bluuebunny/crossref_metadata_2025 huggingface dataset:
Accumulation of Soybean Glycinin and Its Assembly with the Glutelins in Rice1
Authors: Katsube Tomoyuki , Kurisaka Nobuyuki , Ogawa Masahiro , Maruyama Nobuyuki , Ohtsuka Reiko , Utsumi Shigeru , Takaiwa Fumio
Published: July 2002

Saline-soluble glycinins and insoluble glutelins are the major storage proteins in soybean (Glycine max) and rice (Oryza sativa), respectively. In spite of their differences in solubility properties, both proteins are members of the 11S globulin gene family based on their similarities in primary sequences and processing of the coded protein. Wild-type and methionine-modified glycinin coding sequences were expressed in transgenic rice plants under the control of the rice glutelin GluB-1 promoter. Glycinins were specifically synthesized in the endosperm tissue and co-localized with glutelins in type II protein bodies. They assembled into 7S and 11S species, similar to what was observed in developing soybean seeds. This pattern was quite different from that displayed by the rice glutelins in untransformed plants, in which processed subunits sedimenting at 2S were apparent. In glycinin-expressing transgenic plants, however, glutelins were observed sedimenting at 7S and 11S with lesser amounts in the 2S region. A portion of the glycinins was also found associated in the insoluble glutelin fraction. Renaturation experiments suggested that the hybrid glycinin-glutelin oligomers were formed through specific interactions. Overall, these results indicate that despite significant differences in the assembly of soybean glycinin and rice glutelin, both proteins can assemble with each other to form soluble hexameric oligomers or insoluble aggregates.
Biochemical and Immunocytochemical Characterization of Two Types of Myosins in Cultured Tobacco Bright Yellow-2 Cells
Authors: Yokota Etsuo , Yukawa Chiharu , Muto Shoshi , Sonobe Seiji , Shimmen Teruo
Published: July 2002

We have isolated a myosin (referred to as 170-kD myosin) from lily pollen tubes, which consists of 170-kD heavy chain and calmodulin (CaM) light chain and is responsible for cytoplasmic streaming. A 170-kD polypeptide that has similar antigenicity to the 170-kD myosin heavy chain of lily pollen tubes was also present in cultured tobacco (Nicotiana tabacum) Bright Yellow-2 (BY-2) cells, and possessed the ability to interact with F-actin in an ATP-dependent manner. In addition to this myosin, we identified biochemically another kind of myosin in BY-2 cells. This myosin consisted of a CaM light chain and a 175-kD heavy chain with antigenicity different from the 170-kD myosin heavy chain. In the present study, we referred to this myosin as 175-kD myosin. This myosin was able to translocate rhodamine-phalloidin (RP)-labeled F-actin at an average velocity of about 9 μm/s in the motility assay in vitro. In contrast, the sliding velocity of RP-labeled F-actin translocated by fractions containing the 170-kD myosin was 3 to 4 μm/s. The velocity of cytoplasmic streaming in living BY-2 cells ranged from 2 to 9 μm/s. The motile activity of 175-kD myosin in vitro was inhibited by Ca2+ at concentrations higher than 10−6  m. Immunoblot analyses using an antiserum against the heavy chain of 170- or 175-kD myosin revealed that in tobacco plants, the 175-kD myosin was expressed in leaf, stem, and root, but not in germinating pollen, while 170-kD myosin was present in all of these plant parts and in germinating pollen. These results suggest that the two types of myosins, 170 and 175 kD, presumably participate in cytoplasmic streaming in BY-2 cells and other somatic cells of tobacco plants.
GDP-Fucose Uptake into the Golgi Apparatus during Xyloglucan Biosynthesis Requires the Activity of a Transporter-Like Protein Other Than the UDP-Glucose Transporter
Authors: Wulff Cristian , Norambuena Lorena , Orellana Ariel
Published: July 2002

The molecular mechanisms regulating hemicelluloses and pectin biosynthesis are poorly understood. An important question in this regard is how glycosyltransferases are oriented in the Golgi cisternae, and how nucleotide sugars are made available for the synthesis of the polymers. Here we show that the branching enzyme xyloglucan α,1–2 fucosyltransferase (XG-FucTase) from growing pea (Pisum sativum) epicotyls was latent and protected against proteolytic inactivation on intact, right-side-in pea stem Golgi vesicles. Moreover, much of the XG-FucTase activity was membrane associated. These data indicate that XG-FucTase is a membrane-bound luminal enzyme. GDP-Fuc uptake studies demonstrated that GDP-Fuc was taken up into Golgi vesicles in a protein-mediated process, and that this uptake was not competed by UDP-Glc, suggesting that a specific GDP-Fuc transporter is involved in xyloglucan biosynthesis. Once in the lumen, Fuc was transferred onto endogenous acceptors, including xyloglucan. GDPase activity was detected in the lumen of the vesicles, suggesting than the GDP produced upon transfer of Fuc was hydrolyzed to GMP and inorganic phosphate. We suggest than the GDP-Fuc transporter and GDPase may be regulators of xyloglucan fucosylation in the Golgi apparatus from pea epicotyls.
Dg93, a Nodule-Abundant mRNA of Datisca glomerata with Homology to a Soybean Early Nodulin Gene
Authors: Okubara Patricia A. , Fujishige Nancy A. , Hirsch Ann M. , Berry Alison M.
Published: July 2002

