bluuebunny/crossref_metadata_2025 huggingface dataset:
Study of the miscibility and mechanical properties of NBR/HNBR blends
Authors: Hussein Ibnelwaleed A. , Chaudhry Rehan A. , Abu Sharkh Basel F.
Published: November 2004

In this study, hydrogenated acrylonitrile butadiene rubber (HNBR, ZETPOL‐2010L) and nitrile butadiene rubber (NBR, NIPOL‐DN4555) were blended at different ratios in a Haake melt blender at 130°C. The HNBR and the NBR were of very similar acrylonitrile content and Mooney viscosity. The melt miscibility and solid‐state properties were investigated by rheological, thermal, and mechanical testing and scanning electron microscopy (SEM) techniques. The dynamic viscosity of the blends followed the log‐additivity rule, while the flow activation energy closely followed the inverse additivity rule. On the other hand, the storage modulus showed synergistic effects at all compositions, suggesting the presence of emulsion morphology at both ends of the composition range. For the 50/50 HNBR/NBR blend, the SEM micrographs suggest a uniform elongated structure. The thermal analysis showed the presence of two glass transitions, representing the pure components, at all blend ratios, suggesting the absence of segmental miscibility of the blends. The small‐strain mechanical properties such as tensile modulus and yield stress followed linear additivity. However, HNBR and HNBR‐rich blends were observed to strain harden at a rate higher than that of NBR. Induced crystallization of HNBR was suggested to be the reason for the strain hardening. The different rheological, thermal, and mechanical testing techniques agree in suggesting that the structurally similar HNBR and NBR are not thermodynamically miscible but mechanically compatible. Polym. Eng. Sci. 44:2346–2352, 2004. © 2004 Society of Plastics Engineers.
Rapid and selective identification of molecular species in phosphatidylcholine and sphingomyelin by conditional neutral loss scanning and MS3
Authors: Houjou Toshiaki , Yamatani Kotoko , Nakanishi Hiroki , Imagawa Masayoshi , Shimizu Takao , Taguchi Ryo
Published: November 2004

Analyses of molecular species of phospholipids containing choline (Ch), such as phosphatidylcholine (PC) and sphingomyelin (SM), are reported. Neutral loss scanning was applied for the selective detection of these lipids using a quadrupole‐linear ion trap mass spectrometer. By using ammonium formate as an elution buffer, both PC and SM were detected as [M+HCOO]− ions in the negative ion mode. Upon collisional activation, the [M+HCOO]− adduct ions underwent facile elimination of HCO2, to yield an ion which, in turn, readily underwent collisional‐induced dissociation (CID) to eliminate CH3 to yield an [M–CH3]− ion. By selecting the proper conditions for scanning for neutral loss of 60 Da (HCO2+CH3), SM species were identified separately from PCs. Further, by selection of this [M–CH3]− ion as the precursor ion, the identities of the fatty acyl chains of PC species can be effectively obtained by MS3 experiments. Furthermore, by the MS3 analyses of [M–CH3]− specifically obtained from SM molecules, identification of sphingosine or sphinganine derivatives and their N‐acyl species can also be effectively obtained. This systematic analysis of PCs and SMs by conditional neutral loss scanning, with subsequent analyses by MS3, using a linear ion trap mass spectrometer in the negative ion mode, appears to be a very effective and sensitive method. Further, MS/MS in the positive ion mode at relatively low collision energy was also effective for the identification of positional specificities in individual molecular PC species from their lysoPC‐related fragments. The present paper deals only with qualitative identification of individual molecular species, and the related quantitative studies are now underway. Copyright © 2004 John Wiley & Sons, Ltd.
Influence analysis for linear mixed‐effects models
Authors: Demidenko Eugene , Stukel Therese A.
Published: November 2004

In this paper, we extend several regression diagnostic techniques commonly used in linear regression, such as leverage, infinitesimal influence, case deletion diagnostics, Cook's distance, and local influence to the linear mixed‐effects model. In each case, the proposed new measure has a direct interpretation in terms of the effects on a parameter of interest, and collapses to the familiar linear regression measure when there are no random effects. The new measures are explicitly defined functions and do not necessitate re‐estimation of the model, especially for cluster deletion diagnostics. The basis for both the cluster deletion diagnostics and Cook's distance is a generalization of Miller's simple update formula for case deletion for linear models. Pregibon's infinitesimal case deletion diagnostics is adapted to the linear mixed‐effects model. A simple compact matrix formula is derived to assess the local influence of the fixed‐effects regression coefficients. Finally, a link between the local influence approach and Cook's distance is established. These influence measures are applied to an analysis of 5‐year Medicare reimbursements to colon cancer patients to identify the most influential observations and their effects on the fixed‐effects coefficients. Copyright © 2004 John Wiley & Sons, Ltd.
Eine Vollglaskonstruktion – Möglichkeiten und Probleme
Authors: Werner Frank
Published: November 2004

Vorgestellt werden der Entwurf und die Realisierung einer zweigeschossigen Glaskonstruktion, die konsequent auf Stahltrag‐ und Sicherungselemente verzichtet. Stützen und Träger sollten die Transparenz der Hülle nicht beeinträchtigen und sind deshalb aus Glas. Die Gesamttragwirkung wird über die Verklebungen der Bauelemente untereinander und mit dem angrenzenden Baukörper gewährleistet. Die Einleitung örtlicher Lasten in den Rand der Wandscheiben ohne zusätzliche konstruktive Elemente wird numerisch und experimentell untersucht.

