bluuebunny/crossref_metadata_2025 huggingface dataset:
Sagittal tibial translation during exercises in the anterior cruciate ligament‐deficient knee
Authors: Kvist Joanna
Published: May 2005

The aim of this study was to describe and compare the sagittal tibial translation and electromyographic activity of muscles v. medialis and lateralis, gastrocnemius and hamstrings, during common rehabilitation exercises, in patients with anterior cruciate ligament (ACL) deficiency and non‐injured controls. Sagittal tibial translation was registered with the CA‐4000 electrogoniometer, in 12 patients and 17 controls, during Lachman test (static translation) and five exercises (dynamic translation). The exercises were grouped according to muscle work and joint compression (active extension, heel raises, cycling, one‐legged squat and chair squat).

The non‐weight‐bearing exercise with isolated muscle work (active extension) produced a large amount of tibial translation. During weight bearing, the total anterior–posterior tibial motion was halved compared to the non‐weight‐bearing exercises, but tibia was anterior positioned. Heel raising resulted in equal translation as the one‐legged squat and chair squat, whereas cycling produced the smallest amount of tibial translation. The subjects utilized different amounts of their individual joint play (static translation) during the exercises, which may explain why there are no correlation between static translation and the patients functional outcome. These results enhance understanding on tibial translation during activity and gives indications on which exercises can be used early after ACL injury and reconstruction.
Quantitative assessment of the glyoxalase pathway in Leishmania infantum as a therapeutic target by modelling and computer simulation
Authors: Sousa Silva Marta , Ferreira António E. N. , Tomás Ana Maria , Cordeiro Carlos , Ponces Freire Ana
Published: May 2005

The glyoxalase pathway of Leishmania infantum was kinetically characterized as a trypanothione‐dependent system. Using time course analysis based on parameter fitting with a genetic algorithm, kinetic parameters were estimated for both enzymes, with trypanothione derived substrates. A Km of 0.253 mm and a V of 0.21 µmol·min−1·mg−1for glyoxalase I, and a Km of 0.098 mm and a V of 0.18 µmol·min−1·mg−1 for glyoxalase II, were obtained. Modelling and computer simulation were used for evaluating the relevance of the glyoxalase pathway as a potential therapeutic target by revealing the importance of critical parameters of this pathway in Leishmania infantum. A sensitivity analysis of the pathway was performed using experimentally validated kinetic models and experimentally determined metabolite concentrations and kinetic parameters. The measurement of metabolites in L. infantum involved the identification and quantification of methylglyoxal and intracellular thiols. Methylglyoxal formation in L. infantum is nonenzymatic. The sensitivity analysis revealed that the most critical parameters for controlling the intracellular concentration of methylglyoxal are its formation rate and the concentration of trypanothione. Glyoxalase I and II activities play only a minor role in maintaining a low intracellular methylglyoxal concentration. The importance of the glyoxalase pathway as a therapeutic target is very small, compared to the much greater effects caused by decreasing trypanothione concentration or increasing methylglyoxal concentration.
The lipid/protein interface as xenobiotic target site
Authors: Altschuh Joachim , Walcher Sebastian , Sandermann Heinrich
Published: May 2005

High‐resolution X‐ray diffraction structures of integral membrane proteins have revealed various binding modes of lipids, but current spectroscopic studies still use uniform macroscopic binding constants to describe lipid binding. The Adair approach employing microscopic lipid‐binding constants has previously been taken to explain the enhancement of agonist binding to the nicotinic acetylcholine receptor by general anaesthetics in terms of the competitive displacement of essential lipid activator molecules [Walcher S, Altschuh J & Sandermann H (2001) J. Biol. Chem.276, 42191–42195]. This approach was extended to tadpole narcosis induced by alcohols. A single class, or two different classes of lipid activator binding sites, are considered. Microscopic lipid and inhibitor binding constants are derived and allow a close fit to dose–response curves of tadpole narcosis on the basis of a preferential displacement of more loosely bound essential lipid activator molecules. This study illustrates the potential of the Adair approach to resolve protein‐bound lipid populations.
Production and utilization of hydrogen peroxide associated with melanogenesis and tyrosinase‐mediated oxidations of DOPA and dopamine
Authors: Mastore Maristella , Kohler Lara , Nappi Anthony J.
Published: May 2005

The synthesis and involvement of H2O2 during the early stages of melanogenesis involving the oxidations of DOPA and dopamine (diphenolase activity) were established by two sensitive and specific electrochemical detection systems. Catalase‐treated reaction mixtures showed diminished rates of H2O2 production during the autoxidation and tyrosinase‐mediated oxidation of both diphenols. Inhibition studies with the radical scavenger resveratrol revealed the involvement in these reactions of additional reactive intermediate of oxygen (ROI), one of which appears to be superoxide anion. There was no evidence to suggest that H2O2 or any other ROI was produced during the tyrosinase‐mediated conversion of tyrosine to DOPA (monophenolase activity). Establishing by electrochemical methods the endogenous production H2O2 in real time confirms recent reports, based in large part on the use of exogenous H2O2, that tyrosinase can manifest both catalase and peroxidase activities. The detection of ROI in tyrosinase‐mediated in vitro reactions provides evidence for sequential univalent reductions of O2, most likely occurring at the enzyme active site copper. Collectively, these observations focus attention on the possible involvement of peroxidase‐H2O2 systems and related ROI‐mediated reactions in promoting melanocytotoxic and melanoprotective processes.
Oligosaccharide synthesis inFibrobacter succinogenesS85 and its modulation by the substrate
Authors: Nouaille Régis , Matulova Maria , Delort Anne‐Marie , Forano Evelyne
Published: May 2005

