bluuebunny/crossref_metadata_2025 huggingface dataset:
Providing access to electronic information resources in further education
Authors: Banwell Linda , Ray Kathryn , Coulson Graham , Urquhart Christine , Lonsdale Ray , Armstrong Chris , Thomas Rhian , Spink Siân , Yeoman Alison , Fenton Roger , Rowley Jennifer
Published: August 2004

This article aims to provide a baseline for future studies on the provision  and support for the use of digital or electronic information services (EIS) in further education. The analysis presented is based on a multi‐level model of access, which encompasses access to and availability of information and communication technology (ICT) resources, access to and availability of EIS resources, and the third leg of staff skills and their development. The research was conducted within the third cycle of the JISC (Joint Information Services Committee) User Behaviour Monitoring and Evaluation Framework, in 2001/2002. Evidence was gathered from library and information service web sites and various stakeholders, including library and information service staff, academic staff and students to generate insights into the provision of access to EIS in further education. Sector‐wide funding initiatives have had a significant impact on ICT infrastructures, and these attract a positive response from students. EIS are represented on some library web sites but both web site development and EIS availability is very much less advanced than in higher education. Staff, however, lack sufficient dedicated access to ICT to be able to develop their own skills and use. There remains a low level of access to electronic information resources, with only limited access to these resources through library web sites. LIS managers face a number of challenges in enhancing this provision, including licensing arrangements, tight budgets that need to be spread across many discipline areas, and the absence of EIS designed specifically for the further education student. The other key challenge lies in the provision of time and opportunity for academic and LIS staff to develop their ICT and EIS skills, and, more generally in the further development of the role of Information and Learning Technology (ILT) Champions.
Challenges and issues in designing inquiry on the Web
Authors: Lim Byung‐Ro
Published: August 2004

The purpose of this study is to discuss major issues in designing inquiries on the web. Instructional designers face a lot of difficulties and challenges to deal with in the course of designing inquiry experiences for learners in the online environment, a complex and ill‐structured learning environment. Facilitating inquiry or using an inquiry‐based learning approach on the web has both advantages and disadvantages. Instructional designers need to acknowledge the design issues arising in each and every phase of the design process. Utilising appropriate design strategies, instructional designers can provide positive learning experiences for their online learners. For this purpose, three representative cases in the context of teacher professional development were examined and six major issues were identified as follows: (1) seeking a balance between a system‐generated guide and human facilitator, (2) visual representation of the inquiry process, (3) motivating learners with the right question, (4) engaging learners in various learning activities, (5) guiding the inquiry process with various scaffolds, and (6) maximising learning by coordinating resources, tools and the community of inquiry. This study explains each issue in great detail and provides possible guides for instructional designers to handle the issue.
Personality, Identity Styles, and Religiosity: An Integrative Study Among Late Adolescents in Flanders (Belgium)
Authors: Duriez Bart , Soenens Bart , Beyers Wim
Published: August 2004

AbstractThe relationship between the two main dimensions of the religiosity domain (Exclusion vs. Inclusion of Transcendence and Literal vs. Symbolic) and both the Five‐Factor Model of personality andBerzonsky's (1990)identity styles was investigated in a Flemish sample of late adolescents (N=335). The results show that, whereas Exclusion vs. Inclusion is unrelated to any of the personality dimensions, Literal vs. Symbolic was strongly related to Openness to Experience and moderately to Agreeableness. Further, it was shown that Exclusion vs. Inclusion was positively related to the normative identity style, and that Literal vs. Symbolic correlated positively with the informational identity style and negatively with the diffuse/avoidant identity style. As expected, the relation between Openness to Experience and Literal vs. Symbolic was fully mediated by the informational identity style. Once Openness to Experience was taken into account, Agreeableness was no longer an important determinant of Literal vs. Symbolic.
Defensive Copers Show a Deficit in Passive Avoidance Learning on Newman's Go/No‐Go Task: Implications for Self‐Deception and Socialization
Authors: Shane Matthew S. , Peterson Jordan B.
Published: August 2004

Abstract
The present study investigated whether passive avoidance learning was retarded by defensive coping strategies designed to minimize exposure to negatively valenced stimuli. High‐anxious individuals, low‐anxious individuals, and defensive copers completed a computerized go/no‐go task, in which they learned when to press or not to press a button, in response to contingent positive and negative feedback. The duration that feedback remained onscreen was self‐regulated. Defensive copers showed preferential reflection away from negative feedback, committed more passive‐avoidance errors, and were characterized by impaired learning, overall. Further, the ratio of reflection on negative feedback to reflection on positive feedback directly mediated both passive‐avoidance errors and overall learning. Defensive coping strategies, therefore, appear to interfere with passive avoidance learning, thereby fostering perseverative, dysfunctional action patterns by reducing knowledge gained from previous mistakes. Implications for the learning of effective socialization strategies, and for psychopathy—which is commonly characterized by similar passive‐avoidance deficits—are subsequently considered.
Personality, Threat, and Cognitive and Emotional Reactions to Stressful Intercultural Situations
Authors: Van Der Zee Karen , Van Oudenhoven Jan Pieter , De Grijs Ellen
Published: August 2004

The present study examined individual differences in appraisal of and affective reactions to intercultural situations. A sample of 160 students filled out the Multicultural Personality Questionnaire (MPQ) and participated in an experiment in which they received a description of an intercultural situation that was either high or low in potential stressfulness. Individuals with high scores on the intercultural dimensions appraised the potentially stressful situation more positively and showed more positive and less negative reactions to the situation than did individuals with low scores on the MPQ. Interestingly, following a Terror Management Intervention (TMI), individual differences in emotional reactions to intercultural situations disappeared. The results could only partially be replicated using a general personality questionnaire, suggesting that these findings have at least some specificity to intercultural personality dimensions.
Fracture risk of women with primary biliary cirrhosis: no increase compared with general population controls
Authors: Boulton‐Jones J. R. , Fenn R. M. F. , West J. , Logan R. F. A. , Ryder S. D.
Published: August 2004

Aim : Patients with primary biliary cirrhosis may be at increased risk of osteoporosis but to what extent this is reflected in an increased fracture risk is unknown. We have enquired about the fracture experience of female primary biliary cirrhosis patients compared with sex‐ and age‐matched controls.