We have isolated a 590-bp full-length cDNA clone designatedDg93, an mRNA that is highly expressed in symbiotic root nodules of the actinorhizal host Datisca glomerata. Dg93mRNA encodes a deduced polypeptide of 105 amino acids with significant identity (74%) to the soybean (Glycine max) early nodulin (ENOD) gene GmENOD93 (Kouchi and Hata, 1993).Dg93 mRNA is abundant in nodules at 4 weeks post inoculation, the earliest time assayed, and steady-state mRNA levels remain elevated 11 weeks after inoculation. Spatial patterns ofDg93 mRNA expression are complex, with transcript accumulation in the nodule lobe meristem, early infection zone, periderm, and cells of the vascular cylinder, but not in the surrounding uninfected cortical cells. Dg93 is encoded by a small gene family in D. glomerata. To our knowledge, this is the first report of a gene from an actinorhizal host that is expressed in the nodule meristem and that shares sequence homology with an early nodulin gene from a legume.
Purified γ-Glutamyl Transpeptidases from Tomato Exhibit High Affinity for Glutathione and GlutathioneS-Conjugates
Authors: Martin Melinda Neal , Slovin Janet P.
Published: July 2002

γ-Glutamyl transpeptidases (γGTases) are the only enzymes known to hydrolyze the unique N-terminal amide bonds of reduced glutathione (γ-l-glutamyl-cysteinyl-glycine), oxidized glutathione, and glutathione S-conjugates. Two γGTases (I and II) with K  m values for glutathione of 110 and 90 μm were purified 2,977-fold and 2,152-fold, respectively, from ripe tomato (Lycopersicon esculentum) pericarp. Both enzymes also hydrolyze dipeptides and other tripeptides with N-terminal, γ-linked Glu and the artificial substrates γ-l-glutamyl-p-nitroanilide and γ-l-glutamyl(7-amido-4-methylcoumarin). They transfer the glutamyl moiety to water or acceptor amino acids, includingl-Met, l-Phe, l-Trp,l-Ala, or the ethylene precursor 1-aminocyclopropane-1-carboxylic acid. γGTase I and II were released from a wall and membrane fraction of a tomato fruit extract with 1.0 m NaCl, suggesting that they are peripheral membrane proteins. They were further purified by acetone precipitation, Dye Matrex Green A affinity chromatography, and hydrophobic interaction chromatography. The two γGTases were resolved by concanavalin A (Con A) affinity chromatography, indicating that they are differentially glycosylated. The native and SDS-denatured forms of both enzymes showed molecular masses of 43 kD.
Cloning, Developmental, and Tissue-Specific Expression of Sucrose:Sucrose 1-Fructosyl Transferase from Taraxacum officinale. Fructan Localization in Roots
Authors: Van den Ende Wim , Michiels An , Van Wonterghem Dominik , Vergauwen Rudy , Van Laere André
Published: July 2002

Sucrose:sucrose 1-fructosyl transferase (1-SST) is the key enzyme initiating fructan synthesis in Asteraceae. Using reverse transcriptase-PCR, we isolated the cDNA for 1-SST from Taraxacum officinale. The cDNA-derived amino acid sequence showed very high homology to other Asteracean 1-SSTs (Cichorium intybus 86%, Cynara scolymus 82%,Helianthus tuberosus 80%), but homology to 1-SST fromAllium cepa (46%) and Aspergillus foetidus (18%) was much lower. Fructan concentrations, 1-SST activities, 1-SST protein, and mRNA concentrations were compared in different organs during vegetative and generative development ofT. officinale plants. Expression of 1-SST was abundant in young roots but very low in leaves. 1-SST was also expressed at the flowering stages in roots, stalks, and receptacles. A good correlation was found between northern and western blots showing transcriptional regulation of 1-SST. At the pre-flowering stage, 1-SST mRNA concentrations and 1-SST activities were higher in the root phloem than in the xylem, resulting in the higher fructan concentrations in the phloem. Fructan localization studies indicated that fructan is preferentially stored in phloem parenchyma cells in the vicinity of the secondary sieve tube elements. However, inulin-like crystals occasionally appeared in xylem vessels.
Potassium-Efflux Channels in Extensor and Flexor Cells of the Motor Organ of Samanea saman Are Not Identical. Effects of Cytosolic Calcium
Authors: Moshelion Menachem , Moran Nava
Published: July 2002