Dieser Experimentalbau dient dazu, konstruktive Lösungen zu testen und fundierte Aussagen zur Nutzung und zu den Kosten zu erhalten.
Decision‐making capacity and disaster research
Authors: Rosenstein Donald L.
Published: November 2004

The extent to which victims of a disaster are able to make capacitated and voluntary decisions to enroll in research is an important and virtually unexplored question. Although there are no compelling data to suggest that experiencing a severe trauma, in and of itself, renders all or even most individuals incapable of making autonomous decisions, the assessment of decision‐making capacity (DMC) for research participation warrants serious consideration. This paper provides a framework for and procedural approach to the assessment of DMC in research with individuals exposed to disaster. Particular attention is paid to the implementation of additional safeguards to protect subjects who are vulnerable by virtue of impaired DMC. Recommendations are offered to clinical investigators, ethical review boards, and policymakers with regard to the design, review, and conduct of research in the aftermath of disaster.
Interannual variability of the upper ocean carbon cycle at station ALOHA near Hawaii
Authors: Brix Holger , Gruber Nicolas , Keeling Charles D.
Published: November 2004

We investigate interannual variability of the upper ocean carbon cycle in the subtropical North Pacific on the basis of a 14‐year time series (1988–2002) of carbon parameters from Station ALOHA, the site of the U.S. JGOFS Hawaii Ocean Time series program (HOT). The data reveal substantial interannual variability in near‐surface concentrations of dissolved inorganic carbon normalized to constant salinity (sDIC, peak‐to‐peak amplitude of ±4 μmol kg−1), computed ocean surface partial pressure of CO2 (pCO2, ±6 ppm), and the 13C/12C ratio of DIC (±0.07‰). A strong anticorrelation (r = −0.50) between interannual anomalies in sea‐surface temperature (SST) and sDIC is found, which tends to suppress the correlation of either of these properties with pCO2. In contrast, no significant correlation (p < 0.05) is found between anomalies of the 13C/12C ratio of DIC and any other parameter. A diagnostic box model analysis reveals that interannual variability of near‐surface ocean sDIC is driven by air‐sea gas exchange, net community production, and lateral transport. In warmer than normal years the seasonal carbon cycle tends to be weakened, with a sDIC reduction in the mixed layer caused by diminished gas exchange and lateral transport outweighing the effect of less intense sDIC removal by net community production. This explains the observed anticorrelation between SST and sDIC. Interannual (peak‐to‐peak) variability of air‐sea gas exchange (±0.4 mol m−2 yr−1, i.e., 40% of the annual mean value) is primarily governed by strongly covarying changes in SST and wind speeds. Net community production varies interannually by up to ±0.9 mol m−2 yr−1 (39%) and tends to be associated with changes in horizontal transport. Less than 20% of the interannual variance in sDIC near Hawaii can be explained by the Pacific Decadal Oscillation (PDO), and an even smaller fraction (less than 5%) by the El Niño‐Southern Oscillation (ENSO). Because SST variations over a sizable fraction of the North Pacific subtropical gyre vary in concert with those at Station ALOHA, it is plausible that air‐sea fluxes in this region vary also synchronously, resulting in a variability of the atmospheric CO2 sink strength of the North Pacific subtropical gyre of up to ±0.2 PgC yr−1.
Variability of the aerosol number size distribution in Beijing, China: New particle formation, dust storms, and high continental background
Authors: Wehner B. , Wiedensohler A. , Tuch T. M. , Wu Z. J. , Hu M. , Slanina J. , Kiang C. S.
Published: November 2004

Continuous measurements of aerosol number size distributions from 3 nm to 10 μm have been first performed within the city area of Beijing since March 2004. Size distributions of the first 45 measurement days (March 05 to April 18, 2004) were investigated in terms of their high variability. Two dust storm events were observed indicated by high number concentrations greater than 1 μm and mass concentrations around 1 mg m−3. Continental highly polluted air was observed during 12 days indicated by a number peak in the accumulation mode range, and submicrometer volume concentrations above 150 μm3 cm−3 were observed. Newly formed particles with more than 100,000 cm−3 were observed on 25 days when the particle surface area concentration drops below a critical value (100–2000 μm2 cm−3) because of clean air from the north. Measurements show only a slight growth (∼1 nm h−1) of the particles indicating that they are produced within the city area of Beijing.
Nontidal oceanic excitation of nutation and diurnal/semidiurnal polar motion revisited
Authors: Brzeziński Aleksander , Ponte Rui M. , Ali Ahmed H.
Published: November 2004

A new 7.5‐yearlong time series of hourly estimates of nontidal ocean angular momentum, representing signals forced by 6‐hourly surface atmospheric pressure and winds including those forced by the atmospheric tides, is used to assess the influence of a dynamic ocean on nutation and on diurnal and semidiurnal polar motions. Using available atmospheric angular momentum data, we estimate the total excitation by the dynamically coupled atmosphere‐ocean system and compare it to that of the atmosphere alone. In case of the retrograde diurnal excitation, significant contributions are found for the retrograde annual, prograde annual, and prograde semiannual nutations and for the constant offset of the celestial pole. For the prograde annual nutation corresponding to the retrogradeS1component of excitation, the atmosphere‐ocean model does not improve the agreement with VLBI observations beyond that obtained using the atmosphere model alone. We also considered separately an irregular contribution to nutation, which can excite both the free core nutation signal and other broadband variability. The irregular component of the atmosphere‐ocean model is found to be significantly correlated with that derived from the atmosphere model. For the prograde diurnal component of excitation, which contributes to polar motion, the only nonnegligible effect (∼9 microarc seconds (μas)) is associated with the progradeS1harmonic. The only significant component of the semidiurnal excitation found is the elliptical oscillationS2in polar motion with major semiaxis of 9 μas, but this estimate is uncertain because the 6‐hourly atmospheric fields used to force the ocean do not properly resolve semidiurnal signals. In any case, this contribution is much smaller than that from theS2ocean gravitational tide.
Impact of absorbing aerosol on precipitation: Dynamic aspects in association with convective available potential energy and convective parameterization closure and dependence on aerosol heating profile
Authors: Chung Chul Eddy , Zhang Guang J.
Published: November 2004