In this article we compared the metabolism of phosphorylated and unphosphorylated oligosaccharides (cellodextrins and maltodextrins) inFibrobacter succinogenesS85 resting cells incubated with the following substrates: glucose; cellobiose; a mixture of glucose and cellobiose; and cellulose. Intracellular and extracellular media were analysed by1H‐NMR and by TLC. The first important finding is that no cellodextrins were found to accumulate in the extracellular media of cells, regardless of the substrate; this contrasts to what is generally reported in the literature. The second finding of this work is that maltodextrins of degree of polymerization > 2 are synthesized regardless of the substrate, and can be used by the bacteria. Maltotriose plays a key role in this metabolism of maltodextrin. Maltodextrin‐1‐phosphate was detected in all the incubations, and a new metabolite, corresponding to a phosphorylated glucose derivative, was produced in the extracellular medium when cells were incubated with cellulose. The accumulation of these phosphorylated sugars increased with the degree of polymerization of the substrate.
Characterization of a prokaryotic haemerythrin from the methanotrophic bacterium Methylococcus capsulatus (Bath)
Authors: Karlsen Odd A. , Ramsevik Linda , Bruseth Live J. , Larsen Øivind , Brenner Annette , Berven Frode S. , Jensen Harald B. , Lillehaug Johan R.
Published: May 2005

For a long time, the haemerythrin family of proteins was considered to be restricted to only a few phyla of marine invertebrates. When analysing differential protein expression in the methane‐oxidizing bacterium, Methylococcus capsulatus (Bath), grown at a high and low copper‐to‐biomass ratio, respectively, we identified a putative prokaryotic haemerythrin expressed in high‐copper cultures. Haemerythrins are recognized by a conserved sequence motif that provides five histidines and two carboxylate ligands which coordinate two iron atoms. The diiron site is located in a hydrophobic pocket and is capable of binding O2. We cloned the M. capsulatus haemerythrin gene and expressed it in Escherichia coli as a fusion protein with NusA. The haemerythrin protein was purified to homogeneity cleaved from its fusion partner. Recombinant M. capsulatus haemerythrin (McHr) was found to fold into a stable protein. Sequence similarity analysis identified all the candidate residues involved in the binding of diiron (His22, His58, Glu62, His77, His81, His117, Asp122) and the amino acids forming the hydrophobic pocket in which O2 may bind (Ile25, Phe59, Trp113, Leu114, Ile118). We were also able to model a three‐dimensional structure of McHr maintaining the correct positioning of these residues. Furthermore, UV/vis spectrophotometric analysis demonstrated the presence of conjugated diiron atoms in McHr. A comprehensive genomic database search revealed 21 different prokaryotes containing the haemerythrin signature (PROSITE 00550), indicating that these putative haemerythrins may be a conserved prokaryotic subfamily.
The effect of replacing the axial methionine ligand with a lysine residue in cytochrome c‐550 from Paracoccus versutus assessed by X‐ray crystallography and unfolding
Authors: Worrall Jonathan A. R. , van Roon Anne‐Marie M. , Ubbink Marcellus , Canters Gerard W.
Published: May 2005

The structure of cytochrome c‐550 from the nonphotosynthetic bacteria Paraccocus versutus has been solved by X‐ray crystallography to 1.90 Å resolution, and reveals a high structural homology to other bacterial cytochromes c2. The effect of replacing the axial heme‐iron methionine ligand with a lysine residue on protein structure and unfolding has been assessed using the M100K variant. From X‐ray structures at 1.95 and 1.55 Å resolution it became clear that the amino group of the lysine side chain coordinates to the heme‐iron. Structural differences compared to the wild‐type protein are confined to the lysine ligand loop connecting helices four and five. In the heme cavity an additional water molecule is found which participates in an H‐bonding interaction with the lysine ligand. Under cryo‐conditions extra electron density in the lysine ligand loop is revealed, leading to residues K97 to T101 being modeled with a double main‐chain conformation. Upon unfolding, dissociation of the lysine ligand from the heme‐iron is shown to be pH dependent, with NMR data consistent with the occurrence of a ligand exchange mechanism similar to that seen for the wild‐type protein.
Interaction of synthetic peptides corresponding to hepatitis G virus (HGV/GBV‐C) E2 structural protein with phospholipid vesicles
Authors: Larios Cristina , Christiaens Bart , Gómara M. José , Alsina M. Asunción , Haro Isabel
Published: May 2005

The interaction with phospholipid bilayers of two synthetic peptides with sequences corresponding to a segment next to the native N‐terminus and an internal region of the E2 structural hepatitis G virus (HGV/GBV‐C) protein [E2(7–26) and E2(279–298), respectively] has been characterized. Both peptides are water soluble but associate spontaneously with bilayers, showing higher affinity for anionic than zwitterionic membranes. However, whereas the E2(7–26) peptide is hardly transferred at all from water to the membrane interface, the E2(279–298) peptide is able to penetrate into negatively charged bilayers remaining close to the lipid/water interface. The nonpolar environment clearly induces a structural transition in the E2(279–298) peptide from random coil to α‐helix, which causes bilayer perturbations leading to vesicle permeabilization. The results indicate that this internal segment peptide sequence is involved in the fusion of HGV/GBV‐C to membrane.
Solution structure and backbone dynamics of the XPC‐binding domain of the human DNA repair protein hHR23B
Authors: Kim Byoungkook , Ryu Kyoung‐Seok , Kim Hyun‐Jin , Cho Sung‐Jae , Choi Byong‐Seok
Published: May 2005