Leaflet movements in the mimosa-family tree Samanea saman stem from coordinated volume changes of cells in the leaf motor organs in the adaxial and abaxial motor cells (“flexors” and “extensors”). Shrinking, initiated by dissimilar light signals in extensors and in flexors, depends in both cell types on K+efflux via depolarization-dependent potassium (KD) channels. To compare between flexor and extensor KDchannels and to test for a possible interaction of these channels with the Ca2+-mobilizing phosphoinositide cascade evoked in these motor cells by the “shrinking signals,” we probed the channels with varying (5 nm–3 mm) cytosolic free-Ca2+ concentration ([Ca2+]cyt) in patch-clamped inside-out excised membrane patches. Ca2+ was not required for KD channel activation. [Ca2+]cytof 600 nm decreased the mean number of open KD channels in flexors, as monitored at −30 mV. Detailed analysis revealed that in flexors millimolar [Ca2+]cyt decreased the maximum number of open channels, but simultaneously increased KD channel opening probability by negatively shifting the half-maximum-activation voltage by 40 to 50 mV. Thus, the promoting and the inhibitory effects at millimolar [Ca2+]cyt practically cancelled-out. In contrast to flexors, none of the gating parameters of the extensor KD channels were affected by [Ca2+]cyt. Irrespective of [Ca2+]cyt, the steady-state gating of extensor KD channels was slightly but significantly more voltage sensitive than that of flexors. The unitary conductances of flexor and extensor KD channels were similar and decreased by approximately 20% at millimolar [Ca2+]cyt. It is intriguing that the extensor KD channels were significantly less K+selective than those in flexors.
Involvement of a Nuclear-Encoded Basic Helix-Loop-Helix Protein in Transcription of the Light-Responsive Promoter ofpsbD
Authors: Baba Kyoko , Nakano Takeshi , Yamagishi Kazutoshi , Yoshida Shigeo
Published: July 2002

In the chloroplast psbD light-responsive promoter (LRP), a highly conserved sequence exists upstream from the bacterial −10/−35 elements. Multiple sequence-specific DNA binding proteins are predicted to bind to the conserved sequence as transcription factors. Using yeast one-hybrid screening of an Arabidopsis cDNA library, a possible DNA binding protein of the psbD LRP upstream sequence was identified. The protein, designated PTF1, is a novel protein of 355 amino acids (estimated molecular weight of 39.6) that contains a basic helix-loop-helix DNA binding motif in the predicted N-terminal region of the mature protein. Transient expression assay of PTF1-GFP fusion protein showed that PTF1 was localized in chloroplasts. Using the modified DNA sequence in the one-hybrid system, the ACC repeat was shown to be essential for PTF1 binding. The rate of psbD LRP mRNA accumulation was reduced in a T-DNA-inserted Arabidopsis ptf1 mutant. Compared with wild-type plants, the mutant had pale green cotyledons and its growth was inhibited under short-day conditions. These results suggest that PTF1 is a trans-acting factor of the psbD LRP.
The Internal Propeptide of the Ricin Precursor Carries a Sequence-Specific Determinant for Vacuolar Sorting
Authors: Frigerio Lorenzo , Jolliffe Nicholas A. , Di Cola Alessandra , Felipe Doramys Hernández , Paris Nadine , Neuhaus Jean-Marc , Lord J. Michael , Ceriotti Aldo , Roberts Lynne M.
Published: July 2002

Ricin is a heterodimeric toxin that accumulates in the storage vacuoles of castor bean (Ricinus communis) endosperm. Proricin is synthesized as a single polypeptide precursor comprising the catalytic A chain and the Gal-binding B chain joined by a 12-amino acid linker propeptide. Upon arrival in the vacuole, the linker is removed. Here, we replicate these events in transfected tobacco (Nicotiana tabacum) leaf protoplasts. We show that the internal linker propeptide is responsible for vacuolar sorting and is sufficient to redirect the ricin heterodimer to the vacuole when fused to the A or the B chain. This internal peptide can also target two different secretory protein reporters to the vacuole. Moreover, mutation of the isoleucine residue within an NPIR-like motif of the propeptide affects vacuolar sorting in proricin and in the reconstituted A-B heterodimer. This is the first reported example of a sequence-specific vacuolar sorting signal located within an internal propeptide.
Environmental training: emerging products
Authors: Beard Colin
Published: July 2002

Describes how environmental training is increasingly being recognized as a crucial element of any corporate, environmental strategy. In turn, corporate strategies will be an essential component to the achievement of sustainable development. But how do companies respond to the need to change the everyday practices of staff at all levels? Introduces a three‐layered approach to environmental training. Also examines the evolution and progress of two very different new training products. First, Expert Training Systems plc joined forces with BTCV Enterprises to create a Developing People Programme using the outdoors and incorporating specified environmental projects. More outdoor development suppliers are now offering such packages and they are selling well. Second, Earthwise, produced in 1995 in collaboration with ICI, Sheffield Hallam University and Sanderson CBT, provides an interactive toolkit approach to exploring the environmental aspects of producing a fictitious plastic, “prolene”. The product requires the user to assume a decision‐making role through a matrix of six main issues and 21 sub‐issues. At each sub‐issue there are over 34 events, 200 pieces of advice on offer, over 200 generic principles and around 122 possible courses of action.
A customer orientation checklist: a model
Authors: Reyes Pacios Lozano Ana
Published: July 2002

A brief review of the concept connotations in the area of information services is carried out, trying to stress the implications of being “customer oriented” as the most important component of marketing. A model to measure the level of the library’s orientation toward its market – as perceived by its managers – is presented. It was designed taking into account the models that major specialists prepared in order to evaluate marketing activity in companies. It should allow librarians to establish to what extent the library is properly user oriented and, at the same time, what specific marketing‐related aspects it is failing in or should improve at.
The Northern Ireland retailing environment and its effect on ethnic food consumption
Authors: McIlveen Heather , Chestnutt Samantha
Published: July 2002