The Indian Ocean Experiment (INDOEX), conducted from 1995 to 2000 to document aerosols in south Asia during winter monsoon season, revealed the existence of a layer of highly absorbing aerosols in the lower troposphere. The observed aerosol has one of the two distinctly different vertical distributions: (1) aerosols concentrated in the planetary boundary layer (PBL) below 1.5 km and (2) elevated aerosol profile peaking around 3 km. Here we provide the dynamical basis for understanding the direct effects of absorbing aerosols on the large‐scale precipitation and the role of the aerosol vertical distribution. This was done through a series of the south Asian aerosol experiments with the National Center for Atmospheric Research Community Climate Model (CCM3), together with different convection parameterization closures, and the Community Atmospheric Model (CAM2). It is found that the land surface temperature underneath the aerosol layer is sensitive to the aerosol vertical distribution: The lifted layer of aerosols results in a significant cooling of the underlying land surface, while the PBL profile makes very little cooling. The mechanism of the aerosol effect on precipitation distribution is investigated by examining the correspondence between the aerosol heating profile, changes of precipitation, and the atmospheric convective instability. The direct aerosol heating of the near‐surface air increases the convective available potential energy (CAPE), whereas the heating above the boundary layer decreases CAPE. Meanwhile, the regionally concentrated low‐level aerosol heating tends to cause large‐scale rising motion over time, which increases CAPE by decreasing the midlevel temperature. The net CAPE change is small for the lifted profile (i.e., profile elevated above PBL) because the CAPE increase by the midlevel cooling is counteracted by the CAPE decrease through the direct haze heating above the PBL. The precipitation increase averaged over the aerosol area is much larger when the PBL profile is used than when the lifted profile is used in the CCM3 with a CAPE‐based convective parameterization closure. The sensitivity of the aerosol effect to convective parameterization closure is tested using a new closure, which is based on the environmental contribution to CAPE (CAPEe). It is shown that when this closure is used in CCM3, the precipitation increase averaged over the aerosol area is small regardless of the vertical profile. This is because the direct heating of either profile decreases CAPEe, opposing the CAPEe increase by the midlevel cooling.
Mapping annual mean ground‐level PM2.5 concentrations using Multiangle Imaging Spectroradiometer aerosol optical thickness over the contiguous United States
Authors: Liu Yang , Park Rokjin J. , Jacob Daniel J. , Li Qinbin , Kilaru Vasu , Sarnat Jeremy A.
Published: November 2004

We present a simple approach to estimating ground‐level fine particulate matter (PM2.5, particles smaller than 2.5 μm in diameter) concentrations by applying local scaling factors from a global atmospheric chemistry model (GEOS‐CHEM with GOCART dust and sea salt data) to aerosol optical thickness (AOT) retrieved by the Multiangle Imaging Spectroradiometer (MISR). The resulting MISR PM2.5 concentrations are compared with measurements from the U.S. Environmental Protection Agency's (EPA) PM2.5 compliance network for the year 2001. Regression analyses show that the annual mean MISR PM2.5 concentration is strongly correlated with EPA PM2.5 concentration (correlation coefficient r = 0.81), with an estimated slope of 1.00 and an insignificant intercept, when three potential outliers from Southern California are excluded. The MISR PM2.5 concentrations have a root mean square error (RMSE) of 2.20 μg/m3, which corresponds to a relative error (RMSE over mean EPA PM2.5 concentration) of approximately 20%. Using simulated aerosol vertical profiles generated by the global models helps to reduce the uncertainty in estimated PM2.5 concentrations due to the changing correlation between lower and upper tropospheric aerosols and therefore to improve the capability of MISR AOT in estimating surface‐level PM2.5 concentrations. The estimated seasonal mean PM2.5 concentrations exhibited substantial uncertainty, particularly in the west. With improved MISR cloud screening algorithms and the dust simulation of global models, as well as a higher model spatial resolution, we expect that this approach will be able to make reliable estimation of seasonal average surface‐level PM2.5 concentration at higher temporal and spatial resolution.
A study of mesoscale gravity waves over the North Atlantic with satellite observations and a mesoscale model
Authors: Wu Dong L. , Zhang Fuqing
Published: November 2004

Satellite microwave data are used to study gravity wave properties and variabilities over the northeastern United States and the North Atlantic in the December–January periods. The gravity waves in this region, found in many winters, can reach the stratopause with growing amplitude. The Advanced Microwave Sounding Unit‐A (AMSU‐A) observations show that the wave occurrences are correlated well with the intensity and location of the tropospheric baroclinic jet front systems. To further investigate the cause(s) and properties of the North Atlantic gravity waves, we focus on a series of wave events during 19–21 January 2003 and compare AMSU‐A observations to simulations from a mesoscale model (MM5). The simulated gravity waves compare qualitatively well with the satellite observations in terms of wave structures, timing, and overall morphology. Excitation mechanisms of these large‐amplitude waves in the troposphere are complex and subject to further investigations.
Use of hydraulic tests at different scales to characterize fracture network properties in the weathered‐fractured layer of a hard rock aquifer
Authors: Maréchal J. C. , Dewandel B. , Subrahmanyam K.
Published: November 2004