Methods : Patients aged 30–75 with primary biliary cirrhosis and age‐matched controls were sent a postal questionnaire asking about their fracture history and details of risk factors for osteoporosis.

Results : 85 eligible patients with primary biliary cirrhosis and 116 controls responded. Forty‐one per cent of patients with primary biliary cirrhosis and 30% of controls reported ever having had a fracture odds ratio 1.5 (95% confidence interval: 0.80–2.89). Twenty‐eight per cent of primary biliary cirrhosis patients and 23.3% of controls reported a fracture after the age of 30, odds ratio 1.2 (95% confidence interval: 0.57–2.56), and 14.1% of primary biliary cirrhosis patients and 12.1% of controls reported a low impact fracture of the long bones or of the vertebrae odds ratio 1.0 (95% confidence interval: 0.31–2.68).

Conclusions : No overall increased fracture risk in patients with primary biliary cirrhosis was observed. As a group, unselected patients with primary biliary cirrhosis do not represent a population at particularly high risk of osteoporotic fracture and thus targeting them for osteoporosis screening and treatment is not justified. Further work investigating subgroups of patients with primary biliary cirrhosis at potentially high risk of osteoporosis, such as those with advanced disease or severe cholestasis is required.
Effect of acid on duodenal blood flow and mucus secretion measured by reflectance spectrophotometry: a prospective, randomized‐controlled study
Authors: Ramirez F. C. , Holland J. F. , Harker J. , Leung F. W.
Published: August 2004

Background : In animals, hydrochloric acid increases blood flow and mucus secretion in the duodenal mucosa. A significant correlation between index of haemoglobin oxygen saturation and mucosal blood flow, and between change in index of haemoglobin concentration and mucus thickness, respectively, has been demonstrated by reflectance spectrophotometry.

Aim : To examine the effect of topical hydrochloric acid upon mucosal blood flow and mucus secretion in the human duodenum.

Methods : This prospective study of 120 patients undergoing routine upper endoscopy, examined the effect of topical 0.1 n hydrochloric acid or 0.9% saline on the duodenal bulb in a randomized, double‐blind fashion. Duodenal mucosal index of haemoglobin oxygen saturation and index of haemoglobin concentration were measured by endoscopic reflectance spectrophotometry before and after hydrochloric acid or saline.

Results : Baseline index of haemoglobin oxygen saturation, calculated blood flow and index of haemoglobin concentration measurements were comparable between hydrochloric acid (n = 60) and saline (n =60) treated groups. A history of current use of non‐steroidal anti‐inflammatory drug was associated with a significantly lower baseline index of haemoglobin oxygen saturation and calculated blood flow. Hydrochloric acid resulted in a significant increase in index of haemoglobin oxygen saturation and calculated blood flow, but a decrease in index of haemoglobin concentration, reflecting an increase in mucus thickness compared with saline.

Conclusions : Our observations in humans confirm data in animal studies that topical exposure to hydrochloric acid induces an increase in duodenal mucosal blood flow and mucus secretion. Post hoc analysis of the data also revealed that attenuation of basal duodenal mucosal blood flow is associated with a history of current non‐steroidal anti‐inflammatory drug use. Endoscopic reflectance spectrophotometry appears to be adequate to assess factors that influence duodenal defence mechanisms of blood flow and mucus secretion in humans.
Helicobacter pyloriinfection prevents the occurrence of the tolerance phenomenon of histamine H2 receptor antagonists
Authors: Fujisawa T. , Adachi K. , Komazawa Y. , Mihara T. , Azumi T. , Katsube T. , Furuta K. , Kazumori H. , Kinoshita Y.
Published: August 2004

Background: The attenuated anti‐secretory activity of H2 receptor antagonists (H2RA) during continuous administration is referred to as the tolerance phenomenon. However, it is not clarified whetherHelicobacter pyloriinfection affects the occurrence of tolerance to H2RA. It is also not clarified whether the tolerance phenomenon occurs to a new H2RA, lafutidine.

Aim: To investigate the occurrence of the tolerance phenomenon in subjects with and withoutH. pyloriinfection during the continuous administration of lafutidine and famotidine.

Subjects and Methods: Subjects were 20 healthy male volunteers (sevenH. pyloripositive and 13H. pylorinegative cases). All subjects were examined by ambulatory intragastric pH monitoring five times without medication, on the first and 15th day of the administration of 20 mg b.d. famotidine and 10 mg b.d. lafutidine in a cross‐over fashion.

Results: The tolerance phenomenon was not observed inH. pylori‐positive subjects during the 15‐day‐long administration of both H2RAs. In contrast, the tolerance phenomenon was observed inH. pylorinegative subjects, which has been previously reported.

Conclusions: This study demonstrated thatH. pyloriinfection affects the tolerance phenomenon during continuous administration of H2RAs.
Relationship between body mass index, diet, exercise and gastro‐oesophageal reflux symptoms in a community
Authors: Nandurkar S. , Locke G. R. , Fett S. , Zinsmeister A. R. , Cameron A. J. , Talley N. J.
Published: August 2004

Background : Body mass index (BMI) is a risk factor for gastro‐oesophageal reflux but may simply be explained by diet and lifestyle.