The recent arrival of the UK multiples has had a significant effect on the Northern Ireland market, particularly in terms of product range and choice. Ethnic foods offer a potentially lucrative sector, as evidenced in Great Britain. However, retailers in Northern Ireland are only beginning to develop range and ethnicity and are familiarising themselves with a market which has been characterised more often by traditional eating habits. The preliminary study considered consumer perceptions of ethnic food products in Northern Ireland. It would appear that product awareness needs to be developed carefully, and the onus is on the retailer (and indeed, the hospitality establishments) to inform consumers and respond to their needs and wants, particularly with the more recent trend towards regional cuisine and greater authenticity. Only then can a larger range of truly ethnic food products penetrate the Northern Ireland market successfully.
Finding better solutions faster ‐ the issue resolution process
Authors: Barnett Alex
Published: July 2002

Decision making is an inherently simple process, but soon becomes complicated when a number of people are involved in the decision‐making process. How can a group move from a situation of disagreement or conflict to compromise or consensus, yet have confidence that the ultimate decision reflects the best solution to a problem? Describes the “traditional” methodology for managing the resolution of issues within groups and goes on to point out the limitations of this approach and ways in which it can be improved. The resulting methodology is a time‐efficient issue resolution process, which ensures that all team members are given the opportunity to contribute their ideas and opinions of the selected solution. This results in full commitment.
The nascent European policy towards older workers: can the European Union help the older worker?
Authors: McDonald Frank , Potton Margaret
Published: July 2002

Considers the reasons for the lack of consensus among the member states of the European Union (EU) on both the scope and extent of an ageism policy. The EU has become increasingly concerned with the problems that are raised by the ageing of the European population. In response to these concerns the EU has began to develop a policy towards older employees. The expectation of the EU is that this policy will help the member states effectively to adjust to the demographic challenge of an ageing population. Assessment of the effectiveness of the approach taken by the EU is made by utilizing economic theories that have been devised by neo‐classical and equal opportunities economists. The prospects for the development of a common EU policy on ageism is discussed and its possible impact on companies operating in the EU is explored.
Seeing distances – a fast time‐of‐flight 3D camera
Authors: Lange Robert , Seitz Peter
Published: July 2002

The world’s first time‐of‐flight (TOF) 3D range camera without moving parts, based on a custom solid‐state imaging device is described in this article. With the single exception of the smart‐pixel array, only electronic and optical standard components are used. The range camera does not need any mechanically scanning parts. It acquires 10 range images per second and achieves a resolution of a few centimeters over a distance range of 10‐20 meters for non‐cooperative targets. The illumination source consists of 160 LEDs, modulated at 20MHz, with a total optical power of about 800mW. The description of the smart pixel’s working principle and the presentation of our first 3D measurements are completed with a discussion of the theoretical limitations and a comparison with the system performance.
Why does reengineering fail? A practical guide for successful implementation
Authors: Attaran Mohsen
Published: July 2002

Process redesign is not always successful and is almost always accompanied by pain or at least unpleasant side effects. Many companies have undertaken reengineering efforts only to abandon them with little or no positive results. As important as it is to understand how to reengineer, it is crucial to understand how to avoid the traps. A disciplined, step‐by‐step approach to implementing process redesign is critical to the effort. This article offers a framework for the successful design and construction of the reengineering solution.
Modelling total energy costs of sport centres
Authors: Boussabaine A.H. , Kirkham R.J. , Grew R.J.
Published: July 2002

Providing and maintaining safe and comfortable conditions in sport centres raises many issues, particularly cost. The paper gives an overview of the factors associated with sport centre servicing and attempts to highlight the governing factors associated with this, particularly energy costs. A total of 19 sport centres in the City of Liverpool in the UK are investigated, using data elicited from the Liverpool Leisure Services Directorate. The energy operating costs were analysed using statistical methods. Six models were developed to predict total energy costs. Testing and validation results showed a high level of model accuracy. The models would be of use to professionals involved in feasibility studies at the design stage.
Utilizing thermal fatigue testing to differentiate the performance of epoxy materials at various glass transition temperature levels
Authors: Reutzel Kathleen M.
Published: July 2002

A common method for understanding the thermal performance of epoxy laminate materials is to analyze the glass transition temperature using instruments such as differential scanning calorimetry (DSC), dynamic mechanical analysis (DMA) and thermomechanical analysis (TMA). In order to characterize the long‐term performance of a finished printed circuit board, more advanced reliable test methods have been developed. This paper will discuss interconnect stress testing (IST), an accelerated fatigue test used for evaluating the failure modes of a printed circuit board (PCB) interconnect. IST utilizes a DC current to heat the PCB to the recommended temperatures within the interconnect. The plated through hole integrity and the post‐interconnect integrity can be monitored simultaneously. The test matrix compares the performance of various AlliedSignal epoxy laminate materials as a function of glass transition temperature (Tg) and board design.
The Pygmalion effect reconsidered: its implications for education, training and workplace learning
Authors: Murphy Daragh , Campbell Clifton , Garavan Thomas N.
Published: July 2002