The hydrodynamic properties of the weathered‐fractured layer of a hard‐rock pilot watershed in a granitic terrain are characterized using hydraulic tests at different scales. The interpretation of numerous slug tests leads us to characterize the statistical distribution of local permeabilities in the wells. The application of flowmeter profiles during injection tests determines the vertical distribution of conductive fracture zones and their permeabilities. It appears that the extension of the most conductive part of the weathered‐fractured layer is limited down to 35 m depth. The partition of drainage porosity between blocks (90%) and fractures (10%) is determined thanks to the interpretation of pumping tests using a double‐porosity model. The application of anisotropic and single‐fracture analytical solutions on pumping test data allows us to determine, respectively, the degree of anisotropy of permeability (Kr/Kz = 10) and the radius (4–16 m) of the horizontal conductive fractures crossed by the wells. Two different scales of fracture networks are identified: the primary fracture network (PFN), which affects the matrix on a decimeter scale by contributing to an increase in the permeability and storage capacity of the blocks, and the secondary fracture network (SFN), which affects the blocks at the borehole scale. SFN is composed of two sets of fractures. The main set of horizontal fractures is responsible for the subhorizontal permeability of the weathered‐fractured layer. A second set of less permeable subvertical fractures insures the connectivity of the aquifer at the borehole scale. The good connectivity of fracture networks is shown by fractional‐dimension flow solutions. The absence of scale effect in the study area suggests that the hydraulic conductivity at the borehole scale is laterally homogeneous. Finally, the analysis and synthesis of the hydrodynamic properties allow us to propose a comprehensive hydrodynamic model of the fractured‐weathered layer. Many geological and hydrogeological indicators suggest that a continuous and laterally homogeneous weathering process is responsible for the origin of the fractures and permeability encountered in the aquifer. These results confirm the major role played by weathering in the origin of fractures and on resulting hydrodynamic parameters in the shallow part of hard‐rock aquifers.
Platelet–cancer interactions: mechanisms and pharmacology of tumour cell‐induced platelet aggregation
Authors: Jurasz Paul , Alonso‐Escolano David , Radomski Marek W
Published: November 2004

During haematogenous metastasis, cancer cells migrate to the vasculature and interact with platelets resulting in tumour cell‐induced platelet aggregation (TCIPA). We review:

The biological and clinical significance of TCIPA;

Molecular mechanisms involved in platelet aggregation by cancer cells;

Strategies for pharmacological regulation of these interactions.

We conclude that pharmacological regulation of platelet–cancer cell interactions may reduce the impact of TCIPA on cancer biology.

British Journal of Pharmacology (2004) 143, 819–826. doi:10.1038/sj.bjp.0706013
KATP‐channel‐induced vasodilation is modulated by the Na,K‐pump activity in rabbit coronary small arteries
Authors: Glavind‐Kristensen Marianne , Matchkov Vladimir , Hansen Vibeke Brogaard , Forman Axel , Nilsson Holger , Aalkjær Christian
Published: November 2004

The purpose of the study was to evaluate the importance of the Na,K‐pump in relaxations induced by KATP‐channel openers in rabbit coronary small arteries.

Arterial segments were mounted in myographs for recording of isometric tension. Whole‐cell patch clamp was used to assess KATP‐channel currents in isolated smooth muscle cells from the arteries.

In arteries preconstricted with the thromboxane A2 analogue U46619 pinacidil and cromakalim induced concentration‐dependent relaxations. In arteries preconstricted with potassium (124 mM) only high concentrations of pinacidil had a small relaxant effect.

In arteries preconstricted with U46619 pinacidil‐induced relaxations were unaffected by pretreatment with Nω‐nitro‐L‐arginine (L‐NNA) and only slightly reduced after mechanical removal of the endothelium.

Pinacidil induced relaxations were not significantly affected by 1 μM glibenclamide. However, the relaxations were partly inhibited in potassium‐free media and by 1 μM ouabain.

In contrast, the concentration‐dependent relaxation to cromakalim was partly blocked by 1 μM glibenclamide and partly by 1 μM ouabain and when both drugs were present the inhibition increased.

Ouabain (1 μM) and glibenclamide (1 μM) each partly inhibited an ATP‐sensitive current induced by pinacidil and cromakalim. In the presence of both inhibitors a greater inhibition was seen. When the solution in the patch pipette was sodium‐free the current was reduced and ouabain had no effect.

The study suggests that the relaxation to cromakalim and most likely pinacidil is mediated through opening of KATP channels. Inhibition of the Na,K‐pump, however, may change the local environment for the KATP channels (i.e. increases the ATP/ADPratio and/or decreases the transmembrane potassium gradient), which partly prevents the activation of the KATP‐channel current.