Human cells contain two homologs of the yeast RAD23 protein, hHR23A and hHR23B, which participate in the DNA repair process. hHR23B houses a domain (residues 277–332, called XPCB) that binds specifically and directly to the xeroderma pigmentosum group C protein (XPC) to initiate nucleotide excision repair (NER). This domain shares sequence homology with a heat shock chaperonin‐binding motif that is also found in the stress‐inducible yeast phosphoprotein STI1. We determined the solution structure of a protein fragment containing amino acids 275–342 of hHR23B (termed XPCB–hHR23B) and compared it with the previously reported solution structures of the corresponding domain of hHR23A. The periodic positioning of proline residues in XPCB–hHR23B produced kinked α helices and assisted in the formation of a compact domain. Although the overall structure of the XPCB domain was similar in both XPCB–hHR23B and XPCB–hHR23A, the N‐terminal part (residues 275–283) of XPCB–hHR23B was more flexible than the corresponding part of hHR23A. We tried to infer the characteristics of this flexibility through 15N‐relaxation studies. The hydrophobic surface of XPCB–hHR23B, which results from the diverse distribution of N‐terminal region, might give rise to the functional pleiotropy observed in vivo for hHR23B, but not for hHR23A.
Characterization of structural and catalytic differences in rat intestinal alkaline phosphatase isozymes
Authors: Harada Tsuyoshi , Koyama Iwao , Matsunaga Toshiyuki , Kikuno Akira , Kasahara Toshihiko , Hassimoto Masatoshi , Alpers David H. , Komoda Tsugikazu
Published: May 2005

To understand the differences between the rat intestinal alkaline phosphatase isozymes rIAP‐I and rIAP‐II, we constructed structural models based on the previously determined crystal structure for human placental alkaline phosphatase (hPLAP). Our models of rIAP‐I and rIAP‐II displayed a typical α/β topology, but the crown domain of rIAP‐I contained an additional β‐sheet, while the embracing arm region of rIAP‐II lacked the α‐helix, when each model was compared to hPLAP. The representations of surface potential in the rIAPs were predominantly positive at the base of the active site. The coordinated metal at the active site was predicted to be a zinc triad in rIAP‐I, whereas the typical combination of two zinc atoms and one magnesium atom was proposed for rIAP‐II. Using metal‐depleted extracts from rat duodenum or jejunum and hPLAP, we performed enzyme assays under restricted metal conditions. With the duodenal and jejunal extract, but not with hPLAP, enzyme activity was restored by the addition of zinc, whereas in nonchelated extracts, the addition of zinc inhibited duodenal IAP and hPLAP, but not jejunal IAP. Western blotting revealed that nearly all of the rIAP in the jejunum extracts was rIAP‐I, whereas in duodenum the percentage of rIAP‐I (55%) correlated with the degree of AP activation (60% relative to that seen with jejunal extracts). These data are consistent with the presence of a triad of zinc atoms at the active site of rIAP‐I, but not rIAP‐II or hPLAP. Although no differences in amino acid alignment in the vicinity of metal‐binding site 3 were predicted between the rIAPs and hPLAP, the His153 residue of both rIAPs was closer to the metal position than that in hPLAP. Between the rIAPs, a difference was observed at amino acid position 317 that is indirectly related to the coordination of the metal at metal‐binding site 3 and water molecules. These findings suggest that the side‐chain position of His153, and the alignment of Q317, might be the major determinants for activation of the zinc triad in rIAP‐I.
Adaptation of intronic homing endonuclease for successful horizontal transmission
Authors: Kurokawa Sayuri , Bessho Yoshitaka , Higashijima Kyoko , Shirouzu Mikako , Yokoyama Shigeyuki , Watanabe Kazuo I. , Ohama Takeshi
Published: May 2005

Group I introns are thought to be self‐propagating mobile elements, and are distributed over a wide range of organisms through horizontal transmission. Intron invasion is initiated through cleavage of a target DNA by a homing endonuclease encoded in an open reading frame (ORF) found within the intron. The intron is likely of no benefit to the host cell and is not maintained over time, leading to the accumulation of mutations after intron invasion. Therefore, regular invasional transmission of the intron to a new species at least once before its degeneration is likely essential for its evolutionary long‐term existence. In many cases, the target is in a protein‐coding region which is well conserved among organisms, but contains ambiguity at the third nucleotide position of the codon. Consequently, the homing endonuclease might be adapted to overcome sequence polymorphisms at the target site. To address whether codon degeneracy affects horizontal transmission, we investigated the recognition properties of a homing enzyme, I‐CsmI, that is encoded in the intronic ORF of a group I intron located in the mitochondrial COB gene of the unicellular green alga Chlamydomonas smithii. We successfully expressed and purified three types of N‐terminally truncated I‐CsmI polypeptides, and assayed the efficiency of cleavage for 81 substrates containing single nucleotide substitutions. We found a slight but significant tendency that I‐CsmI cleaves substrates containing a silent or tolerated amino acid change more efficiently than nonsilent or nontolerated ones. The published recognition properties of I‐SpomI, I‐ScaI, and I‐SceII were reconsidered from this point of view, and we detected proficient adaptation of I‐SpomI, I‐ScaI, and I‐SceII for target site sequence degeneracy. Based on the results described above, we propose that intronic homing enzymes are adapted to cleave sequences that might appear at the target region in various species, however, such adaptation becomes less prominent in proportion to the time elapsed after intron invasion into a new host.
ERS1 encodes a functional homologue of the human lysosomal cystine transporter
Authors: Gao Xiao‐Dong , Wang Ji , Keppler‐Ross Sabine , Dean Neta
Published: May 2005