Reviews the literature on the Pygmalion effect and in particular its implications for workplace learning and training. The article considers the wide ranging debate within the education literature on the value of the Pygmalion concept and also considers research conducted in training and workplace settings. The implications of the concept for learning design, trainee self‐esteem, trainer behaviours, and workplace learning are considered.
Eutectic Sn/Pb solder bump and under bump metallurgy: interfacial reactions and adhesion
Authors: Jang Se‐Young , Paik Kyung‐Wook
Published: July 2002

In flip‐chip interconnection on organic substrates using eutectic tin/lead solder bumps, a highly reliable under bump metallurgy (UBM) is required to maintain adhesion and solder wettability. Various UBM systems such as 1μm Al/0.2μm Ti/5μm Cu, 1μm Al/02μm Ti/1μm Cu, 1μm Al/0.2μm Ni/1μm Cu and 1μm Al/0.2μm Pd/1μm Cu, applied under eutectic tin/lead solder bumps, have been investigated with regard to their interfacial reactions and adhesion properties. The effects of the number of solder reflow cycles and the aging time on the growth of intermetallic compounds (IMCs) and on the solder ball shear strength were investigated. Good ball shear strength was obtained with 1μm Al/0.2μm Ti5μm Cu and 1μm Al/0.2μm Ni/1μm Cu even after four solder reflows or seven‐day aging at 150∞C. In contrast, 1μm Al/0.2μm Ti/1μm Cu and 1μm Al/0.2μm Pd/1μm Cu showed poor ball shear strength. The decrease of the shear strength was mainly due to the direct contact between solder and non‐wettable metals such as Ti and AL, resulting in a delamination. In this case, thin 1μm Cu and 0.2μm Pd diffusion barrier layers were completely consumed by Cu‐Sn and Pd‐Sn reaction.
ICT‐enabled coordination of dynamic supply webs
Authors: Christiaanse Ellen , Kumar Kuldeep
Published: July 2002

Increasingly, two key trends – value‐ and cost‐conscious customers, and globalization of markets and supply sources – are shifting the competitive focus from the competitive advantage of firms to competitive advantages of entire supply chains. At the same time, the possibilities for flexibility and coordination inherent in modern information and communication technologies are making it possible to design and follow a much greater variety of organization and governance strategies for delivering customer value. Contributes primarily to the development of general principles for ICT‐enabled redesign of supply chains. Rather than examine the individual impact and design implications of each new ICT innovation, proposes principles of abstraction that can be used to frame supply‐chain redesign options and decisions. Concludes with how these principles lead to flexibility in the design of various supply chain options, thereby providing cost and value advantages for the supply chain.
Non‐convergence versus non‐conservation in effective heat capacity methods for phase change problems
Authors: Gong Zhen‐Xiang , Mujumdar Arun S.
Published: July 2002

In the original effective heat capacity method the latent heat effect is approximated by a large effective heat capacity over a small temperature range. This method is simple in concept and easy to implement. However, it is so sensitive to the choice of the phase change temperature interval and the chosen time integration scheme that non‐convergence always occurs when it is implemented with implicit time integration schemes. Presents the cause and cure of non‐convergence in effective heat capacity methods and discusses sample results.
A good league table guide?
Authors: Yorke Mantz
Published: July 2002

Subjects the league table of universities published in The Times and The Times Good University Guide in 1995 and 1996 to statistical and conceptual analyses. Shows the “measures” used by the compilers of the tables to be problematic in a number of technical respects. Shows that the number of underlying variables is largely reducible to a single factor which discriminates between pre‐1992 and post‐1992 universities. Casts serious doubt on the validity of the tables from both technical and conceptual perspectives.
Smart card technology for deploying a secure information management framework
Authors: Lambrinoudakis Costas
Published: July 2002

The continuously increasing need for de‐centralized information systems offering data to the people who need them irrespective of their physical location, as well as the requirement for exchanging information between different but interoperable systems, make the system’s architectural and functional design more complex and in many cases extremely vulnerable in respect to its security attributes. The concept of a “secure portable information file”, that can nowadays be easily implemented through the available smart card technology, can significantly ease information management and ensure maximum data protection in respect to their integrity, confidentiality and availability. This paper presents the use of smart cards in an educational environment as a case‐study example for demonstrating the above mentioned benefits, focussing on the utilization of the smart card’s cryptographic functions for implementing mechanisms capable of providing an extremely secure operational framework in terms of user and application provider authenticity, management of access privileges and data integrity and confidentiality.
Providing feedback to distant students
Authors: Ley Kathryn
Published: July 2002

Describes a feedback system for distant students including the tools and documents to provide feedback on frequent fixed response assignments and on projects or essay response assignments. A distance feedback system depends upon carefully planned, written assignments; specified evaluation criteria; and technology. It includes documentation, progress tracking, standardized responses based upon the specified evaluation criteria, and multiple assessments during the course. The proposed feedback system accommodates technological limitations yet provides useful feedback to facilitate distant student learning. The system could be adapted to benefit students enrolled in traditional instruction. A description of the feedback system components follows a brief introduction to instructional feedback and feedback levels and functions.
The effectiveness of commercial Internet Web sites: a user’s perspective
Authors: Bell Hudson , Tang Nelson K.H.
Published: July 2002