British Journal of Pharmacology (2004) 143, 872–880. doi:10.1038/sj.bjp.0706016
Selective inhibition of sodium–calcium exchanger by SEA‐0400 decreases early and delayed afterdepolarization in canine heart
Authors: Nagy Zsolt A , Virág László , Tóth András , Biliczki Péter , Acsai Károly , Bányász Tamás , Nánási Péter , Papp Julius Gy , Varró András
Published: November 2004

The sodium–calcium exchanger (NCX) was considered to play an important role in arrhythmogenesis under certain conditions such as heart failure or calcium overload. In the present study, the effect of SEA‐0400, a selective inhibitor of the NCX, was investigated on early and delayed afterdepolarizations in canine ventricular papillary muscles and Purkinje fibres by applying conventional microelectrode techniques at 37°C. The amplitude of both early and delayed afterdepolarizations was markedly decreased by 1 μM SEA‐0400 from 26.6±2.5 to 14.8±1.8 mV (n=9, P<0.05) and from 12.5±1.7 to 5.9±1.4 mV (n=3, P<0.05), respectively. In enzymatically isolated canine ventricular myocytes, SEA‐0400 did not change significantly the L‐type calcium current and the intracellular calcium transient, studied using the whole‐cell configuration of the patch‐clamp technique and Fura‐2 ratiometric fluorometry. It is concluded that, through the reduction of calcium overload, specific inhibition of the NCX current by SEA‐0400 may abolish triggered arrhythmias.

British Journal of Pharmacology (2004) 143, 827–831. doi:10.1038/sj.bjp.0706026
Evaluation of the diffusion coefficient for controlled release of oxytetracycline from alginate/chitosan/poly(ethylene glycol) microbeads in simulated gastrointestinal environments
Authors: Cruz Maria C. Pinto , Ravagnani Sergio P. , Brogna Fabio M. S. , Campana Sérgio P. , Triviño Galo Cardenas , Lisboa Antonio C. Luz , Mei Lucia H. Innocentini
Published: November 2004

Diffusion studies of OTC (oxytetracycline) entrapped in microbeads of calcium alginate, calcium alginate coacervated with chitosan (of high, medium and low viscosity) and calcium alginate coacervated with chitosan of low viscosity, covered with PEG [poly(ethylene glycol) of molecular mass 2, 4.6 and 10 kDa, were carried out at 37±0.5 °C, in pH 7.4 and pH 1.2 buffer solutions – conditions similar to those found in the gastrointestinal system. The diffusion coefficient, or diffusivity (D), of OTC was calculated by equations provided by Crank [(1975) Mathematics in Diffusion, p. 85, Clarendon Press, Oxford] for diffusion, which follows Fick's [(1855) Ann. Physik (Leipzig) 170, 59] second law, considering the diffusion from the inner parts to the surface of the microbeads. The least‐squares and the Newton–Raphson [Carnahan, Luther and Wilkes (1969) Applied Numerical Methods, p. 319, John Wiley & Sons, New York] methods were used to obtain the diffusion coefficients. The microbead swelling at pH 7.4 and OTC diffusion is classically Fickian, suggesting that the OTC transport, in this case, is controlled by the exchange rates of free water and relaxation of calcium alginate chains. In case of acid media, it was observed that the phenomenon did not follow Fick's law, owing, probably, to the high solubility of the OTC in this environment. It was possible to modulate the release rate of OTC in several types of microbeads. The presence of cracks formed during the process of drying the microbeads was observed by scanning electron microscopy.
Kinetic and structural optimization to catalysis at low temperatures in a psychrophilic cellulase from the Antarctic bacterium Pseudoalteromonas haloplanktis
Authors: GARSOUX Geneviève , LAMOTTE Josette , GERDAY Charles , FELLER Georges
Published: November 2004

The cold-adapted cellulase CelG has been purified from the culture supernatant of the Antarctic bacterium Pseudoalteromonas haloplanktis and the gene coding for this enzyme has been cloned, sequenced and expressed in Escherichia coli. This cellulase is composed of three structurally and functionally distinct regions: an N-terminal catalytic domain belonging to glycosidase family 5 and a C-terminal cellulose-binding domain belonging to carbohydrate-binding module family 5. The linker of 107 residues connecting both domains is one of the longest found in cellulases, and optimizes substrate accessibility to the catalytic domain by drastically increasing the surface of cellulose available to a bound enzyme molecule. The psychrophilic enzyme is closely related to the cellulase Cel5 from Erwinia chrysanthemi. Both kcat and kcat/Km values at 4 °C for the psychrophilic cellulase are similar to the values for Cel5 at 30–35 °C, suggesting temperature adaptation of the kinetic parameters. The thermodynamic parameters of activation of CelG suggest a heat-labile, relatively disordered active site with low substrate affinity, in agreement with the experimental data. The structure of CelG has been constructed by homology modelling with a molecule of cellotetraose docked into the active site. No structural alteration related to cold-activity can be found in the catalytic cleft, whereas several structural factors in the overall structure can explain the weak thermal stability, suggesting that the loss of stability provides the required active-site mobility at low temperatures.
New insights into the structure–function relationships of Rho-associated kinase: a thermodynamic and hydrodynamic study of the dimer-to-monomer transition and its kinetic implications
Authors: DORAN John D. , LIU Xun , TASLIMI Paul , SAADAT Ahmad , FOX Ted
Published: November 2004