Cystinosis is a lysosomal storage disease caused by an accumulation of insoluble cystine in the lumen of the lysosome. CTNS encodes the lysosomal cystine transporter, mutations in which manifest as a range of disorders and are the most common cause of inherited renal Fanconi syndrome. Cystinosin, the CTNS product, is highly conserved among mammals. Here we show that the yeast Ers1 protein and cystinosin are functional orthologues, despite sharing only limited sequence homology. Ers1 is a vacuolar protein whose loss of function results in growth sensitivity to hygromycin B. This phenotype can be complemented by the human CTNS gene but not by mutant ctns alleles that were previously identified in cystinosis patients. A genetic screen for multicopy suppressors of an ers1Δ yeast strain identified a novel gene, MEH1, which is implicated in regulating Ers1 function. Meh1 localizes to the vacuolar membrane and loss of MEH1 results in a defect in vacuolar acidification, suggesting that the vacuolar environment is critical for normal ERS1 function. This genetic system has also led us to identify Gtr1 as an Meh1 interacting protein. Like Meh1 and Ers1, Gtr1 associates with vacuolar membranes in an Meh1‐dependent manner. These results demonstrate the utility of yeast as a model system for the study of CTNS and vacuolar function.
Phosphatidylinositol 3,4,5‐trisphosphate modulation in SHIP2‐deficient mouse embryonic fibroblasts
Authors: Blero Daniel , Zhang Jing , Pesesse Xavier , Payrastre Bernard , Dumont Jacques E. , Schurmans Stéphane , Erneux Christophe
Published: May 2005

SHIP2, the ubiquitous SH2 domain containing inositol 5‐phosphatase, includes a series of protein interacting domains and has the ability to dephosphorylate phosphatidylinositol 3,4,5‐trisphosphate [PtdIns(3,4,5)P3]in vitro. The present study, which was undertaken to evaluate the impact of SHIP2 on PtdIns(3,4,5)P3 levels, was performed in a mouse embryonic fibroblast (MEF) model using SHIP2 deficient (–/–) MEF cells derived from knockout mice. PtdIns(3,4,5)P3 was upregulated in serum stimulated –/– MEF cells as compared to +/+ MEF cells. Although the absence of SHIP2 had no effect on basal PtdIns(3,4,5)P3 levels, we show here that this lipid was significantly upregulated in SHIP2 –/– cells but only after short‐term (i.e. 5–10 min) incubation with serum. The difference in PtdIns(3,4,5)P3 levels in heterozygous fibroblast cells was intermediate between the +/+ and the –/– cells. In our model, insulin‐like growth factor‐1 stimulation did not show this upregulation. Serum stimulated phosphoinositide 3‐kinase (PI 3‐kinase) activity appeared to be comparable between +/+ and –/– cells. Moreover, protein kinase B, but not mitogen activated protein kinase activity, was also potentiated in SHIP2 deficient cells stimulated by serum. The upregulation of protein kinase B activity in serum stimulated cells was totally reversed in the presence of the PI 3‐kinase inhibitor LY‐294002, in both +/+ and –/– cells. Altogether, these data establish a link between SHIP2 and the acute control of PtdIns(3,4,5)P3 levels in intact cells.
Dual role of oxygen during lipoxygenase reactions
Authors: Ivanov Igor , Saam Jan , Kuhn Hartmut , Holzhütter Hermann‐Georg
Published: May 2005

Studying the oxygenation kinetics of (19R/S,5Z,8Z,11Z,14Z)‐19‐hydroxyeicosa‐5,8,11,14‐tetraenoic acid (19‐OH‐AA) by rabbit 15‐lipoxygenase‐1 we observed a pronounced oxygen dependence of the reaction rate, which was not apparent with arachidonic acid as substrate. Moreover, we found that peroxide‐dependent activation of the lipoxygenase depended strongly on the oxygen concentration. These data can be described with a kinetic model that extends previous schemes of the lipoxygenase reaction in three essential aspects: (a) the product of 19‐OH‐AA oxygenation is a less effective lipoxygenase activator than (13S,9Z,11E)‐13‐hydroperoxyoctadeca‐9,11‐dienoic acid; (b) molecular dioxygen serves not only as a lipoxygenase substrate, but also impacts peroxide‐dependent enzyme activation; (c) there is a leakage of radical intermediates from the catalytic cycle, which leads to the formation of inactive ferrous lipoxygenase. This enzyme inactivation can be reversed by another round of peroxide‐dependent activation. Taken together our data indicate that both peroxide activation and the oxygen affinity of lipoxygenases depend strongly on the chemistry of the lipid substrate. These findings are of biological relevance as variations of the reaction conditions may turn the lipoxygenase reaction into an efficient source of free radicals.
Production and characterization of a noncytotoxic deletion variant of the Aspergillus fumigatus allergen Aspf1 displaying reduced IgE binding
Authors: Garciá‐Ortega Lucía , Lacadena Javier , Villalba Mayte , Rodríguez Rosalía , Crespo Jesús F. , Rodríguez Julia , Pascual Cristina , Olmo Nieves , Oñaderra Mercedes , del Pozo Álvaro Martínez , Gavilanes José G.
Published: May 2005

