,593 bp) revealed that both phages have linear double-stranded DNA genomes with direct terminal repeats of 428 and 298 bp and encode 54 and 56 genes, respectively. The majority of the predicted structural proteins were experimentally confirmed as part of the phage particle using mass spectrometry. Phage LKD16 is closely related to bacteriophage φKMV (83% overall DNA homology), allowing a more thoughtful gene annotation of both genomes. In contrast, LKA1 is more distantly related, lacking significant DNA homology and showing protein similarity to φKMV in 48% of its gene products. The early region of the LKA1 genome has diverged strongly from φKMV and LKD16, and intriguing differences in tail fiber genes of LKD16 and LKA1 likely reflect the observed discrepancy in infection-related properties. Nonetheless, general genome organization is clearly conserved among φKMV, LKD16, and LKA1. The three phages carry a single-subunit RNA polymerase gene adjacent to the structural genome region, a feature which distinguishes them from other members of the T7 supergroup. Therefore, we propose that φKMV represents an independent and widespread group of lytic
P. aeruginosa
phages within the T7 supergroup.
Human Immunodeficiency Virus Type 1 V1-V2 Envelope Loop Sequences Expand and Add Glycosylation Sites over the Course of Infection, and These Modifications Affect Antibody Neutralization Sensitivity
Authors: Sagar Manish , Wu Xueling , Lee Sandra , Overbaugh Julie
Published: September 2006

Over the course of infection, human immunodeficiency virus type 1 (HIV-1) continuously adapts to evade the evolving host neutralizing antibody responses. Changes in the envelope variable loop sequences, particularly the extent of glycosylation, have been implicated in antibody escape. To document modifications that potentially influence antibody susceptibility, we compared envelope variable loops 1 and 2 (V1-V2) from multiple sequences isolated at the primary phase of infection to those isolated around 2 to 3 years into the chronic phase of infection in nine women with HIV-1 subtype A. HIV-1 sequences isolated during chronic infection had significantly longer V1-V2 loops, with a significantly higher number of potential N-linked glycosylation sites, than the sequences isolated early in infection. To assess the effects of these V1-V2 changes on antibody neutralization and infectivity, we created chimeric envelope sequences, which incorporated a subject's V1-V2 sequences into a common subtype A envelope backbone and then used them to generate pseudotyped viruses. Compared to the parent virus, the introduction of a subject's early-infection V1-V2 envelope variable loops rendered the chimeric envelope more sensitive to that subject's plasma samples but only to plasma samples collected >6 months after the sequences were isolated. Neutralization was not detected with the same plasma when the early-infection V1-V2 sequences were replaced with chronic-infection V1-V2 sequences, suggesting that changes in V1-V2 contribute to antibody escape. Pseudotyped viruses with V1-V2 segments from different times in infection, however, showed no significant difference in neutralization sensitivity to heterologous pooled plasma, suggesting that viruses with V1-V2 loops from early in infection were not inherently more neutralization sensitive. Pseudotyped viruses bearing chimeric envelopes with early-infection V1-V2 sequences showed a trend in infecting cells with low CD4 concentrations more efficiently, while engineered viruses with V1-V2 sequences isolated during chronic infection were moderately better at infecting cells with low CCR5 concentrations. These studies suggest that changes within the V1-V2 envelope domains over the course of an infection influence sensitivity to autologous neutralizing antibodies and may also impact host receptor/coreceptor interactions.
Characterization of Antibody Responses to Combinations of a Dengue Virus Type 2 DNA Vaccine and Two Dengue Virus Type 2 Protein Vaccines in Rhesus Macaques
Authors: Simmons Monika , Porter Kevin R. , Hayes Curtis G. , Vaughn David W. , Putnak Robert
Published: September 2006