The effect of the length of ROCK (Rho-associated kinase) on its oligomerization state has been investigated by analysing full-length protein and four truncated constructs using light-scattering and analytical ultracentrifugation methods. Changes in size correlate with the kinetic properties of the kinase. Sedimentation velocity, sedimentation equilibrium and light-scattering data analyses revealed that protein constructs of size Ser6–Arg415 and larger exist predominantly as dimers, while smaller constructs are predominantly monomeric. The amino acid segments comprising residues 379–415 and 47–78 are shown to be necessary to maintain the dimeric ROCK structure. kcat values ranged from 0.7 to 2.1 s−1 and from 1.0 to 5.9 s−1 using ROCK peptide (KKRNRTLSV) and the 20000 Da subunit of myosin light chain respectively as substrate, indicating that the effect of the ROCK oligomerization state on the kcat is minor. Values of ATP Km for monomeric constructs were increased by 50–80-fold relative to the dimeric constructs, and Ki comparisons using the specific competitive ROCK inhibitor Y-27632 also showed increases of at least 120-fold, demonstrating significant perturbations in the ATP binding site. The corresponding Km values for the ROCK peptide and myosin light chain substrates increased in the range 1.4–16-fold, demonstrating that substrate binding is less sensitive to the ROCK oligomerization state. These results show that the oligomerization state of ROCK may influence both its kinase activity and its interactions with inhibitors, and suggest that the dimeric structure is essential for normal in vivo function.
Human replication-dependent histone H3 genes are activated by a tandemly arranged pair of two CCAAT boxes
Authors: KOESSLER Heiner , KAHLE Joerg , BODE Christa , DOENECKE Detlef , ALBIG Werner
Published: November 2004

We have analysed the transcriptional regulation of the human histone H3 genes using promoter deletion series, scanning mutagenesis, specific mutagenesis and electrophoretic mobility-shift assay experiments. The promoters of five of the six examined histone H3 genes showed near-maximal activity at lengths of 133–227 bp: H3/d 198 bp, H3/h 147 bp, H3/k 133 bp, H3/m 227 bp, H3/n 140 bp (exception H3/i). To search for functional cis-elements within these regions, we performed scanning mutagenesis of the two histone H3 promoters H3/k and H3/m. Mutagenesis revealed that the functional framework of the histone H3 promoters consists of a TATA box and two tandemly arranged CCAAT boxes in relatively fixed positions. Alterations of the distance between the CCAAT boxes and of the distance between the CCAAT boxes and the TATA box resulted in significant loss of activity. In electrophoretic mobility-shift assay experiments, the factor CBF (CCAAT-binding factor)/NF-Y (nuclear factor-Y) bound to isolated CCAAT boxes of the H3/k promoter. This suggests that an initiation complex is formed on the histone H3 promoter that has a defined structure and limited flexibility, consisting of two molecules of CBF/NF-Y and further (general or specific) transcription factors.
Structural diversity and specific distribution of O-glycans in normal human mucins along the intestinal tract
Authors: ROBBE Catherine , CAPON Calliope , CODDEVILLE Bernadette , MICHALSKI Jean-Claude
Published: November 2004

Purified human mucins from different parts of the intestinal tract (ileum, cecum, transverse and sigmoid colon and rectum) were isolated from two individuals with blood group ALeb (A-Lewisb). After alkaline borohydride treatment the released oligosaccharides were structurally characterized by nano-ESI Q-TOF MS/MS (electrospray ionization quadrupole time-of-flight tandem MS) without prior fractionation or derivatization. More than 100 different oligosaccharides, with up to ten monosaccharide residues, were identified using this technique. Oligosaccharides based on core 3 structures, GlcNAc(β1-3)GalNAc (where GlcNAc is N-acetyl-D-glucosamine and GalNAc is N-acetylgalactosamine), were widely distributed in human intestinal mucins. Core 5 structures, GalNAc(α1-3)GalNAc, were also recovered in all fractions. Moreover, a comparison of the oligosaccharide repertoire, with respect to size, diversity and expression of glycans and terminal epitopes, showed a high level of mucin-specific glycosylation: highly fucosylated glycans, found specifically in the small intestine, were mainly based on core 4 structures, GlcNAc-(β1-3)[GlcNAc(β1-6)]GalNAc, whereas the sulpho-LeX determinant carrying core 2 glycans, Gal(β1-3)[GlcNAc(β1-6)]-GalNAc (where Gal is galactose), was recovered mainly in the distal colon. Blood group H and A antigenic determinants were present exclusively in the ileum and cecum, whereas blood group Sda/Cad related epitopes, GalNAc(β1-4)[NeuAc(α2-3)]Gal (where NeuAc is N-acetylneuraminate), were found to increase along the length of the colon. Our findings suggest that mucins create an enormous repertoire of potential binding sites for micro-organisms that could explain the regio-specific colonization of bacteria in the human intestinal tract.
Interactions among p22, glyceraldehyde-3-phosphate dehydrogenase and microtubules
Authors: ANDRADE Josefa , PEARCE Sandy Timm , ZHAO Hu , BARROSO Margarida
Published: November 2004

Previously, we have shown that p22, an EF-hand Ca2+-binding protein, interacts indirectly with microtubules in an N-myristoylation-dependent and Ca2+-independent manner. In the present study, we report that N-myristoylated p22 interacts with several microtubule-associated proteins within the 30–100 kDa range using overlay blots of microtubule pellets containing cytosolic proteins. One of those p22-binding partners, a 35–40 kDa microtubule-binding protein, has been identified by MS as GAPDH (glyceraldehyde-3-phosphate dehydrogenase). Several lines of evidence suggest a functional relationship between GAPDH and p22. First, endogenous p22 interacts with GAPDH by immunoprecipitation. Secondly, p22 and GAPDH align along microtubule tracks in analogous punctate structures in BHK cells. Thirdly, GAPDH facilitates the p22-dependent interactions between microtubules and microsomal membranes, by increasing the ability of p22 to bind microtubules but not membranes. We have also shown a direct interaction between N-myristoylated p22 and GAPDH in vitro with a KD of ∼0.5 μM. The removal of either the N-myristoyl group or the last six C-terminal amino acids abolishes the binding of p22 to GAPDH and reduces the ability of p22 to associate with microtubules. In summary, we report that GAPDH is involved in the ability of p22 to facilitate microtubule–membrane interactions by affecting the p22–microtubule, but not the p22–membrane, association.
Localization of ferrochelatase in Plasmodium falciparum
Authors: VARADHARAJAN Sundaramurthy , SAGAR B. K. Chandrashekar , RANGARAJAN Pundi N. , PADMANABAN Govindarajan
Published: November 2004