Aspergillus fumigatus is responsible for many allergic respiratory diseases, the most notable of which − due to its severity − is allergic bronchopulmonary aspergillosis. Aspf1 is a major allergen of this fungus: this 149‐amino acid protein belongs to the ribotoxin family, whose best characterized member is α‐sarcin (EC 3.1.27.10). The proteins of this group are cytotoxic ribonucleases that degrade a unique bond in ribosomal RNA impairing protein biosynthesis. Aspf1 and its deletion mutant Aspf1Δ(7–22) have been produced as recombinant proteins; the deleted region corresponds to an exposed β‐hairpin. The conformation of these two proteins has been studied by CD and fluorescence spectroscopy. Their enzymatic activity and cytotoxicity against human rhabdomyosarcoma cells was also measured and their allergenic properties have been studied by using 58 individual sera of patients sensitized to Aspergillus. Aspf1Δ(7–22) lacks cytotoxicity and shows a remarkably reduced IgE reactivity. From these studies it can be concluded that the deleted β‐hairpin is involved in ribosome recognition and is a significant allergenic region.
Nature, nurture and neurology: gene–environment interactions in neurodegenerative disease
Authors: Spires Tara L. , Hannan Anthony J.
Published: May 2005

Neurodegenerative disorders, such as Huntington's, Alzheimer's, and Parkinson's diseases, affect millions of people worldwide and currently there are few effective treatments and no cures for these diseases. Transgenic mice expressing human transgenes for huntingtin, amyloid precursor protein, and other genes associated with familial forms of neurodegenerative disease in humans provide remarkable tools for studying neurodegeneration because they mimic many of the pathological and behavioural features of the human conditions. One of the recurring themes revealed by these various transgenic models is that different diseases may share similar molecular and cellular mechanisms of pathogenesis. Cellular mechanisms known to be disrupted at early stages in multiple neurodegenerative disorders include gene expression, protein interactions (manifesting as pathological protein aggregation and disrupted signaling), synaptic function and plasticity. Recent work in mouse models of Huntington's disease has shown that enriching the environment of transgenic animals delays the onset and slows the progression of Huntington's disease‐associated motor and cognitive symptoms. Environmental enrichment is known to induce various molecular and cellular changes in specific brain regions of wild‐type animals, including altered gene expression profiles, enhanced neurogenesis and synaptic plasticity. The promising effects of environmental stimulation, demonstrated recently in models of neurodegenerative disease, suggest that therapy based on the principles of environmental enrichment might benefit disease sufferers and provide insight into possible mechanisms of neurodegeneration and subsequent identification of novel therapeutic targets. Here, we review the studies of environmental enrichment relevant to some major neurodegenerative diseases and discuss their research and clinical implications.
Affinity of S100A1 protein for calcium increases dramatically upon glutathionylation
Authors: Goch Grażyna , Vdovenko Sergiusz , Kozłowska Hanna , Bierzyñski Andrzej
Published: May 2005

S100A1 is a typical representative of a group of EF‐hand calcium‐binding proteins known as the S100 family. The protein is composed of two α subunits, each containing two calcium‐binding loops (N and C). At physiological pH (7.2) and NaCl concentration (100 mm), we determined the microscopic binding constants of calcium to S100A1 by analysing the Ca2+‐titration curves of Trp90 fluorescence for both the native protein and its Glu32 → Gln mutant with an inactive N‐loop. Using a chelator method, we also determined the calcium‐binding constant for the S100A1 Glu73 → Gln mutant with an inactive C‐loop. The protein binds four calcium ions in a noncooperative way with binding constants of K1 =4 ± 2 × 103 m−1 (C‐loops) and K2≈ 102 m−1 (N‐loops). Only when both loops are saturated with calcium does the protein change its global conformation, exposing to the solvent hydrophobic patches, which can be detected by 2‐p‐toluidinylnaphthalene‐6‐sulfonic acid – a fluorescent probe of protein‐surface hydrophobicity. S‐Glutathionylation of the single cysteine residue (85) of the α subunits leads to a 10‐fold increase in the affinity of the protein C‐loops for calcium and an enormous – four orders of magnitude – increase in the calcium‐binding constants of its N‐loops, owing to a cooperativity effect corresponding to ΔΔG = −6 ± 1 kcal·mol−1. A similar effect is observed upon formation of the mixed disulfide with cysteine and 2‐mercaptoethanol. The glutathionylated protein binds TRTK‐12 peptide in a calcium‐dependent manner. S100A1 protein can act, therefore, as a linker between the calcium and redox signalling pathways.
Quantitative interdependence of coeffectors, CcpA and cre in carbon catabolite regulation of Bacillus subtilis
Authors: Seidel Gerald , Diel Marco , Fuchsbauer Norbert , Hillen Wolfgang
Published: May 2005

The phosphoproteins HPrSerP and CrhP are the main effectors for CcpA‐mediated carbon catabolite regulation (CCR) in Bacillus subtilis. Complexes of CcpA with HPrSerP or CrhP regulate genes by binding to the catabolite responsive elements (cre). We present a quantitative analysis of HPrSerP and CrhP interaction with CcpA by surface plasmon resonance (SPR) revealing small and similar equilibrium constants of 4.8 ± 0.4 µm for HPrSerP–CcpA and 19.1 ± 2.5 µm for CrhP–CcpA complex dissociation. Forty millimolar fructose‐1,6‐bisphosphate (FBP) or glucose‐6‐phosphate (Glc6‐P) increases the affinity of HPrSerP to CcpA at least twofold, but have no effect on CrhP–CcpA binding. Saturation of binding of CcpA to cre as studied by fluorescence and SPR is dependent on 50 µm of HPrSerP or > 200 µm CrhP. The rate constants of HPrSerP–CcpA–cre complex formation are ka = 3 ± 1 × 106 m−1·s−1 and kd = 2.0 ± 0.4 × 10−3·s−1, resulting in a KD of 0.6 ± 0.3 nm. FBP and Glc6‐P stimulate CcpA–HPrSerP but not CcpA‐CrhP binding to cre. Maximal HPrSerP‐CcpA–cre complex formation in the presence of 10 mm FBP requires about 10‐fold less HPrSerP. These data suggest a specific role for FBP and Glc6‐P in enhancing only HPrSerP‐mediated CCR.
Calpain 1–titin interactions concentrate calpain 1 in the Z‐band edges and in the N2‐line region within the skeletal myofibril
Authors: Raynaud Fabrice , Fernandez Eric , Coulis Gerald , Aubry Laurent , Vignon Xavier , Bleimling Nathalie , Gautel Mathias , Benyamin Yves , Ouali Ahmed
Published: May 2005