The main thrust of this article is to discuss a survey of 60 companies, predominantly from the user’s perspective, that use the Internet currently; and to examine the effectiveness of their current Internet Web sites. The results of an online survey were subjected to statistical analysis and this revealed some interesting findings. It was found that 30 percent of the companies had facilities for conducting transactions online and only 7 percent charged users for Web site access. Overall, the Web sites rated highly in terms of ease of access, content and structure but scored poorly for their number of unique features. Of the six industry sectors surveyed (electronic commerce, entertainment and leisure, financial and banking services, information services, retailing and travel and tourism), it was the retailing sector that showed the best overall performance. The best individual Web site in the survey was the Financial Times (http://www.ft.com). It is suggested that this initial survey work could be advanced by extending the sample size and by devising a programme of more extensive statistical analysis.
Rationality and implementation analysis
Authors: Ryan Neal
Published: July 2002

This paper examines the prospects of developing rational policy processes. The approach taken is to examine two components of policy processes. First, the paper analyses the way in which rationality has been applied to three different models, or modes of public administration: Weberian bureaucracy; market or rational actor political behaviour; and managerialism. The analysis suggests that “rational” approaches to public administration are inherently value‐laden, emphasising norms such as institutional integrity, representation or efficiency. Second, analysis is undertaken of policy implementation which is one phase of the policy process. The paper examines “top‐down”, “bottom‐up”, institutional and statutory‐coherence approaches to policy implementation. Contrasts amongst these competing models of policy implementation reinforce previous findings that there appears to be little prospect of achieving policy rationality because of the inability of the current approaches to policy analysis to enable reconciliation of fundamental normative assumptions underpinning the approaches. The current methods utilised by policy analysts do not appear to be able to provide either the tools or the structures required to achieve instrumental rationality in policy sciences.
Using consumer benchmarking criteria to improve service sector competitiveness
Authors: Smith Anne M.
Published: July 2002

The adoption of benchmarking techniques has increased over recent years, yet service organisations have been relatively reluctant to adopt the practice. It is widely acknowledged that elements of “service quality” play a key role in the performance and competitiveness of service organisations and thus provide potential benchmarking criteria. Yet perceived service quality must be defined from the consumer’s perspective and, unlike manufacturing organisations, the consumer is involved in the production process. This study examines the potential for the generation and evaluation of consumer focused benchmarking criteria. Consumers of three service sectors – health (family planning); education/professional (accountancy training) and retail (supermarkets) completed measurement scales relating to potentially deterministic attributes and assessed these for current and previous suppliers. The findings indicate that, although management must be aware of a number of potential problems and issues, information derived from consumers can provide a valuable input into a comprehensive external benchmarking programme involving both competitive and generic measures.
Correlated conformational fluctuations during enzymatic catalysis: Implications for catalytic rate enhancement
Authors: Alper Kutay O. , Singla Manish , Stone Jennifer L. , Bagdassarian Carey K.
Published: July 2002

Correlated enzymatic conformational fluctuations are shown to contribute to the rate of enhancement achieved during catalysis. Cytidine deaminase serves as a model system. Crystallographic temperature factor data for this enzyme complexed with substrate analog, transition‐state analog, and product are available, thereby establishing a measure of atomic scale conformational fluctuations along the (approximate) reaction coordinate. First, a neural network‐based algorithm is used to visualize the decreased conformational fluctuations at the transition state. Second, a dynamic diffusion equation along the reaction coordinate is solved and shows that the flux velocity through the associated enzymatic conformation space is greatest at the transition state. These results suggest (1) that there are both dynamic and energetic restrictions to conformational fluctuations at the transition state, (2) that enzymatic catalysis occurs on a fluctuating potential energy surface, and (3) a form for the potential energy. The Michaelis‐Menten equations are modified to describe catalysis on this fluctuating potential energy profile, leading to enhanced catalytic rates when fluctuations along the reaction coordinate are appropriately correlated. This represents a dynamic tuning of the enzyme for maximally effective transformation of the ES complex into EP.
Inactivation mechanism of the membrane protein diacylglycerol kinase in detergent solution
Authors: Zhou Yufeng , Lau Francis W. , Nauli Sehat , Yang Dawn , Bowie James U.
Published: July 2002

We have examined the irreversible inactivation mechanism of the membrane protein diacylglycerol kinase in the detergents n‐octyl‐β‐D‐glucopyranoside (OG) at 55°C and n‐decyl‐maltopyranoside (DM) at 80°C. Under no inactivation conditions did we find any direct evidence for the chemical modifications that are commonly found in soluble proteins. Moreover, protein inactivated at 55°C in OG could be reactivated by an unfolding and refolding protocol, suggesting that the protein is inactivated by a stable conformational change, not a covalent modification. We also found that the inactivation rate decreased with both increasing protein concentration and increasing thermodynamic stability, consistent with an inactivation pathway involving transient dissociation and/or unfolding of the protein. Our results suggest that the primary cause of diacylglycerol kinase inactivation is not low solubility, but poor intrinsic stability in the detergent environment.
Optimization of binding electrostatics: Charge complementarity in the barnase‐barstar protein complex
Authors: Lee Lee‐Peng , Tidor Bruce
Published: July 2002