Our previous studies have demonstrated de novo haem biosynthesis in the malarial parasite (Plasmodium falciparum and P. berghei). It has also been shown that the first enzyme of the pathway is the parasite genome-coded ALA (δ-aminolaevulinate) synthase localized in the parasite mitochondrion, whereas the second enzyme, ALAD (ALA dehydratase), is accounted for by two species: one species imported from the host red blood cell into the parasite cytosol and another parasite genome-coded species in the apicoplast. In the present study, specific antibodies have been raised to PfFC (parasite genome-coded ferrochelatase), the terminal enzyme of the haem-biosynthetic pathway, using recombinant truncated protein. With the use of these antibodies as well as those against the hFC (host red cell ferrochelatase) and other marker proteins, immunofluorescence studies were performed. The results reveal that P. falciparum in culture manifests a broad distribution of hFC and a localized distribution of PfFC in the parasite. However, PfFC is not localized to the parasite mitochondrion. Immunoelectron-microscopy studies reveal that PfFC is indeed localized to the apicoplast, whereas hFC is distributed in the parasite cytoplasm. These results on the localization of PfFC are unexpected and are at variance with theoretical predictions based on leader sequence analysis. Biochemical studies using the parasite cytosolic and organellar fractions reveal that the cytosol containing hFC accounts for 80% of FC enzymic activity, whereas the organellar fraction containing PfFC accounts for the remaining 20%. Interestingly, both the isolated cytosolic and organellar fractions are capable of independent haem synthesis in vitro from [4-14C]ALA, with the cytosol being three times more efficient compared with the organellar fraction. With [2-14C]glycine, most of the haem is synthesized in the organellar fraction. Thus haem is synthesized in two independent compartments: in the cytosol, using the imported host enzymes, and in the organellar fractions, using the parasite genome-coded enzymes.
Cryptococcal phospholipases: a novel lysophospholipase discovered in the pathogenic fungus Cryptococcus gattii
Authors: WRIGHT Lesley C. , PAYNE Jackie , SANTANGELO Rosemary T. , SIMPANYA Mukoma F. , CHEN Sharon C. A. , WIDMER Fred , SORRELL Tania C.
Published: November 2004

The pathogenic fungus Cryptococcus neoformans produces an extracellular PLB1 (phospholipase B1), shown previously to be a virulence factor. A novel phospholipase (LPL1) with only LPL (lysophospholipase) and LPTA (transacylase) activities has now been characterized in C. gattii, and found to be a 66-kDa glycoprotein (by SDS/PAGE), with a native molecular mass of 670 kDa. The pI was 6.3, and it was active at high temperatures (to 70 °C), as well as at both acidic and neutral pH values. It was stimulated by calcium and palmitoyl carnitine at pH 7.0, but not at pH 5.0, and palmitoyl lysophosphatidylcholine was the preferred substrate. Sequencing indicated that LPL1 is a novel cryptococcal lysophospholipase, and not the gene product of CnLYSO1 or PLB1. A protein with only LPL and LPTA activities was subsequently isolated from two strains of C. neoformans var. grubii. A PLB1 enzyme was isolated from both C. gattii and a highly virulent strain of C. neoformans var. grubii (H99). In both cases, all three enzyme activities (PLB, LPL and LPTA) were present in one 95–120 kDa glycoprotein (by SDS/PAGE) with pI 3.9–4.3. Characterization of PLB1 from C. gattii showed that it differed from that of C. neoformans in its larger native mass (275 kDa), high PLB activity relative to LPL and LPTA, and preference for saturated lipid substrates. Differences in the properties between the secreted phospholipases of the two cryptococcal species could contribute to phenotypic differences that determine their respective environmental niches and different clinical manifestations.
The protein structures that shape caspase activity, specificity, activation and inhibition
Authors: FUENTES-PRIOR Pablo , SALVESEN Guy S.
Published: November 2004

The death morphology commonly known as apoptosis results from a post-translational pathway driven largely by specific limited proteolysis. In the last decade the structural basis for apoptosis regulation has moved from nothing to ‘quite good’, and we now know the fundamental structures of examples from the initiator phase, the pre-mitochondrial regulator phase, the executioner phase, inhibitors and their antagonists, and even the structures of some substrates. The field is as well advanced as the best known of proteolytic pathways, the coagulation cascade. Fundamentally new mechanisms in protease regulation have been disclosed. Structural evidence suggests that caspases have an unusual catalytic mechanism, and that they are activated by apparently unrelated events, depending on which position in the apoptotic pathway they occupy. Some naturally occurring caspase inhibitors have adopted classic inhibition strategies, but other have revealed completely novel mechanisms. All of the structural and mechanistic information can, and is, being applied to drive therapeutic strategies to combat overactivation of apoptosis in degenerative disease, and underactivation in neoplasia. We present a comprehensive review of the caspases, their regulators and inhibitors from a structural and mechanistic point of view, and with an aim to consolidate the many threads that define the rapid growth of this field.
The human SNARE protein Ykt6 mediates its own palmitoylation at C-terminal cysteine residues
Authors: VEIT Michael
Published: November 2004