Calpain 1, a ubiquitous calcium‐dependent intracellular protease, was recently found in a tight association with myofibrils in skeletal muscle tissue [Delgado EF, Geesink GH, Marchello JA, Goll DE & Koohmaraie M (2001) J Anim Sci79, 2097–2107). Our immunofluorescence and immunoelectron microscopy investigations restrain the protease location at the periphery of the Z‐band and at the midpoint of the I‐band. Furthermore, calpain 1 is found to localize in myofibril fractures, described as proteolysis sites, in postmortem bovine skeletal red muscles, near the calcium deposits located at the N1 and N2 level. This in situ localization of calpain 1 is substantiated by binding assays with two titin regions covering the I‐band region: a native fragment of 150 kDa (identified by mass spectrometry) that includes the N‐terminal Z8–I5 region and the N1‐line region of titin, and an 800 kDa fragment external to the N1 line that bears the PEVK/N2 region. These two titin fragments are shown to tightly bind calpain 1 in the presence of CaCl2 and E64, a calpain inhibitor. In the absence of E64, they are cleaved by calpain 1. We conclude that titin affords binding sites to calpain 1, which concentrates the protease in the regions restrained by the Z‐band edge and the N1‐line as well as at the N2‐line level, two sarcomeric regions where early postmortem proteolysis is detected.
The binding of foot‐and‐mouth disease virus leader proteinase to eIF4GI involves conserved ionic interactions
Authors: Foeger Nicole , Kuehnel Elisabeth , Cencic Regina , Skern Tim
Published: May 2005

The leader proteinase (Lpro) of foot‐and‐mouth disease virus (FMDV) initially cleaves itself from the polyprotein. Subsequently, Lpro cleaves the host proteins eukaryotic initiation factor (eIF) 4GI and 4GII. This prevents protein synthesis from capped cellular mRNAs; the viral RNA is still translated, initiating from an internal ribosome entry site. Lpro cleaves eIF4GI between residues G674 and R675. We showed previously, however, that Lpro binds to residues 640–669 of eIF4GI. Binding was substantially improved when the eIF4GI fragment contained the eIF4E binding site and eIF4E was present in the binding assay. Lpro interacts with eIF4GI via residue C133 and residues 183–195 of the C‐terminal extension. This binding domain lies about 25 Å from the active site. Here, we examined the binding of Lpro to eIF4GI fragments generated by in vitro translation to narrow the binding site down to residues 645–657 of human eIF4GI. Comparison of these amino acids with those in human eIF4GII as well as with sequences of eIF4GI from other organisms allowed us to identify two conserved basic residues (K646 and R650). Mutation of these residues was severely detrimental to Lpro binding. Similarly, comparison of the sequence between residues 183 and 195 of Lpro with those of other FMDV serotypes and equine rhinitis A virus showed that acidic residues D184 and E186 were highly conserved. Substitution of these residues in Lpro significantly reduced eIF4GI binding and cleavage without affecting self‐processing. Thus, FMDV Lpro has evolved a domain that specifically recognizes a host cell protein.
Grade III bleeding haemorrhoids in a 6‐year‐old boy
Authors: Gupta Pravin J.
Published: May 2005

We report a case of grade 3 bleeding haemorrhoids in a 6‐year‐old boy, who had a history of bleeding per rectum since he was 1‐year‐old. The diagnosis was made on anoscopic examination after ruling out other colorectal pathologies by a full‐length colonoscopy. There was no evidence of portal hypertension.

He was successfully treated with haemorrhoidectomy.

A congenital weakness of the haemorrhoid vein wall was thought to be the probable cause of disease for want of any other causative factor. Though rare to be found at the age of 6 years without any obvious aetiology, haemorrhoids should be considered as one of the causes of bleeding per rectum in children.
Catheter‐related sepsis in ultrasound‐guided percutaneously inserted long‐term tunnelled central venous catheter: A review of 50 patients
Authors: Li Wing‐Hong , Tsang Shiu‐Hei , Tsao Joanne Pui‐Yue , Tong Wai‐Chung , Tang Lap‐Fai
Published: May 2005

Objective:  Ultrasound‐guided percutaneous insertion of a long‐term tunneled central venous catheter (TCVC) is now a very common procedure, but catheter‐related sepsis remains a significant problem. This study aims to review our results concerning this complication.

Methodology:  Fifty‐seven central venous catheters were inserted into 50 patients percutaneously under ultrasound guidance in Queen Elizabeth hospital for between 1 March 2003 and 30 April 2004. Data was retrospectively collected and analysed from hospital records.