Theoretical and experimental studies have shown that the large desolvation penalty required for polar and charged groups frequently precludes their involvement in electrostatic interactions that contribute strongly to net stability in the folding or binding of proteins in aqueous solution near room temperature. We have previously developed a theoretical framework for computing optimized electrostatic interactions and illustrated use of the algorithm with simplified geometries. Given a receptor and model assumptions, the method computes the ligand‐charge distribution that provides the most favorable balance of desolvation and interaction effects on binding. In this paper the method has been extended to treat complexes using actual molecular shapes. The barnase‐barstar protein complex was investigated with barnase treated as a target receptor. The atomic point charges of barstar were varied to optimize the electrostatic binding free energy. Barnase and natural barstar form a tight complex (Kd ∼ 10−14 M) with many charged and polar groups near the interface that make this a particularly relevant system for investigating the role of electrostatic effects on binding. The results show that sets of barstar charges (resulting from optimization with different constraints) can be found that give rise to relatively large predicted improvements in electrostatic binding free energy. Principles for enhancing the effect of electrostatic interactions in molecular binding in aqueous environments are discussed in light of the optima. Our findings suggest that, in general, the enhancements in electrostatic binding free energy resulting from modification of polar and charged groups can be substantial. Moreover, a recently proposed definition of electrostatic complementarity is shown to be a useful tool for examining binding interfaces. Finally, calculational results suggest that wild‐type barstar is closer to being affinity optimized than is barnase for their mutual binding, consistent with the known roles of these proteins.
Exploring protein aggregation and self‐propagation using lattice models: Phase diagram and kinetics
Authors: Dima R.I. , Thirumalai D.
Published: July 2002

Many seemingly unrelated neurodegenerative disorders, such as amyloid and prion diseases, are associated with propagating fibrils whose structures are dramatically different from the native states of the corresponding monomers. This observation, along with the experimental demonstration that any protein can aggregate to form either fibrils or amorphous structures (inclusion bodies) under appropriate external conditions, suggest that there must be general principles that govern aggregation mechanisms. To probe generic aspects of prion‐like behavior we use the model of Harrison, Chan, Prusiner, and Cohen. In this model, aggregation of a structure, that is conformationally distinct from the native state of the monomer, occurs by three parallel routes. Kinetic partitioning, which leads to parallel assembly pathways, occurs early in the aggregation process. In all pathways transient unfolding precedes polymerization and self‐propagation. Chain polymerization is consistent with templated assembly, with the dimer being the minimal nucleus. The kinetic effciency of Rn−1 + G → Rn (R is the aggregation prone state and G is either U, the unfolded state, or N, the native state of the monomer) is increased when polymerization occurs in the presence of a “seed” (a dimer). These results support the seeded nucleated‐polymerization model of fibril formation in amyloid peptides. To probe generic aspects of aggregation in two‐state proteins, we use lattice models with side chains. The phase diagram in the (T,C) plane (T is the temperature and C is the polypeptide concentration) reveals a bewildering array of “phases” or structures. Explicit computations for dimers show that there are at least six phases including ordered structures and amorphous aggregates. In the ordered region of the phase diagram there are three distinct structures. We find ordered dimers (OD) in which each monomer is in the folded state and the interaction between the monomers occurs via a well‐defined interface. In the domain‐swapped structures a certain fraction of intrachain contacts are replaced by interchain contacts. In the parallel dimers the interface is stabilized by favorable intermolecular hydrophobic interactions. The kinetics of folding to OD shows that aggregation proceeds directly from U in a dynamically cooperative manner without populating partially structured intermediates. These results support the experimental observation that ordered aggregation in the two‐state folders U1A and CI2 takes place from U. The contrasting aggregation processes in the two models suggest that there are several distinct mechanisms for polymerization that depend not only on the polypeptide sequence but also on external conditions (such as C, T, pH, and salt concentration).
Covalent cross‐linking of proteins without chemical reagents
Authors: Simons Brigitte L. , King Mary C. , Cyr Terry , Hefford Mary Alice , Kaplan Harvey
Published: July 2002

A facile method for the formation of zero‐length covalent cross‐links between protein molecules in the lyophilized state without the use of chemical reagents has been developed. The cross‐linking process is performed by simply sealing lyophilized protein under vacuum in a glass vessel and heating at 85°C for 24 h. Under these conditions, approximately one‐third of the total protein present becomes cross‐linked, and dimer is the major product. Chemical and mass spectroscopic evidence obtained shows that zero‐length cross‐links are formed as a result of the condensation of interacting ammonium and carboxylate groups to form amide bonds between adjacent molecules. For the protein examined in the most detail, RNase A, the cross‐linked dimer has only one amide cross‐link and retains the enzymatic activity of the monomer. The in vacuo cross‐linking procedure appears to be general in its applicability because five different proteins tested gave substantial cross‐linking, and co‐lyophilization of lysozyme and RNase A also gave a heterogeneous covalently cross‐linked dimer.
Amide hydrogen exchange shows that malate dehydrogenase is a folded monomer at pH 5
Authors: Chen Jiwen , Smith David L.
Published: July 2002