The yeast SNARE (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor) protein Ykt6 was shown to mediate palmitoylation of the fusion factor Vac8 in a reaction essential for the fusion of vacuoles. Here I present evidence that hYkt6 (human Ykt6) has self-palmitoylating activity. Incubation of recombinant hYkt6 with [3H]Pal-CoA ([3H]palmitoyl-CoA) leads to covalent attachment of palmitate to C-terminal cysteine residues. The N-terminal domain of human Ykt6 contains a Pal-CoA binding site and is required for the reaction.
Skeletal muscle morphology and capillarization of renal failure patients receiving different dialysis therapies
Authors: SAKKAS Giorgos K. , BALL Derek , SARGEANT Anthony J. , MERCER Thomas H. , KOUFAKI Pelagia , NAISH Patrick F.
Published: November 2004

The morphology of gastrocnemius muscles was examined in RFPs (renal failure patients) being treated using HD (haemodialysis) and CAPD (continuous ambulatory peritoneal dialysis). RFPs (n=24) volunteered to participate in the present study. Twelve RFPs (five women and seven men; mean age, 55 years) were undergoing CAPD treatment and 12 RFPs (two women and ten men; mean age, 62 years) were undergoing HD treatment. Muscle biopsies from gastrocnemius muscles were found not to differ (P>0.05) in fibre type distribution, MyHC (myosin heavy chain) expression or fibre CSA (cross-sectional area) between the two groups. There were, however, significant differences (P<0.05) in CC/F (capillary contact/fibre), C/F (capillary to fibre ratio) and cytochrome c oxidase activity. The HD group had 33% more CC/F, with a 19% higher C/F and 33% greater cytochrome c activity in glycolytic fibres (II) than the CAPD group. There were no apparent differences in age, gender, co-morbidity, self-reported physical activity or physical functioning between the two groups, which could account for the difference in muscle capillarity between the groups. The HD patients were, however, administered heparin as a routine part of the dialysis therapy. The possibility is discussed that heparin in combination with mild anaemia and acidosis may have augmented angiogenesis in the HD patients.
Elevated temporal QT variability index in patients with chronic renal failure
Authors: JOHANSSON Mats , GAO Sinsia A. , FRIBERG Peter , ANNERSTEDT Marita , BERGSTRÖM Göran , CARLSTRÖM Jan , IVARSSON Ted , JENSEN Gert , LJUNGMAN Susanne , MATHILLAS Öivind , NIELSEN Finn-David , STRÖMBOM Ulf
Published: November 2004

Patients with CRF (chronic renal failure) are at increased risk of cardiovascular diseases, and 60% of cardiovascular mortality in CRF is attributed to sudden death. Various abnormalities in myocardial repolarization are associated with the risk of ventricular arrhythmia. The aim of this study was to evaluate an index of temporal myocardial repolarization lability, the temporal QTVI (QT variability index), in patients with CRF. ECGs were recorded in 153 patients with CRF on haemodialysis (n=67), continuous ambulatory peritoneal dialysis (n=43) or conservative treatment (n=43) during 30 min of rest. QTVI was calculated as the logarithm of the ratio between the variances of the normalized QT and RR intervals. Age-matched healthy subjects (n=39) were examined for comparison. QTVI was increased by 47% in CRF patients compared with healthy subjects (−0.82±0.56 compared with −1.54±0.27 respectively; P<0.01). QTVI did not differ among patients on dialysis or conservative treatment, whereas QTVI was elevated further in patients with diabetes compared with non-diabetic CRF patients (−0.56±0.54 compared with −0.94±0.52 respectively; P<0.01). In a multiple linear regression analysis, diabetes and a history of coronary artery disease were the only independent predictors of QTVI in the CRF population. The present study demonstrates that elevated QTVI in patients with CRF is associated with diabetes and coronary disease. The present findings are important given that repolarization instability may predispose to ventricular arrhythmia and sudden death, events that occur frequently in CRF patients.
Running for health: how much running for how much health?
Authors: McCULLY Kevin
Published: November 2004

Increasing physical activity has been shown to reduce physiological markers of cardiovascular disease, such as high blood pressure, vascular endothelial cell reactivity and arterial stiffness. In this issue of Clinical Science, Hägg and colleagues have chosen the spontaneously hypertensive rat model to investigate the effect of exercise on vascular function. They found that spontaneous running increased aortic compliance and antioxidant capacity with decreased oxidative stress in mesenteric arteries, presenting support for the cardiovascular protective effects of physical activity. Two important aspects of their study include the magnitude of the running stimulus and the choice of artery to evaluate.
Strain hardening and large tensile elongation in ultrahigh-strength nano-twinned copper
Authors: Ma E. , Wang Y. M. , Lu Q. H. , Sui M. L. , Lu L. , Lu K.
Published: November 2004

A high density of growth twins in pure Cu imparts high yield strength while preserving the capacity for efficient dislocation storage, leading to high strain hardening rates at high flow stresses, especially at 77K. Uniform tensile deformation is stabilized to large plastic strains, resulting in an ultrahigh tensile strength of ∼1GPa together with an elongation to failure of ∼30%.
Scanning probe energy loss spectroscopy below 50nm resolution
Authors: Festy F. , Palmer R. E.
Published: November 2004

We report scanning probe energy loss spectroscopy (SPELS) measurements from a roughened Si(111) surface in ultrahigh vacuum. The experiments, which utilize a scanning tunneling microscope tip in the field emission mode as the electron source, establish that the spatial resolution in SPELS is better than 50nm. The spectral maps acquired indicate different contrast mechanisms for the inelastically scattered and secondary electrons identified in the energy loss spectrum.