We evaluated three nonreplicating dengue virus type 2 (DENV-2) vaccines: (i) a DNA vaccine containing the prM-E gene region (D), (ii) a recombinant subunit protein vaccine containing the B domain (i.e., domain III) of the E protein as a fusion with theEscherichia colimaltose-binding protein (R), and (iii) a purified inactivated virus vaccine (P). Groups of four rhesus macaques each were primed once and boosted twice using seven different vaccination regimens. After primary vaccination, enzyme-linked immunosorbent assay (ELISA) antibody levels increased most rapidly for groups inoculated with the P and DP combination, and by 1 month after the second boost, ELISA titers were similar for all groups. The highest plaque reduction neutralization test (PRNT) titers were seen in those groups that received the DR/DR/DR combination (geometric mean titer [GMT], 510), the P/P/P vaccine (GMT, 345), the DP/DP/DP combination (GMT, 287), and the R/R/R vaccine (GMT, 200). The next highest titers were seen in animals that received the D/R/R vaccine (GMT, 186) and the D/P/P vaccine (GMT, 163). Animals that received the D/D/D vaccine had the lowest neutralizing antibody titer (GMT, 49). Both ELISA and PRNT titers declined at variable rates. The only significant protection from viremia was observed in the P-vaccinated animals (mean of 0.5 days), which also showed the highest antibody concentration, including antibodies to NS1, and highest antibody avidity at the time of challenge.
Kaposi's Sarcoma-Associated Herpesvirus Lytic Switch Protein Stimulates DNA Binding of RBP-Jk/CSL To Activate the Notch Pathway
Authors: Carroll Kyla Driscoll , Bu Wei , Palmeri Diana , Spadavecchia Sophia , Lynch Stephen J. , Marras Salvatore A. E. , Tyagi Sanjay , Lukac David M.
Published: September 2006

Kaposi's sarcoma-associated herpesvirus (KSHV) lytic switch protein, Rta, is a ligand-independent inducer of the Notch signal transduction pathway, and KSHV cannot reactivate from latency in cells null for the Notch target protein RBP-Jk. Here we show that Rta promotes DNA binding of RBP-Jk, a mechanism that is fundamentally different from that established for the RBP-Jk-activating proteins, Notch intracellular domain (NICD) and Epstein-Barr virus EBNA2. Although constitutively active RBP-Jk and NICD do not transactivate KSHV promoters independently, cotransfection of an Rta mutant lacking its transactivation domain robustly restores transcriptional activation. Cooperation requires intact DNA binding sites for Rta and RBP-Jk and trimeric complex formation between the three molecules in vitro. In infected cells, RBP-Jk is virtually undetectable on a series of viral and cellular promoters during KSHV latency but is significantly enriched following Rta expression during viral reactivation. Accordingly, Rta, but not EBNA2 and NICD, reactivates the complete viral lytic cycle.
Induction of Human Immunodeficiency Virus Type 1 (HIV-1)-Specific T-Cell Responses in HIV Vaccine Trial Participants Who Subsequently Acquire HIV-1 Infection
Authors: Horton Helen , Havenar-Daughton Colin , Lee Deborah , Moore Erin , Cao Jianhong , McNevin John , Andrus Thomas , Zhu Haiying , Rubin Abbe , Zhu Tuofu , Celum Connie , McElrath M. Juliana
Published: September 2006

Candidate human immunodeficiency virus type 1 (HIV-1) vaccines designed to elicit T-cell immunity in HIV-1-uninfected persons are under investigation in phase I to III clinical trials. Little is known about how these vaccines impact the immunologic response postinfection in persons who break through despite vaccination. Here, we describe the first comprehensive characterization of HIV-specific T-cell immunity in vaccine study participants following breakthrough HIV-1 infection in comparison to 16 nonvaccinated subjects with primary HIV-1 infection. Whereas none of the 16 breakthrough infections possessed vaccine-induced HIV-1-specific T-cell responses preinfection, 85% of vaccinees and 86% of nonvaccinees with primary HIV-1 infection developed HIV-specific T-cell responses postinfection. Breakthrough subjects' T cells recognized 43 unique HIV-1 T-cell epitopes, of which 8 are newly described, and 25% were present in the vaccine. The frequencies of gamma interferon (IFN-γ)-secreting cells recognizing epitopes within gene products that were and were not encoded by the vaccine were not different (P= 0.64), which suggests that responses were not anamnestic. Epitopes within Nef and Gag proteins were the most commonly recognized in both vaccinated and nonvaccinated infected subjects. One individual controlled viral replication without antiretroviral therapy and, notably, mounted a novel HIV-specific HLA-C14-restricted Gag LYNTVATL-specific T-cell response. Longitudinally, HIV-specific T cells in this individual were able to secrete IFN-γ and tumor necrosis factor alpha, as well as proliferate and degranulate in response to their cognate antigenic peptides up to 5 years postinfection. In conclusion, a vaccinee's ability to mount an HIV-specific T-cell response postinfection is not compromised by previous immunization, since the CD8+T-cell responses postinfection are similar to those seen in vaccine-naïve individuals. Finding an individual who is controlling infection highlights the importance of comprehensive studies of breakthrough infections in vaccine trials to determine whether host genetics/immune responses and/or viral characteristics are responsible for controlling viral replication.
Adenovirus Core Protein pVII Is Translocated into the Nucleus by Multiple Import Receptor Pathways
Authors: Wodrich Harald , Cassany Aurelia , D'Angelo Maximiliano A. , Guan Tinglu , Nemerow Glen , Gerace Larry
Published: September 2006