Aim : We aimed to determine the contribution of BMI, diet and exercise to GER.

Methods : Community subjects (n = 211, mean age =36 years, 43% males) completed validated questionnaires on gastro‐oesophageal reflux, energy expenditure (Harvard Alumni Activity Survey), dietary intake (Harvard Food Frequency Questionnaire) and measures of personality and life event stress. Diet, exercise, BMI and other potential risk factors for reflux were analysed using logistic regression analyses.

Results : The overall mean (±s.d.) BMI was 26.6 (±5.7); 79 (37%) reported infrequent ( 25, 10% reported frequent reflux compared to 4% of those with BMI ≤ 25. In a model which included age, sex and Symptom Checklist‐90 somatisation T‐score, BMI was associated with reflux (OR per 5 units = 1.9, 95% CI: 1.2, 3.0). In models which included diet and exercise variables, BMI but not diet or exercise was associated with reflux.

Conclusion : BMI may be associated with symptomatic gastro‐oesophageal reflux independent of diet and exercise.
Structure and Activity of a Chimeric Interleukin‐8‐Melanoma‐Growth‐Stimulatory‐Activity Protein
Authors: Sticht Heinrich , Auer Manfred , Schmitt Bettina , Besemer Jürgen , Horcher Markus , Kirsch Thomas , Lindley Ivan J. D. , Rösch Paul
Published: August 2004

A 72‐amino‐acid chimeric protein, Chi1, was constructed from the N‐terminal part of interleukin 8, IL‐8‐(1–53), and the C‐terminal part of melanoma growth stimulatory activity, MGSA‐(54–72). Chi1 protein showed receptor‐binding specificity and biological activity similar, but not identical to IL‐8 and decidedly different from MGSA. The structure of Chi1 was determined in solution by two‐dimensional NMR and molecular‐dynamics calculations. The structure resembled the structures of MGSA and IL‐8 closely, containing a triple‐stranded β‐sheet in the IL‐8 part and an amphipathic α‐helix in the MGSA part. Chi1 formed dimers at millimolar concentrations via the first strand from the N‐terminus, analogous to IL‐8 and MGSA. In contrast to the latter molecules, however, the α‐helix of Chi1 did not pack against the β‐sheet part, but was an independent structural element. This structural difference could be explained mainly by the modulation of hydrophobic interactions between the helix and the rest of the protein in Chi1 as compared to IL‐8 and MGSA. It is concluded that tight helix packing is not required for receptor binding and biological activity of Chi1.
Asp70 in the Peripheral Anionic Site of Human Butyrylcholinesterase
Authors: Masson Patrick , Froment Marie‐Thérèse , Bartels Cynthia F. , Lockridge Oksana
Published: August 2004

The goal of this work was to determine what amino acids at the mouth of the active‐site gorge are important for the function of human butyrylcholinesterase. Mutants D70G, Q119Y, G283D, A277W, A277H and A277W/G283D were expressed in human embryonal kidney cells and the secreted enzymes were assayed by steady‐state kinetics. The result was that only one amino acid, D70 was found to be important for function. When D70 was mutated to G, the same mutation as in the naturally occurring atypical butyrylcholinesterase, the affinity for positively charged substrates and positively charged inhibitors decreased 5–30‐fold. The D70G mutant had another striking abnormality in that it was virtually devoid of the phenomenon of substrate activation by excess butyrylthiocholine. Thus, though kcat, was the same for wild‐type and D70G mutant, being 24000 min−1 at low butyrylthiocholine concentrations (0.01–0.1 mM), it failed to increase for the D70G mutant at 40 mM butyrylthiocholine, whereas it increased threefold for wild type. The D70C mutant was more sensitive to changes in salt concentration, its catalytic rate decreasing more than that of the wild type. The D70G mutant appeared to have a greater surface negative charge than wild type suggesting that the D70G mutant had a conformation different from that of the wild type. That D70 affects the function of butyrylcholinesterase, together with its location at the mouth of the active‐site gorge, supports the hypothesis that D70 is a component of the peripheral anionic site of butyrylcholinesterase. Mutants containing aromatic amino acids at the mouth of the gorge had increased binding affinity for propidium and fasciculin, but unaltered function, suggesting that aromatic amino acids are not important to the function of the peripheral anionic site of butyrylcholinesterase.
Glutamine Increases Argininosuccinate Synthetase mRNA Levels in Rat'Hepatocytes
Authors: Quillard Muriel , Husson Annie , Lavoinne Alain
Published: August 2004

Glutamine, one of the most efficient substrates in the urea cycle, was found to induce the accumulation of argininosuccinate synthetase (ASS) mRNA in primary cultured rat hepatocytes. The inducing action of glutamine was obtained at various stages of development and a half‐maximal effect was observed at about 3 mM glutamine. This effect was apparent from 6 h and persisted for at least 48 h. NH4Cl addition up to 5 mM did not significantly change the ASS mRNA level. Like glutamine, hypoosmotic medium and aminoisobutyric acid (conditions known to increase cell volume) also increased the ASS mRNA level, which was, in contrast, decreased by hyperosmotic medium. These results demonstrate that the glutamine‐induced swelling may participate in the observed increase of the ASS mRNA level. This increase in the ASS mRNA level was associated with an increase in ASS activity.
Effects of Pancreatic Spasmolytic Polypeptide (PSP) on Epithelial Cell Function
Authors: Otto William R. , Rao Jagdish , Cox Helen M. , Kotzian Ewald , Lee Chung. Y. , Goodlad Robert A. , Lane Andrew , Gorman Michael , Freemont Paul A. , Hansen Hans F. , Pappin Darryl , Wright Nicholas A.
Published: August 2004