Results:  This study had a successful placement rate of 100%. The overall complication rate was 31% (18 out of 57), comprised of four mechanical complications (all occurring within 30 days of insertion). There was no complication documented within the first 24 h. Fourteen patients developed infective complications, and of these eight patients were classified as catheter‐related sepsis, all requiring premature removal of their catheters. There were in total 6161 catheter days, with the incidence rate of catheter‐related sepsis of 1.2 per 1000 catheter days. The infecting organisms were heterogeneous, with the majority being Gram‐positive cocci (47%). Our study found that patients who developed neutropaenia within 30 days after insertion were significantly associated with catheter‐related sepsis (P < 0.05).

Conclusion:  Insertion of TCVC under ultrasound guidance is a safe method with a high success rate. Incidence of catheter‐related sepsis of TCVC under percutaneous guidance was comparable to the open method. Neutropaenia developing within 30 days after ultrasound‐guided percutaneous insertion of central venous catheter may be associated with increased risk of catheter‐related sepsis.
AN EXPLORATION OF MEMBER ROLES AS A MULTILEVEL LINKING MECHANISM FOR INDIVIDUAL TRAITS AND TEAM OUTCOMES
Authors: STEWART GREG L. , FULMER INGRID S. , BARRICK MURRAY R.
Published: May 2005

We use data from 220 individuals in 45 teams to examine team member roles as a cross‐level linking mechanism between personality traits and team‐level outcomes. At the individual level, peer ratings of task role behavior relate positively with Conscientiousness and negatively with Neuroticism and Extraversion. Peer ratings of social role behavior relate positively with Agreeableness and negatively with Openness to Experience. At the team level, a composition process of aggregation operates such that the mean for social roles corresponds with social cohesion. Compilation processes of aggregation also occur, as the variance of social roles corresponds negatively with task performance, and the variance of task roles corresponds negatively with cohesion. Skew of the distribution for social roles within each team—a measure of critical mass of members individually enacting the role—also correlates with social cohesion.
THE RELATIONSHIP BETWEEN HR PRACTICES AND FIRM PERFORMANCE: EXAMINING CAUSAL ORDER
Authors: WRIGHT PATRICK M. , GARDNER TIMOTHY M. , MOYNIHAN LISA M. , ALLEN MATHEW R.
Published: May 2005

Significant research attention has been devoted to examining the relationship between HR practices and firm performance, and research support has assumed HR as the causal variable. Using data from 45 business units (with 62 data points), this study examines how measures of HR practices correlate with past, concurrent, and future operational performance measures. The results indicate that correlations with performance measures at all 3 times are both high and invariant, and that controlling for past or concurrent performance virtually eliminates the correlation of HR practices with future performance. Implications are discussed.
PREDICTORS OF OBJECTIVE AND SUBJECTIVE CAREER SUCCESS: A META‐ANALYSIS
Authors: NG THOMAS W. H. , EBY LILLIAN T. , SORENSEN KELLY L. , FELDMAN DANIEL C.
Published: May 2005

Using the contest‐ and sponsored‐mobility perspectives as theoretical guides, this meta‐analysis reviewed 4 categories of predictors of objective and subjective career success: human capital, organizational sponsorship, sociodemographic status, and stable individual differences. Salary level and promotion served as dependent measures of objective career success, and subjective career success was represented by career satisfaction. Results demonstrated that both objective and subjective career success were related to a wide range of predictors. As a group, human capital and sociodemographic predictors generally displayed stronger relationships with objective career success, and organizational sponsorship and stable individual differences were generally more strongly related to subjective career success. Gender and time (date of the study) moderated several of the relationships examined.
SYNTHETIC VALIDITY: PAST, PRESENT, AND FUTURE
Authors: SCHERBAUM CHARLES A.
Published: May 2005

Practitioners in the area of personnel selection can be confronted with situations in which the use of traditional validation strategies is not feasible. This situation can result from small numbers of incumbents, rapidly changing jobs, insufficient resources, or the lack of high‐quality criterion data. Although traditional strategies are not possible, a set of techniques falling under the rubric of synthetic validity can be used to estimate predictor validity in these situations. Synthetic validity is a logical process of inferring validity on the basis of the relationships between components of a job and tests of the attributes that are needed to perform the job components. Unfortunately, synthetic validity is infrequently used and not well understood. This article provides an overview and conceptualization of synthetic validity, reviews the historical approaches to synthetic validity, describes the recent developments and trends, and suggests future directions for synthetic validity application and research. The relationship of synthetic validity with other validity concepts, the advantages, the limitations, and the legal issues are considered throughout.
CONSEQUENCES OF INDIVIDUALS' FIT AT WORK: A META‐ANALYSIS OF PERSON–JOB, PERSON–ORGANIZATION, PERSON–GROUP, AND PERSON–SUPERVISOR FIT
Authors: KRISTOF‐BROWN AMY L. , ZIMMERMAN RYAN D. , JOHNSON ERIN C.
Published: May 2005

This meta‐analysis investigated the relationships between person–job (PJ), person–organization (PO), person–group, and person–supervisor fit with preentry (applicant attraction, job acceptance, intent to hire, job offer) and postentry individual‐level criteria (attitudes, performance, withdrawal behaviors, strain, tenure). A search of published articles, conference presentations, dissertations, and working papers yielded 172 usable studies with 836 effect sizes. Nearly all of the credibility intervals did not include 0, indicating the broad generalizability of the relationships across situations. Various ways in which fit was conceptualized and measured, as well as issues of study design, were examined as moderators to these relationships in studies of PJ and PO fit. Interrelationships between the various types of fit are also meta‐analyzed. 25 studies using polynomial regression as an analytic technique are reviewed separately, because of their unique approach to assessing fit. Broad themes emerging from the results are discussed to generate the implications for future research on fit.
New aspects of vascular remodelling: the involvement of all vascular cell types
Authors: McGrath John C. , Deighan Clare , Briones Ana M. , Shafaroudi Majid Malekzadeh , McBride Melissa , Adler Jeremy , Arribas Silvia M. , Vila Elisabet , Daly Craig J.
Published: May 2005