Adenoviruses are nonenveloped viruses with an ∼36-kb double-stranded DNA genome that replicate in the nucleus. Protein VII, an abundant structural component of the adenovirus core that is strongly associated with adenovirus DNA, is imported into the nucleus contemporaneously with the adenovirus genome shortly after virus infection and may promote DNA import. In this study, we evaluated whether protein VII uses specific receptor-mediated mechanisms for import into the nucleus. We found that it contains potent nuclear localization signal (NLS) activity by transfection of cultured cells with protein VII fusion constructs and by microinjection of cells with recombinant protein VII fusions. We identified three NLS-containing regions in protein VII by deletion mapping and determined important NLS residues by site-specific mutagenesis. We found that recombinant protein VII and its NLS-containing domains strongly and specifically bind to importin α, importin β, importin 7, and transportin, which are among the most abundant cellular nuclear import receptors. Moreover, these receptors can mediate the nuclear import of protein VII fusions in vitro in permeabilized cells. Considered together, these data support the hypothesis that protein VII is a major NLS-containing adaptor for receptor-mediated import of adenovirus DNA and that multiple import pathways are utilized to promote efficient nuclear entry of the viral genome.
Vaccinia Virus G9 Protein Is an Essential Component of the Poxvirus Entry-Fusion Complex
Authors: Ojeda Suany , Domi Arban , Moss Bernard
Published: September 2006

The vaccinia virus G9R gene (VACWR087) encodes a protein of 340 amino acids with the following structural features that are conserved in all poxviruses: a site for N-terminal myristoylation, 14 cysteines, and a C-terminal transmembrane domain. Previous studies showed that G9 is one of eight proteins associated in a putative entry-fusion complex. Our attempt to isolate a mutant without the G9R gene was unsuccessful, suggesting that it is essential for virus replication. To further investigate its role, we constructed a recombinant vaccinia virus in which G9R is regulated by addition of an inducer. Induced G9 protein was associated with mature infectious virions and could be labeled with a membrane-impermeant biotinylation reagent, indicating surface exposure. Omission of inducer reduced the infectious-virus yield by about 1.5 logs; nevertheless, all stages of virus morphogenesis appeared normal and extracellular virions were present on the cell surface. Purified virions assembled without inducer had a specific infectivity of less than 5% of the normal level and a comparably small amount of G9, whereas their overall polypeptide composition, including other components of the entry-fusion complex, was similar to that of virions made in the presence of inducer or of wild-type virions. G9-deficient virions bound to cells, but penetration of cores into the cytoplasm and early viral RNA synthesis were barely detected, and cell-cell fusion was not triggered by low pH. Of the identified components of the multiprotein complex, G9 is the sixth that has been shown to be required for entry and membrane fusion.
Proteasome Inhibition Reveals that a Functional Preintegration Complex Intermediate Can Be Generated during Restriction by Diverse TRIM5 Proteins
Authors: Anderson Jenny L. , Campbell Edward M. , Wu Xiaolu , Vandegraaff Nick , Engelman Alan , Hope Thomas J.
Published: September 2006