Trefoil peptides are expressed near endodermal ulcerations and may modulate epithelial repair. The trefoil pancreatic spasmolytic polypeptide (PSP) was tested for growth activity in vitro on epithelial cells and in vivo following intragastric or intravenous infusion in parenterally fed intact rats. Ion transport was assessed as changes in short‐circuit current in rat intestine and adenocarcinoma cells in Ussing chambers. PSP stimulated growth of MCF‐7 and Colo‐357 cells, but only in the presence of extracellular glutathione (GSH). The effect was attenuated by GSH depletion with buthionine sulphoximine, even in GSH‐containing media. When GSH‐reduced PSP was carboxymethylated with iodoacetic acid, it still depended on extracellular GSH for its growth effect. Intestinal epithelial proliferation in rats was not affected by either intravenous or intraluminal infusion. PSP had no effect on basal or stimulated ion flux in rat jejunum or epithelial monolayers. The peptide did not compete with 125I‐labeled epidermal growth factor for its receptor. [14C]Iodoacetamide treatment of PSP, followed by prolonged tryptic digestion yielded predominantly a 14C‐labeled tetrapeptide fragment containing Cys104, with a lesser quantity of a 14C‐labeled 15‐amino‐acid peptide containing Cys95 (molar ratio 15:1). GSH may predominantly reduce the Cys6‐Cys104 terminal disulphide bond in PSP. We conclude that some epithelia may exhibit a growth response to PSP if extracellular GSH is present. Reduction of PSP by GSH is not necessary for this response, suggesting that the trefoil receptor or its signal transduction is GSH sensitive. PSP could assist wound healing by interactions with epithelial cells exposed concurrently to a local high GSH concentration.
Diradylglycerol Formation in Cholecystokinin‐Stimulated Rabbit Pancreatic Acini
Authors: Hermans Stefanie W. G. , Engelmann Bernd , Reinhardt Ulrike , Bartholomeus‐Van Nooij Irene G. P. , De Pont Jan Joep H. H. M. , Willems Peter H. G. M.
Published: August 2004

The aim of the present study was to assess the origin of the 1,2‐diradylglycerols produced during prolonged hormonal stimulation of rabbit pancreatic acini by comparison of their relative molecular species composition with that of the major acinar phospholipids. Both phosphatidylcholine (PtdCho) and phosphatidylethanolamine (PtdEtn) consisted of 1,2‐diacyl as well as 1‐alk‐1‐enyl‐2‐acyl species. In contrast, phosphatidylinositol (PtdIns), phosphatidylserine and phosphatidic acid existed only in the 1,2‐diacyl form. Acinar cells did not contain detectable amounts of 1‐alkyl‐2‐acyl phospholipids. Similarly, the acinar 1,2‐diradylglycerol fraction consisted of 1,2‐diacylglycerols and 1‐alk‐1‐enyl‐2‐acylglycerols. Mass 1,2‐diradylglycerol measurements revealed that prolonged stimulation with cholecystokinin resulted in a marked and sustained increase in acinar 1,2‐diradylglycerol content. Based on the relative amounts of the 1,2‐diacyl species present in both the 1,2‐diradylglycerol fraction and the individual phospholipids, it is calculated that under control conditions 60% of the 1,2‐diacylglycerols originate from PtdCho and 40% from PtdIns, whereas under stimulatory conditions 53% is calculated to be derived from PtdCho, 46% from PtdIns and 1% from PtdEtn. Likewise, it is calculated that in control as well as stimulated acini 100% of the 1‐alk‐1‐enyl‐2‐acylglycerols originate from plasmenylcholine. Further evidence in favour of the idea that at least a considerable part of the 1,2‐diacylglycerols produced during prolonged hormonal stimulation originate from inositolphospholipids is provided by the observation that labeling of phosphatidylinositol 4,5‐bisphosphate with inorganic phosphate reached isotopic equilibrium markedly faster under stimulatory conditions as compared to the control situation, which is in agreement with an elevated turnover rate. The data presented support the idea that PtdCho and inositolphospholipids are the major precursors in basal and stimulated 1,2‐diradylglycerol production in rabbit pancreatic acini.
Site‐Directed Mutagenesis of Conserved Charged Residues in the Helical Region of the Human C5a Receptor
Authors: Raffetseder Ute , Röper Detlef , Mery Laurence , Gietz Claudia , Klos Andreas , Grötzinger Joachim , Wollmer Axel , Boulay François , Köhl Jörg , Bautsch Wilfried
Published: August 2004

The human C5a receptor (C5aR) belongs to the family of G‐protein‐coupled receptors with seven transmembrane helices. This part of the molecule is thought to contain part of the ligand‐binding pocket, specifically to bind the C‐terminal Arg of human C5a. Guided by sequence similarity and molecular modelling studies, several residues including polar (Asn119, Thr168, Gln259) as well as all conserved charged amino acids in the upper transmembrane region of the C5aR (Asp37, Asp82, Arg175, Arg206, Asp282) were exchanged by site‐directed mutagenesis. Receptor mutants were transiently expressed in COS cells and analyzed for altered binding behaviour and/or localization at the cell surface by immunofluorescence. For all residues, suitable mutants could be found that exhibited wild‐type affinity towards the ligand, providing evidence against a major contribution of these residues to high‐affinity ligand binding. Some mutants, however, exhibited a complete (Asp282→Ala) or partial loss of ligand‐binding capacity (Arg175→Ala, Arg206→Gln) despite adequate expression levels on the cell surface. This phenotype was further analyzed in the [Gln206]C5aR mutant: quantitative flow cytometric analysis of epitope‐tagged receptor derivatives in 293 cells confirmed an equal level of wild‐type and mutant C5aR on the cell surface. Competitive binding curves revealed the presence of only a small population (<10%) of high‐affinity sites (Kd≈2nM), which was functionally active at 20 nM in the heterologous Xenopus oocyte expression system after coexpression of Gα‐16. The number of high‐affinity sites of wild‐type and [Gln206]C5aR in 293 cells could be up‐regulated by coexpression of Giα‐2 and down‐regulated by GTP[γS]‐mediated uncoupling of the G‐protein receptor interaction in membrane preparations. These findings are compatible with a model in which the Arg206 residue located in the upper third of transmembrane helix V determines high‐affinity binding in the human C5aR by affecting the intracellular G‐protein coupling.
NMR Data Show that the Carcinogen N‐2‐Acetylaminofluorene Stabilises an Intermediate of −2 Frameshift Mutagenesis in a Region of High Mutation Frequency
Authors: Milhé Catherine , Fuchs Robert P. P. , Lefèvre Jean‐François
Published: August 2004