Conventionally, the architecture of arteries is based around the close‐packed smooth muscle cells and extracellular matrix. However, the adventitia and endothelium are now viewed as key players in vascular growth and repair. A new dynamic picture has emerged of blood vessels in a constant state of self‐maintenance. Recent work raises fundamental questions about the cellular heterogeneity of arteries and the time course and triggering of normal and pathological remodelling. A common denominator emerging in hypertensive remodelling is an early increase in adventitial cell density suggesting that adventitial cells drive remodelling and may initiate subsequent changes such as re‐arrangement of smooth muscle cells and extracellular matrix. The organization of vascular smooth muscle cells follows regular arrangements that can be modelled mathematically. In hypertension, new patterns can be quantified in these terms and give insights to how structure affects function. As with smooth muscle, little is known about the organization of the vascular endothelium, or its role in vascular remodelling. Current observations suggest that there may be a close relationship between the helical organization of smooth muscle cells and the underlying pattern of endothelial cells. The function of myoendothelial connections is a topic of great current interest and may relate to the structure of the internal elastic lamina through which the connections must pass. In hypertensive remodelling this must present an organizational challenge. The objective of this paper is to show how the functions of blood vessels depend on their architecture and a continuous interaction of different cell types and extracellular proteins.
Kv7/KCNQ/M and HCN/h, but not KCa2/SK channels, contribute to the somatic medium after‐hyperpolarization and excitability control in CA1 hippocampal pyramidal cells
Authors: Gu Ning , Vervaeke Koen , Hu Hua , Storm Johan F.
Published: May 2005

In hippocampal pyramidal cells, a single action potential (AP) or a burst of APs is followed by a medium afterhyperpolarization (mAHP, lasting ∼0.1 s). The currents underlying the mAHP are considered to regulate excitability and cause early spike frequency adaptation, thus dampening the response to sustained excitatory input relative to responses to abrupt excitation. The mAHP was originally suggested to be primarily caused by M‐channels (at depolarized potentials) and h‐channels (at more negative potentials), but not SK channels. In recent reports, however, the mAHP was suggested to be generated mainly by SK channels or only by h‐channels. We have now re‐examined the mechanisms underlying the mAHP and early spike frequency adaptation in CA1 pyramidal cells by using sharp electrode and whole‐cell recording in rat hippocampal slices. The specific M‐channel blocker XE991 (10 μm) suppressed the mAHP following 1–5 APs evoked by current injection at −60 mV. XE991 also enhanced the excitability of the cell, i.e. increased the number of APs evoked by a constant depolarizing current pulse, reduced their rate of adaptation, enhanced the afterdepolarization and promoted bursting. Conversely, the M‐channel opener retigabine reduced excitability. The h‐channel blocker ZD7288 (4‐ethylphenylamino‐1,2‐dimethyl‐6‐methylaminopyrimidinium chloride; 10 μm) fully suppressed the mAHP at −80 mV, but had little effect at −60 mV, whereas XE991 did not measurably affect the mAHP at −80 mV. Likewise, ZD7288 had little or no effect on excitability or adaptation during current pulses injected from −60 mV, but changed the initial discharge during depolarizing pulses injected from −80 mV. In contrast to previous reports, we found that blockade of Ca2+‐activated K+ channels of the SK/KCa type by apamin (100–400 nm) failed to affect the mAHP or adaptation. A computational model of a CA1 pyramidal cell predicted that M‐ and h‐channels will generate mAHPs in a voltage‐dependent manner, as indicated by the experiments. We conclude that M‐ and h‐channels generate the somatic mAHP in hippocampal pyramidal cells, with little or no net contribution from SK channels.
An Optimality Criterion for the Structural Optimization of Machine Elements
Authors: Teng C.-P. , Angeles J.
Published: May 2005

Methods of structural optimization have been studied and developed over the last three decades. An important aspect of structural optimization pertains to the condition under which the loads are applied. Most machine structures in operation are subject to loads varying as functions of time. In this paper, a novel approach is proposed to cope with loads whose magnitudes vary within given bounds and with variable directions. The underlying ideas are applied to the structural optimization of the roller-carrying disk of a novel class of cam-follower speed reduction devices termed Speed-o-Cam (SoC). Results obtained in this paper are compared with a current prototype and with an intermediate design in which the dimensions of the roller pins are optimized. Combined with the optimum dimension of the roller pins, our structural-optimization results lead to an improvement of almost twice the stiffness with a mass reduction of 40% of the original prototype.
Interactive Product Design Selection With an Implicit Value Function
Authors: Maddulapalli K. , Azarm S. , Boyars A.
Published: May 2005

We present a new method to aid a decision maker (DM) in selecting the “most preferred” from a set of design alternatives. The method is deterministic and assumes that the DM’s preferences reflect an implicit value function that is quasi-concave. The method is interactive, with the DM stating preferences in the form of attribute tradeoffs at a series of trial designs, each a specific design under consideration. The method is iterative and uses the gradient of the value function obtained from the preferences of the DM to eliminate lower value designs at each trial design. We present an approach for finding a new trial design at each iteration. We provide an example, the design selection for a cordless electric drill, to demonstrate the method. We provide results showing that (within the limit of our experimentation) our method needs only a few iterations to find the most preferred design alternative. Finally we extend our deterministic selection method to account for uncertainty in the attributes when the probability distributions governing the uncertainty are known.