The primate TRIM5 proteins constitute a class of restriction factors that prevent host cell infection by retroviruses from different species. The TRIM5 proteins act early after virion entry and prevent viral reverse transcription products from accumulating. We recently found that proteasome inhibitors altered the rhesus monkey TRIM5α restriction of human immunodeficiency virus type 1 (HIV-1), allowing reverse transcription products to accumulate even though viral infection remained blocked. To assess whether sensitivity to proteasome inhibitors was a common feature of primate TRIM5 proteins, we conducted a similar analysis of restriction mediated by owl monkey TRIM-cyclophilin A (CypA) or human TRIM5α. Similar to rhesus monkey TRIM5α restriction, proteasome inhibition prevented owl monkey TRIM-CypA restriction of HIV-1 reverse transcription, even though HIV-1 infection and the output of 2-LTR circles remained impaired. Likewise, proteasome inhibition alleviated human TRIM5α restriction of N-tropic murine leukemia virus reverse transcription. Finally, HIV-1 reverse transcription products escaping rhesus TRIM5α restriction by proteasome inhibition were fully competent for integration in vitro, demonstrating that TRIM5α likely prevents the viral cDNA from accessing chromosomal target DNA. Collectively, these data indicate that the diverse TRIM5 proteins inhibit retroviral infection in multiple ways and that inhibition of reverse transcription products is not necessary for TRIM5-mediated restriction of retroviral infection.
Identification of a 709-Amino-Acid Internal Nonessential Region within the Essential Conserved Tegument Protein (p)UL36 of Pseudorabies Virus
Authors: Böttcher Sindy , Klupp Barbara G. , Granzow Harald , Fuchs Walter , Michael Kathrin , Mettenleiter Thomas C.
Published: September 2006

Tegument proteins homologous to the essential herpes simplex virus type 1 UL36 gene product (p)UL36 are conserved throughout the
Herpesviridae
and constitute the largest herpesvirus-encoded proteins. So far, only limited information is available on their functions, which include complex formation with the (p)UL37 homologs via an N-terminal domain and a deubiquitinating activity in the extreme N terminus. For further analysis we constructed deletion mutants lacking 437, 784, 926, 1,046, 1,217, or 1,557 amino acids (aa) from the C terminus. While none of them supported replication of a pseudorabies virus (PrV) UL36 deletion mutant, a mutant polypeptide with an internal deletion from aa 2087 to 2795, which comprises a proline/alanine-rich region, fully complemented the lethal replication defect. Thus, our data indicate that the extreme C terminus of (p)UL36 fulfills an essential role in PrV replication, while a large internal portion of the C-terminal half of the protein is dispensable for replication in cell culture.
Nonlinear mechanics of cracks subjected to indentation
Authors: Selvadurai A P
Published: September 2006

The paper presents the application of a boundary element technique to study the behaviour of plane cracks that are located at corner regions of an elastic solid and open during indentation. In particular, the surfaces of the planes on which indentation takes place also exhibit Coulomb frictional responses and degradation in the friction angle with plastic energy dissipation. An incremental boundary element formulation, in which special singularity elements model the behaviour at the crack tip, is used to examine the crack problems. The methodology is applied to investigate the mode I stress intensity factor at the crack tip located at the base of a V-notch in a test specimen.Key words: indented cracks, boundary element modelling, Coulomb friction, stress intensity factors
Performance of waterstops at different temperatures
Authors: Mirza Jahangir , Tremblay Stéphane , Sebbagh Frédéric , Ballivy Gérard
Published: September 2006

Waterstops are installed in the body of concrete (to a depth of 30–60 cm) during the construction of hydraulic structures to minimize or eliminate all possible leaks at construction joints. However, they often deteriorate as a result of the thermal variations and hydrostatic pressure to which hydraulic structures are subjected over their lifetime. Since repairing or replacing deteriorated waterstops is very complicated and costly, they are normally left in place and new ones installed close by. Various new products and methods have now been developed for this purpose, but their performance is not well documented. Eighteen different products were initially chosen and subjected to a battery of laboratory tests (viscosity, chemical compatibility with water, and freeze–thaw cycling) at 23 °C, 15 °C, and 5 °C. Four of the 18 products were subsequently tested for permeability and apparent adherence in a specially designed setup. The test data showed that the waterstop that best fulfilled the criteria was a two-component polyurethane resin.Key words: adherence, chemical compatibility with water, different temperatures, hydraulic structures, permeability, viscosity, waterstops.
Factors affecting buildability of building designs
Authors: Wong Franky W.H , Lam Patrick T.I , Chan Edwin H.W , Wong Francis K.W
Published: September 2006