The heteroduplex, d(ACCGGCGCC ACA) · d(TGTGG–CCGGT), containing two bulged bases, a cytosine and the guanine G7, either unmodified or modified with the carcinogen N‐2‐acetylaminofluorene, have been studied by NMR as models of slipped‐mutagenic intermediates (SMI). The melting temperature of the modified heteroduplex is strongly increased compared with that of the unmodified heteroduplex. NMR studies have shown that all the bases of the unmodified heteroduplex are stacked within the helix, without any disruption of the sequential connectivities. The two strands are in a B‐like conformation. Nevertheless, exchangeable‐proton studies have revealed that base pairing is very weak, or even lacking, over two base pairs apart from the bulge. Concerning the modified heteroduplex, no B‐like connectivity is observed in the G5–C9 segment. Moreover, the cytosine C8 is rejected outside the helix, whereas the N‐2‐acetylaminofluorene moiety is inserted within the helix. The G5 · C18, C6 · G17 and C9 · G16 bases are remarkably stable when the temperature is increased, in agreement with the high melting temperature. Some small unassigned peaks reveal the presence of the minor conformation in equilibrium. The strong stabilisation of the N‐2‐acetylaminofluorene‐modified heteroduplex compared with the unmodified duplex is in agreement with the high N‐2‐acetylaminofluorene‐induced mutation frequency compared with the spontaneous frequency and with the hypothesis of mutagenesis occurring during replication.
The NeuAc(α‐2,6)‐Gal/GalNAc‐Binding Lectin from Elderberry (Sambucus Nigra) Bark, a type‐2 Ribosome‐Inactivating Protein with an Unusual Specificity and Structure
Authors: Van Damme Els J. M. , Barre Annick , Rougé Pierre , Van Leuven Fred , Peumans Willy J.
Published: August 2004

The cDNA encoding the NeuAc(α‐2,6)Gal/GalNAc binding lectin from elderberry (Sambucus nigra) bark (SNAI) was isolated from a cDNA library constructed with mRNA from the bark. Sequence analysis of this lectin cDNA revealed a striking similarity to the previously sequenced type‐2 ribosome‐inactivating proteins from Ricinus communis and Abrus precatorius. Molecular modelling of SNAI further indicated that its structure closely resembles that of ricin. Since SNAI strongly inhibits cell‐free protein synthesis in a rabbit reticulocyte lysate it presumably is a type‐2 ribosome‐inactivating protein. However, SNAI differs from all previously described type‐2 ribosome‐inactivating proteins by its specificity towards NeuAc(α‐2,6)Gal/GalNAc and its unusual molecular structure.
Properties of the Human Nuclear Protein p85Mcm
Authors: Schulte Dorothea , Richter Andreas , Rurkhart Roland , Musahl Christine , Knippers Rolf
Published: August 2004

Recently we identified a cDNA fragment encoding a conserved part of a new human minichromosome maintenance (Mcm) protein, provisionally termed P1.1Mcm3. Here, we report that the protein is most highly related to a yeast cell‐division‐cycle protein, Cdc47, encoded by the open reading frame YBR1441 on chromosome II of Saccharomyces cerevisiae. The human protein migrates on a polyacrylamide gel with an apparent molecular mass of 85 kDa and shares areas of significant similarity with the Mcm family of replication proteins. It is, therefore, designated as p85Mcm. Microscopic immuno‐fluorescence studies revealed that protein p85Mcm is located in the nuclei of interphase cells, but is evenly distributed throughout the cell during mitosis. The amounts of p85Mcm do not significantly change during the cell cycle, but mRNA levels rise with the beginning of the S phase. However, in vitro differentiation of HL60 cells results in a striking decrease of both p85Mcm mRNA and protein levels, suggesting a role for p85Mcm in proliferating, but not in differentiated cells. Under physiological salt conditions, p85Mcm is a component of a high molecular‐mass complex including other Mcm proteins. The complex dissociates at high ionic strength given rise to stable subcomplexes, one of which contains protein p85Mcm together with Mcm proteins hCdc21 and p105Mcm.
Quinaldine 4‐Oxidase from Arthrobacter sp. Rü61a, a Versatile Procaryotic Molybdenum‐Containing Hydroxylase Active towards N‐Containing Heterocyclic Compounds and Aromatic Aldehydes
Authors: Stephan Ingrid , Tshisuaka Barbara , Fetzner Susanne , Lingens Franz
Published: August 2004

Quinaldine 4‐oxidase from Arthrobacter sp. Rü61a, an inducible molybdenum‐containing hydroxylase, was purified to homogeneity by an optimized five‐step procedure. Molecular oxygen is proposed as physiological electron acceptor. Electrons are also transferred to artificial electron acceptors with E′o > −8 mV. The molybdo‐iron/sulfur flavoprotein regiospecifically attacks its N‐heterocyclic substrates: isoquinoline and phthalazine are hydroxylated adjacent to the N‐heteroatom at C1, whereas quinaldine, quinoline, cinnoline and quinazoline are hydroxylated at C4. Additionally, the aromatic aldehydes benzaldehyde, salicylaldehyde, vanillin and cinnamaldehyde are oxidized to the corresponding carboxylic acids, whereas short‐chain aliphatic aldehydes are not.