Although there is general agreement that native mitochondrial malate dehydrogenase (MDH) exists as a dimer at pH 7, its aggregation state at pH 5 is less certain. The present amide hydrogen exchange study was performed to determine whether MDH remains a dimer at pH 5. To detect pH‐induced changes in solvent accessibility, MDH was exposed to D2O at pH 5 or 7, then fragmented with pepsin into peptides that were analyzed by mass spectrometry. Even after adjustments for the effect of pH on the intrinsic rate of hydrogen exchange, large increases in deuterium levels were found at pH 5 only in peptic fragments derived from the subunit binding surface of MDH. In parallel experiments, elevated deuterium levels were also found in the same regions of MDH monomer trapped inside a mutant form of the chaperonin GroEL. This selective increase in hydrogen exchange rates, which was attributed to increased solvent accessibility of these regions, provides new evidence that MDH is a monomer at pH 5.
Isoform‐specific effects of sialic acid on voltage‐dependent Na+ channel gating: functional sialic acids are localized to the S5‐S6 loop of domain I
Authors: Bennett Eric S.
Published: July 2002

The isoform specific role of sialic acid in human voltage‐gated sodium channel gating was investigated through expression and chimeric analysis of two human isoforms, Nav1.4 (hSkM1), and Nav1.5 (hH1) in Chinese hamster ovary (CHO) cell lines. Immunoblot analyses indicate that both hSkM1 and hH1 are glycosylated and that hSkM1 is more glycosylated than hH1. Four sets of voltage‐dependent parameters, the voltage of half‐activation (Va), the voltage of half‐inactivation (Vi), the time constants for fast inactivation (τh), and the time constants for recovery from inactivation (τrec), were measured for hSkM1 and hH1 expressed in two CHO cell lines, Pro5 and Lec2, to determine the effect of changing sialylation on channel gating under conditions of full (Pro5) or reduced (Lec2) sialylation. For all parameters measured, hSkM1 gating showed a consistent 11–15 mV depolarizing shift under conditions of reduced sialylation, while hH1 showed no significant change in any gating parameter. Shifts in channel Va with changing external [Ca2+] indicated that sialylation of hSkM1, but not hH1, directly contributes to a negative surface potential. Functional analysis of two chimeras, hSkM1P1 and hH1P1, indicated that the responsible sialic acids are localized to the hSkM1 S5‐S6 loop of domain I. When hSkM1 IS5‐S6 was replaced by the analogous hH1 loop (hSkM1P1), changing sialylation had no significant effect on any voltage‐dependent parameter. Conversely, when hSkM1 IS5‐S6 was added to hH1 (hH1P1), all four parameters shifted by 6–7 mV in the depolarized direction under conditions of reduced sialylation. In summary, the gating of two human sodium channel isoforms show very different dependencies on sialic acid, with hSkM1 gating uniformly altered by sialic acid levels through an apparent electrostatic mechanism, while hH1 gating is unaffected by changing sialylation. Sialic acid‐dependent gating can be removed or created by replacing or inserting hSkM1 IS5‐S6, respectively, indicating that the functionally relevant sialic acid residues are localized to the first domain of the channel.
Muscle contraction under capillaries in hamster muscle induces arteriolar dilatation via KATP channels and nitric oxide
Authors: Cohen Kenneth D. , Sarelius Ingrid H.
Published: July 2002

We tested the hypothesis that adenosine and nitric oxide can be sensed by capillaries and are implicated in the remote arteriolar dilatation initiated by muscle contraction. We also explored a role for KATP channel activity in this response. Small bundles of muscle fibres underlying a group of capillaries in cremaster muscles of anaesthetized hamsters were electrically stimulated to contract for 2 min at each of 2, 4 and 8 Hz. Diameter changes were measured in the inflow arteriole to the group of capillaries after muscle contraction in the presence or absence of 10−6m xanthine amine congener (XAC) to block A1 and A2 adenosine receptors, 10−4 or 10−3mNω‐nitro‐l‐arginine (LNNA) to block nitric oxide production, or 10−5m glibenclamide to block KATP channel activity. Dilatations were unchanged with XAC (3.0 ± 0.5, 3.9 ± 0.7 and 6.1 ± 1.0 μm), but were significantly reduced with LNNA (to 1.8 ± 0.6, 3.5 ± 0.7 and 4.9 ± 0.7 μm) or glibenclamide (to 0.4 ± 0.3, 0.8 ± 0.7 and 1.9 ± 0.6 μm). Neither KATP channel activity nor nitric oxide was required for transmission or manifestation of the dilator response. Thus, muscle contraction can be sensed by capillaries and the signalling mechanism for the ensuing remote dilatation depends on KATP channel activity and on NO, but not adenosine. Local application of 10−4m adenosine, 10−4m sodium nitroprusside or 10−5m pinacidil directly to capillaries initiated remote arteriolar dilatations. Thus, capillaries can respond directly to known mediators of metabolic vasodilatation, but these signalling pathways are not invariably implicated in the response to muscle contraction.