By identifying the succinct attributes of design, the abstract concept of buildability can be expressed in a more defined and tangible way for its improvements. In this research, factor analysis is used to expound the data obtained from a questionnaire survey on buildability attributes. Results show the first three out of nine key buildability factors for building designs are (i) allowing economic use of contractor resources, (ii) enabling design requirements to be easily visualized and coordinated by site staff, and (iii) enabling contractors to develop and adopt alternative construction details. Discriminant analysis has been used to identify significant differences amongst the respondent groups. At the project level, the findings give designers a better understanding of factors affecting the buildability of their outputs, thus enabling design solutions leading to more efficient and safe construction. At the industry level, the identified factors can contribute to sustainable development through the reduction of waste and the economic use of resources.Key words: design appraisal, buildability improvement, factor analysis, discriminant analysis.
Inelastic analysis of a reinforced concrete shear wall building according to the National Building Code of Canada 2005
Authors: Panneton M , Léger P , Tremblay R
Published: September 2006

An eight-storey reinforced concrete shear wall building located in Montréal and designed according to the 1995 National Building Code of Canada (NBCC) and the Canadian Standards Association standard CSA-A23.3-94 is studied to evaluate the impact of new requirements for inclusion in new editions of the NBCC and CSA-A23.3. Static and modal analyses were conducted according to the 2005 NBCC (draft 2003) and CSA-A23.3-04 (draft 4) procedures, and three-dimensional dynamic inelastic time history analysis was performed using three earthquake records. The building is braced by four flat shear walls and three cores. Various estimates of the fundamental period of vibration based on empirical expressions presented in the literature or structural models with different stiffness assumptions were examined. The analysis also permitted the study of the displacement and force demand on the lateral load resisting system. It was found that the base shear from the 2005 NBCC is 29% higher than the 1995 NBCC value when code empirical formulae are used for the fundamental period of vibration.Key words: building, shear wall, inelastic seismic response, NBCC, CSA-A23.3 design of concrete structures.
Modeling the effect of corrosion on bond strength at the steel–concrete interface with finite-element analysis
Authors: Amleh Lamya , Ghosh Alaka
Published: September 2006

This paper examines the basic influence of corrosion on bond strength at the steel–concrete interface and the associated slip and cracking. A nonlinear finite-element model is developed to account for the effect of corrosion on deterioration of the bond. Deterioration of the mechanical interaction between the corroding reinforcing steel and the concrete is modeled with the nonlinear finite-element program ABAQUS. The contact pressure normal to the steel–concrete interface is reduced when the concrete cracks, which occurs along with a decrease in the cross-sectional area of the steel bar and the decrease of the friction coefficient between the steel and the concrete. The loss of contact pressure and the decrease in the friction coefficient with the mass loss of steel bars are evaluated using pullout test specimens with different levels of rebar corrosion. Finally, the relationship between the loss of bond strength and the mass loss of the steel rebar is established. The model gives reasonably accurate predictions of bond strength for three independent experimental studies. Key words: bond, concrete, corrosion, mechanical interaction, reinforcing steels, slip, steel-concrete interface.
A targeted study on possible free ammonia inhibition of Nitrospira
Authors: Simm R A , Mavinic D S , Ramey W D
Published: September 2006

Batch tests were carried out with Nitrospira in mixed and pure cultures using concentrations of free ammonia widely believed to be inhibitory to nitrite-oxidizing organisms. The mixed culture batch tests were conducted with mixed liquor from a bench scale completely stirred tank reactor (CSTR) treating a synthetic wastewater having a low C:N ratio. Nitrospira were confirmed as the dominant nitrite oxidizers via RNA slot blotting. Nitrospira moscoviensis were used for the pure culture trials. The results from this study suggest that free ammonia (NH3-N) concentrations of up to 10 mg/L were not inhibitory to Nitrospira either in situ or in pure culture. Key words: fatty acid methyl esters, free ammonia inhibition, molecular methods, nitrification, nitrite, Nitrobacter, Nitrospira, nitrous acid, nitrous oxide, RNA slot blotting, wastewater treatment.
Recycling of calcium fluoride sludge as additive in the solidification–stabilization of fly ash
Authors: Kim Young-Ju , Qureshi Tahir Imran
Published: September 2006