Quinaldine 4‐oxidase is compared to the two molybdenum‐containing hydroxylases quinoline 2‐oxidoreductase from Pseudomonas putida 86 [Tshisuaka, B., Kappl, R., Hüttermann, J. & Lingens, F. (1993) Biochemistry 32, 12928–12934] and isoquinoline 1‐oxidoreductase from Pseudomonas diminuta 7 [Lehmann, M., Tshisuaka, B., Fetzner, S., Röger, P. & Lingens, F. (1994) J. Biol. Chem. 269, 11254–11260] with respect to the substrates converted and the electron‐acceptor specificities. These dehydrogenases hydroxylate their N‐heterocyclic substrates exclusively adjacent to the heteroatom. Whereas the aldehydes tested are scarcely oxidized by quinoline 2‐oxidoreductase, isoquinoline 1‐oxidoreductase catalyzes the oxidation of the aromatic aldehydes, although being progressively inhibited. Neither quinoline 2‐oxidoreductase nor isoquinoline 1‐oxidoreductase transfer electrons to oxygen. Otherwise, the spectrum of electron acceptors used by quinoline 2‐oxidoreductase and quinaldine 4‐oxidase is identical. However, isoquinoline 1‐oxidoreductase differs in its electron‐acceptor specificity.

Quinaldine 4‐oxidase is unusual in its substrate and electron‐acceptor specificity. This enzyme is able to function as oxidase or dehydrogenase, it oxidizes aldehydes, and it catalyzes the nucleophilic attack of N‐containing heterocyclic compounds at two varying positions depending on the substrate.
Major Structural Differences between Pokeweed Antiviral Protein and Ricin A‐Chain do not Account for their Differing Ribosome Specificity
Authors: Chaddock John A. , Monzingo Arthur F. , Robertus Jon D. , Lord J. Michael , Roberts Lynne M.
Published: August 2004

Pokeweed antiviral protein (PAP) and the A‐chain of ricin (RTA) are two members of a family of ribosome‐inactivating proteins (RIPs) that are characterised by their ability to catalytically depurinate eukaryotic ribosomes, a modification that makes the ribosomes incapable of protein synthesis. In contrast to RTA, PAP can also inactivate prokaryotic ribosomes. In order to investigate the reason for this differing ribosome specificity, a series of PAP/RTA hybrid proteins was prepared to test for their ability to depurinate prokaryotic and eukaryotic ribosomes. Information from the X‐ray structures of RTA and PAP was used to design gross polypeptide switches and specific peptide insertions. Initial gross polypeptide swaps created hybrids that had altered ribosome inactivation properties. Preliminary results suggest that the carboxy‐terminus of the RIPs (PAP 219–262) does not contribute to ribosome recognition, whereas polypeptide swaps in the amino‐terminal half of the proteins did affect ribosome inactivation. Structural examination identified three loop regions that were different in both structure and composition within the amino‐terminal region. Directed substitution of RTA sequences into PAP at these sites, however, had little effect on the ribosome inactivation characteristics of the mutant PAPs, suggesting that the loops were not crucial for prokaryotic ribosome recognition. On the basis of these results we have identified regions of RIP primary sequence that may be important in ribosome recognition. The implications of this work are discussed.
Activation of Human Neutrophil Procollagenase by Stromelysin 2
Authors: Knäuper Vera , Murphy Gillian , Tschesche Harald
Published: August 2004

Neutrophil procollagenase (MMP‐8) was efficiently activated by incubation with active stromelysin 2 (MMP‐10). A single‐step activation mechanism involving the cleavage of the Gly78‐Phe79 peptide bond at the end of the propeptide domain was observed. Determination of the collagenolytic activity revealed the generation of active neutrophil collagenase displaying high specific activity. When compared with the specific activity following mercurial activation, which generates active collagenase by autoproteolytic cleavage of either Phe79‐Met80 or Met80‐Leu81 peptide bonds [Bläser, J., Knäuper, V., Osthues, A., Reinke, H. & Tschesche, H. (1991) Eur J. Biochem. 202, 1223–1230], the specific activity of the stromelysin‐2‐activated enzyme was considerably higher. Thus, human neutrophil procollagenase was ‘superactivated’ by stromelysin 2, as was recently shown for the stromelysin‐1‐activated enzyme [Knäuper, V., Wilhelm, S. M., Seperack, P. K., De Clerck, Y. A., Langley, K. E., Osthues, A. & Tschesche, H. (1993a) Biochem. J. 295, 581–586].
Purification and Characterization of Carbamoyl‐Phosphate Synthetase from the Deep‐Sea Hyperthermophilic Archaebacterium Pyrococcus abyssi
Authors: Purcarea Cristina , Simon Valérie , Prieur Daniel , Hervé Guy
Published: August 2004

Carbamoyl‐phosphate synthetase was purified from the deep‐sea hyperthermophilic archaebacterium Pyrococcus abyssi. This enzyme appears to be monomeric and uses ammonium salts as nitrogen donor. Its activity is inhibited by some nucleotides that compete with ATP. In contrast with the carbamoyl‐phosphate synthetases investigated so far, this enzyme is very resistant to high temperature. Its low molecular mass (46.6 kDa) and its catalytic properties suggest that the gene coding for this enzyme is a previously postulated ancestor whose duplication gave the genes coding for carbamoyl‐phosphate synthetases and carbamate kinases.
Purification and Characterization of an Intracellular Catalase‐Peroxidase from Penicillium Simplicissimum
Authors: Fraaije Marco W. , Roubroeks Hanno P. , Hagen Wilfred R. , Van Berkel Wiliem J. H.
Published: August 2004