Recycling of calcium fluoride sludge as additive in the solidification of fly ash with cement or calcium hydroxide was studied. Fly ash solidified with calcium hydroxide and calcium fluoride sludge failed to achieve a target compressive strength of 200 kN/m2 over a reaction period of 10 d. White Portland cement had produced good results in comparison with high early strength cement or Portland cement. Compressive strength of the material solidified with white Portland cement was found to be 39% higher than the material solidified with Portland cement. The optimum mix was found to be that with 80% fly ash, 5% white Portland cement, and 15% calcium fluoride sludge. Heavy metal ions and fluorides found in the leachate of stabilized matrix were below the discharge limit. Recycling of calcium fluoride as additive reduced the cost of solidification of fly ash to 41% while helping to transform hazardous sludge into an environmentally friendly material. Key words: solidification, additive, cement, lime, recycling.
Behavior of complexing ligands during coagulationflocculation by ferric chloride: A comparative study between sewage water and an engineered colloidal model
Authors: El Samrani A G , Ouaini N , Lartiges B S , Kazpard V , Ibrik A , Saad Z
Published: September 2006

In this work, a comparative study is conducted on sewage as an example of a natural water system and on an engineered colloidal model. The two water systems were treated by coagulationflocculation with ferric chloride. Affinity of phosphate and sulfate used as complexing ligands to coagulant species was studied within the two systems. Aggregation dynamics were followed with jar tests and laser diffraction. Sulfate and phosphate were determined by using ion chromatography. Atomic absorption spectroscopy (AAS) and transmission electron microscopy coupled with energy dispersive X-ray spectrometry were used for elemental and ionic analysis of P, Fe, S, SO4, and PO4 in supernatants and sediments. Investigation of ligands revealed that phosphate and iron(III) species make strong complexes that enhance aggregation velocity, whereas sulfate is weakly complexed with iron(III) species. It can be released in suspensions without any consequences on aggregation dynamics.Key words: aggregation, coagulant species, complexing ligands, coagulation, flocculation.
Characterization of Au and Pd nanoparticles by high-temperature TGA–MS
Authors: Jia Weijuan , McLachlan Jessica , Xu Jiayan , Tadayyon Seyed M , Norton Peter R , Eichhorn S Holger
Published: September 2006

Noble metal nanoparticles (NPs) prepared by a surfactant-free single-phase solution method have been proposed to contain fewer ionic contaminants than similar NPs prepared by a two-phase method. Reported herein is the possible contamination of Au and Pd NPs, prepared by a surfactant-free single-phase method, with Li2CO3 and other Li salts. High-temperature thermal gravimetric analysis measurements coupled with mass spectrometry (TGA–MS) up to 1100 °C were employed to determine the relative amounts of ionic contaminants since protecting thiolate groups and inorganic contaminants were removed in separate weight loss events. Assignment of the different weight loss events was supported by MS analysis of the evolved gases. TGA–MS also revealed the presence of larger amounts of oxidized sulfur species in the Pd NPs. High-resolution transmission electron microscopy (HRTEM), UV–vis, IR, elemental analysis (EA), and X-ray photoelectron spectroscopy (XPS) measurements complemented the characterization of the NPs. The amount of ionic contaminants crucially depended on the workup conditions, and quenching of the reaction mixture with ethanol was found to be essential for the formation of Li2CO3. Workup procedures that avoid the formation of TGA–MS detectable ionic contaminants are proposed along with purification steps for contaminated NPs.Key words: gold, palladium, nanoparticles, TGA, mass spectrometry, ionic contamination.
AN IMPROVEMENT TO THE FOURIER SERIES METHOD FOR INVERSION OF LAPLACE TRANSFORMS APPLIED TO ELASTIC AND VISCOELASTIC WAVES
Authors: LAVERTY RICHARD R. , GAZONAS GEORGE A.
Published: September 2006

A parametric study of composite strips leads to systems of partial differential equations, coupled through interface conditions, that are naturally solved in Laplace transform space. Because of the complexity of the solutions in transform space and the potential variations due to geometry and materials, a systematic approach to inversion is necessarily numerical. The Dubner-Abate-Crump (DAC) algorithm is the standard in such problems and is implemented. The presence of discontinuous wavefronts in the problems considered leads to Gibbs phenomenon; which, in turn, overestimates the values of maximum stress. These errors are mitigated by use of Lanczos' σ-factors, which combine naturally with the DAC algorithm.
PREDICTIONS OF ANISOTROPY AFFECTED FATIGUE CRACK PROPAGATION PATHS IN PURE ALUMINUM SHEETS
Authors: IRAWAN YUDY SURYA , HAGIWARA YOSHIHIKO , OHYA SHIN-ICHI
Published: September 2006