The first dimeric catalase‐peroxidase of eucaryotic origin, an intracellular hydroperoxidase from Penicillium simplicissimum which exhibited both catalase and peroxidase activities, has been isolated. The enzyme has an apparent molecular mass of about 170 kDa and is composed of two identical subunits. The purified protein has a pH optimum for catalase activity at 6.4 and for peroxidase at 5.4. Both activities are inhibited by cyanide and azide whereas 3‐amino‐1,2,4‐triazole has no effect. 3,3′‐Diaminobenzidine, 3,3′‐dimethoxybenzidine, guaiacol, 2,6‐dimethoxyphenol and 2,2′‐azinobis(3‐ethylbenzothiazoline‐6‐sulfonic acid) all serve as substrates. The optical spectrum of the purified enzyme shows a Soret band at 407 nm. Reduction by dithionite results in the disappearance of the Soret band and formation of three absorption maxima at 440, 562 and 595 nm. The prosthetic group was identified as a protoheme IX and EPR spectroscopy revealed the presence of a histidine residue as proximal ligand.

In addition to the catalase‐peroxid, an atypical catalase which is active over a broad pH range was also partially purified from P. simplicissimum. This catalase is located in the periplasm and contains a chlorin‐type heme as prosthetic group.
Isolation and Structures of Glycoprotein‐Derived Free Oligosaccharides from the Unfertilized Eggs of Scyliorhinus Caniculus
Authors: Plancke Yves , Delplace Florence , Wieruszeski Jean‐Michel , Maes Emmanuel , Strecker Gérard
Published: August 2004

As previously reported [Ishii, K., Iwasaki, M., Inoue, S., Kenny, P. T. M., Komura, H. & Inoue, Y. (1989) J. Biol. Chem. 264, 1623–1630; Inoue, S., Iwasaki, M., Ishii, K., Kitajima, K. & Inoue, Y. (1989) J. Biol. Chem. 264, 18520–18526], the unfertilized eggs of two different species of fresh‐water fish, Plecoglossus altivelis and Tribodolon hakonensis, contain relatively large amounts of free sialooligosaccharides. These oligosaccharides were found to derive from glycophosphoproteins, owing to the activity of a peptide –N4‐(N ‐acetyl‐β‐d‐glucosaminyl)asparagine amidase [Iwasaki, M., Seko, A., Kitajima, K., Inoue, Y. & Inoue, S. (1992) J. Biol. Chem. 267, 24287–24296; Seko, A., Kitajima, K., Inoue, Y. & Inoue, S. (1991) J. Biol. Chem. 266, 22110–22114].

Here we describe a new type of free oligosaccharides, isolated from unfertilized eggs of Scyliorhinus caniculus. From the structural analysis, based upon 1H‐NMR spectroscopy, the following glycan units are proposed.
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Covalent Association of Lipopolysaccharide at the Hemocyte Surface of Insects is an Initial Step for its Internalization
Authors: Charalambidis Nektarios D. , Foukas Lazaros C. , Marmaras Vassilis J.
Published: August 2004

It is well known that lipopolysaccharide (LPS) of Gram‐negative bacteria triggers antibacterial responses to mammalian macrophages [Weinstein, S., Gold, M. R. & DeFranco, A. (1991) Proc. Natl Acad. Sci. USA 88, 4148–4152] and insect hemocytes [Charalambidis, N. D., Zervas, C. G., Lambropoulou, M., Katsoris, P. G. & Marmaras, V. J. (1995) Eur. J. Cell Biol. 67, 32–41], via protein‐tyrosine phosphorylation. In this study we show that insect hemocytes in response to LPS facilitate internalization of LPS (either cell‐associated or cell‐free). According to our data, the recognition and covalent association of LPS (either cell‐associated or cell‐free) to the hemocyte surface are essential initial steps for LPS internalization. LPS (Escherichia coli) recognizes membrane effector 47–kDa protein (p47) and then crosslinks to membrane‐associated p47 (mp47) via the intermediacy of tyrosine derivatives generated by the action of phenol oxidase, as is the case for cuticular protein–chitin crosslinks during sclerotization [Shaefer, J., Kramer, K. J., Garbow, J. R., Jacob, G. S., Stejskal, E. O., Hopkins, T. L. & Speirs, R. D. (1987) Science 235, 1200–1204]. The covalent association of LPS to the hemocyte surface appears to be a prerequisite for LPS internalization as judged by the resistance of LPS binding to dissociation by proteinase K. In addition, our results show that the effector molecules participating in LPS covalent association at the cell surface and LPS internalization are not involved in LPS‐induced activation of hemocytes. However, the fact that genistein, as well as the inhibitors of LPS‐dependent secretion, block LPS covalent association at the cell surface and LPS internalization provides a preliminary characterization of an LPS signal‐transduction‐dependent process which is apparently involved.
Changes in the Cellular Energy State Affect the Activity of the Bacterial Phosphotransferase System
Authors: Rohwer Johann M. , Jensen Peter Ruhdal , Shinohara Yasuo , Postma Pieter W. , Westerhoff Hans V.
Published: August 2004

The effect of different cellular free‐energy states on the uptake of methyl α‐d‐glucopyranoside, an analogue of glucose, by the Escherichia coli phosphoenolpyruvate:carbohydrate phosphotransferase system was investigated. The intracellular [ATP]/[ADP] ratio was varied by changing the expression of the atp operon, which codes for the H+‐ATPase, or by adding an uncoupler of oxidative phosphorylation or an inhibitor of respiration. Corresponding initial phosphotransferase uptake rates were determined using an improved uptake assay that works with growing cells in steady state. The results show that the initial uptake rate was decreased under conditions of lowered intracellular [ATP]/[ADP] ratios, irrespective of which method was used to change the cellular energy state. When either the expression of the atp operon was changed or 2,4‐dinitrophenol was added to wild‐type cells, the relationship between initial phosphotransferase uptake rate and the logarithm of the [ATP]/[ADP] ratio was approximately linear. These results suggest that the cellular free‐energy state, as reflected in the intracellular [ATP]/[ADP] ratio, plays an important role in regulating the activity of the phosphotransferase system.
Mitochondrial Sulfide Oxidation in Arenicola Marina
Authors: Völkel Susanne , Grieshaber Manfred K.
Published: August 2004

Sulfide is oxidized in the mitochondria of the lugworm Arenicola marina. Mitochondrial sulfide oxidation is coupled with oxygen consumption and with an equimolar production of thiosulfate [Vökel, S. & Grieshaber, M. K. (1994) Mar. Biol. 118, 137–147], Mitochondrial respiration in the presence of malate (or succinate) and ADP but without sulfide could be completely inhibited by rotenone, antimycin, cyanide, and sulfide. Only 40% inhibition was achieved by salicylhydroxamic acid. Sulfide oxidation (with sulfide as the only substrate) was fully inhibited by antimycin and by salicylhydroxamic acid but not by rotenone or sulfide. Moreover, sulfide oxidation was 3–4‐fold less sensitive to cyanide as compared to normal respiration. The data indicate that sulfide oxidation in A. marina is linked to the respiratory electron transport chain. We suggest that electrons from sulfide enter the respiratory chain via ubiquinone or at the ubiquinol–cytochrome‐c oxidoreductase. At sulfide concentrations higher than 10 μM, the cyto‐chrome‐c oxidase is blocked and electrons from sulfide are transferred to oxygen via an alternative terminal oxidase.
Activation of Gene Transcription by Prostacyclin Analogues is Mediated by the Peroxisome‐Proliferators‐Activated Receptor (PPAR)
Authors: Hertz Rachel , Berman Ina , Keppler Dietrich , Bar‐Tana Jacob
Published: August 2004

Xenobiotic amphipathic carboxylates, known collectively as hypolipidemic peroxisome proliferators (e.g., aryloxyalkanoic acids), or native long‐chain fatty acids induce liver peroxisome proliferation and other biological activities. This broad spectrum of effects results from modulation of transcription of specific genes mediated by binding of peroxisome‐proliferators‐activated receptors (PPAR) to respective sequence‐specific promoter elements (PPRE). The broad specificity and relatively low potency of reported hypolipidemic peroxisome proliferators prompted us to search for specific highly potent peroxisome proliferators. Here we report that stable prostacyclin analogues may act in such a manner.

mPPARα‐mediated expression of a reporter gene linked to the peroxisomal rat acyl‐CoA oxidase promoter was dose‐dependently induced by carbaprostacyclin and iloprost. The ED50 for carbaprostacyclin was 25 nM, and carbaprostacyclin was therefore 25‐fold and 200‐fold more effective than the most potent xenobiotic (5,18,11,14‐eicosatetraynoic acid) and native (arachidonic acid) inducers, respectively. Induction was further increased by cotransfecting the cells with mPPARα and an expression vector for retinoic acid‐X‐receptor.

PPAR‐mediated activation of gene expression by prostacyclin analogues was specific for PPAR and was not observed using other members of the superfamily.

No activation of gene expression was induced by other prostaglandins or leukotrienes at concentrations 100‐fold higher than those of the prostacyclin analogues.

Induction of gene expression by prostacyclin analogues was inhibited in cells transfected with the long‐chain‐acyl‐CoA synthase, indicating that the acidic form of prostacyclin, rather than the respective CoA derivative or a metabolite derived thereof, serves as the activator of the PPAR/PPRE transduction pathway.

Hence, PPAR‐mediated modulation of gene transcription by prostacyclins may form the basis for their novel role as regulators of gene expression. Xenobiotic hypolipidemic peroxisome proliferators and native long‐chain fatty acids seem to exploit the PPAR/PPRE transduction pathway used by prostacyclin.
Pressure‐Tuning the Conformation of Bovine Pancreatic Trypsin Inhibitor Studied by Fourier‐Transform Infrared Spectroscopy
Authors: Goossens Koen , Smeller László , Frank Johannes , Heremans Karel
Published: August 2004

A hydrostatic pressure of 1.5 GPa induces changes in the secondary structure of bovine pancreatic tryspin inhibitor (BPTI) as revealed by the analysis of the amide I' band with Fourier‐transform infrared (FTIR) spectroscopy in the diamond anvil cell. The features of the secondary structure remain distinct at high pressure suggesting that the protein does not unfold. The fitted percentages of the secondary structure elements during compression and decompression strongly suggest that the pressure‐induced changes are reversible. The pressure‐induced changes in the tyrosine side chain band are also reversible. The results demonstrate that the infrared technique explores different aspects of the behaviour of proteins in comparison with two published molecular dynamics studies performed up to 1 GPa [Kitchen, D. B., Reed, L. H. & Levy, R. M. (1992) Biochemistry 31, 10083–10093] and 500 MPa [Brunne, R. M. & van Gunsteren, W. F. (1993) FEBS Lett. 323, 215–217]. A possible explanation for the difference is the time scale of the experiments.