The anisotropy affected fatigue cracks that propagate on the rolling direction as mixed mode I-II cracks in CCT-specimens of pure aluminum sheets were predicted using Finite Element Method. The crack growth paths were estimated based on the value of mode I, II, total stress intensity factor range, and the ratio of mode II and mode I energy release rate of the crack. The predictions of crack propagation paths in no-hole specimens and 6-type holed specimens with 0, 30, 45 and 60 degrees of rolling direction towards loading direction were performed and the results were coincided with experimental results.
Interactions Between Mosquito Larvae and Species that Share the Same Trophic Level
Authors: Blaustein Leon , Chase Jonathan M.
Published: September 2006

Ecological theory predicts, and empirical research shows, that species sharing the same trophic level as a target species (hereafter controphic species) can have large direct and indirect effects on the target species by sharing resources and/or by serving as alternative prey to predators. Yet, the roles of controphic species of mosquito larvae in affecting mosquito populations have received little attention. Published empirical evidence, although scarce, suggests that controphic species such as zooplankton and anuran larvae compete with mosquito larvae, can positively affect mosquito larvae by consuming bacteria that are pathogenic to mosquito larvae, reduce predation on mosquito larvae by serving as alternative prey, and ultimately cause increased predation on mosquito larvae by causing a numerical response in the predator. We conclude that more extensive theoretical and empirical studies in elucidating the roles of controphic species will better allow us to predict mosquito population dynamics and allow for better management of mosquitoes.
Specificity and Regulation of Renal Sulfate Transporters
Authors: Markovich Daniel , Aronson Peter S.
Published: September 2006

Sulfate is essential for normal cellular function. The kidney plays a major role in sulfate homeostasis. Sulfate is freely filtered and then undergoes net reabsorption in the proximal tubule. The apical membrane Na+/sulfate cotransporter NaS1 (SLC13A1) has a major role in mediating proximal tubule sulfate reabsorption, as demonstrated by the findings of hyposulfatemia and hypersulfaturia in Nas1-null mice. The anion exchanger SAT1 (SLC26A1), the founding member of the SLC26 sulfate transporter family, mediates sulfate exit across the basolateral membrane to complete the process of transtubular sulfate reabsorption. Another member of this family, CFEX (SLC26A6), is present at the apical membrane of proximal tubular cells. It also can transport sulfate by anion exchange, which probably mediates backflux of sulfate into the lumen. Knockout mouse studies have demonstrated a major role of CFEX as an apical membrane Cl−/oxalate exchanger that contributes to NaCl reabsorption in the proximal tubule. Several additional SLC26 family members mediate sulfate transport and show some level of renal expression (e.g., SLC26A2, SLC26A7, SLC26A11). Their roles in mediating renal tubular sulfate transport are presently unknown. This paper reviews current data available on the function and regulation of three sulfate transporters (NaS1, SAT1, and CFEX) and their physiological roles in the kidney.
Alterations in cardiac contractile performance and sarcoplasmic reticulum function in sucrose-fed rats is associated with insulin resistance
Authors: Vasanji Zainisha , Cantor Elliott J. F. , Juric Danijel , Moyen Mellissa , Netticadan Thomas
Published: September 2006

Diabetes mellitus (DM) causes the development of a specific cardiomyopathy that results from the metabolic derangements present in DM and manifests as cardiac contractile dysfunction. Although myocardial dysfunction in Type 1 DM has been associated with defects in the function and regulation of the sarcoplasmic reticulum (SR), very little is known about SR function in Type 2 DM. Accordingly, this study examined whether abnormalities in cardiac contractile performance and SR function occur in the prestage of Type 2 DM (i.e., during insulin resistance). Sucrose feeding was used to induce whole body insulin resistance, whereas cardiac contractile performance was assessed by echocardiography and SR function was measured by SR calcium (Ca2+) uptake. Sucrose-fed rats exhibited hyperinsulinemia, hyperglycemia, and hyperlipidemia relative to control rats. Serial echocardiographic assessments in the sucrose-fed rats revealed early abnormalities in diastolic function followed by late systolic dysfunction and concurrent alterations in myocardial structure. The hearts of the 10-wk sucrose-fed rats showed depressed SR function demonstrated by a significant reduction in SR Ca2+ uptake. The decline in SR Ca2+ uptake was associated with a significant decrease in the cAMP-dependent protein kinase and Ca2+/calmodulin-dependent protein kinase II-mediated phosphorylation of phospholamban. The results show that abnormalities in cardiac contractile performance and SR function occur at an insulin-resistant stage before the manifestation of overt Type 2 DM.
Analysis of DHE-derived oxidation products by HPLC in the assessment of superoxide production and NADPH oxidase activity in vascular systems
Authors: Fernandes Denise C. , Wosniak João , Pescatore Luciana A. , Bertoline Maria A. , Liberman Marcel , Laurindo Francisco R. M. , Santos Célio X. C.
Published: September 2006

Dihydroethidium (DHE) is a widely used sensitive superoxide (O2•−) probe. However, DHE oxidation yields at least two fluorescent products, 2-hydroxyethidium (EOH), known to be more specific for O2•−, and the less-specific product ethidium. We validated HPLC methods to allow quantification of DHE products in usual vascular experimental situations. Studies in vitro showed that xanthine/xanthine oxidase, and to a lesser degree peroxynitrite/carbon dioxide system led to EOH and ethidium formation. Peroxidase/H2O2 but not H2O2 alone yielded ethidium as the main product. In vascular smooth muscle cells incubated with ANG II (100 nM, 4 h), we showed a 60% increase in EOH/DHE ratio, prevented by PEG-SOD or SOD1 overexpression. We further validated a novel DHE-based NADPH oxidase assay in vascular smooth muscle cell membrane fractions, showing that EOH was uniquely increased after ANG II. This assay was also adapted to a fluorescence microplate reader, providing results in line with HPLC results. In injured artery slices, shown to exhibit increased DHE-derived fluorescence at microscopy, there was ∼1.5- to 2-fold increase in EOH/DHE and ethidium/DHE ratios after injury, and PEG-SOD inhibited only EOH formation. We found that the amount of ethidium product and EOH/ethidium ratios are influenced by factors such as cell density and ambient light. In addition, we indirectly disclosed potential roles of heme groups and peroxidase activity in ethidium generation. Thus HPLC analysis of DHE-derived oxidation products can improve assessment of O2•− production or NADPH oxidase activity in many vascular experimental studies.
Multiparameter metabolic analysis reveals a close link between attenuated mitochondrial bioenergetic function and enhanced glycolysis dependency in human tumor cells
Authors: Wu Min , Neilson Andy , Swift Amy L. , Moran Rebecca , Tamagnine James , Parslow Diane , Armistead Suzanne , Lemire Kristie , Orrell Jim , Teich Jay , Chomicz Steve , Ferrick David A.
Published: September 2006

Increased conversion of glucose to lactic acid associated with decreased mitochondrial respiration is a unique feature of tumors first described by Otto Warburg in the 1920s. Recent evidence suggests that the Warburg effect is caused by oncogenes and is an underlying mechanism of malignant transformation. Using a novel approach to measure cellular metabolic rates in vitro, the bioenergetic basis of this increased glycolysis and reduced mitochondrial respiration was investigated in two human cancer cell lines, H460 and A549. The bioenergetic phenotype was analyzed by measuring cellular respiration, glycolysis rate, and ATP turnover of the cells in response to various pharmacological modulators. H460 and A549 cells displayed a dependency on glycolysis and an ability to significantly upregulate this pathway when their respiration was inhibited. The converse, however, was not true. The cell lines were attenuated in oxidative phosphorylation (OXPHOS) capacity and were unable to sufficiently upregulate mitochondrial OXPHOS when glycolysis was disabled. This observed mitochondrial impairment was intimately linked to the increased dependency on glycolysis. Furthermore, it was demonstrated that H460 cells were more glycolytic, having a greater impairment of mitochondrial respiration, compared with A549 cells. Finally, the upregulation of glycolysis in response to mitochondrial ATP synthesis inhibition was dependent on AMP-activated protein kinase activity. In summary, our results demonstrate a bioenergetic phenotype of these two cancer cell lines characterized by increased rate of glycolysis and a linked attenuation in their OXPHOS capacity. These metabolic alterations provide a mechanistic explanation for the growth advantage and apoptotic resistance of tumor cells.