bluuebunny/crossref_metadata_2025 huggingface dataset:
Tetracycline tolerance mediated by gene amplification in Bacillus subtilis
Authors: Wannarat Wannasiri , Motoyama Shiori , Masuda Kenta , Kawamura Fujio , Inaoka Takashi
Published: August 2014

Bacillus subtilis can acquire a higher tolerance to tetracycline by increasing the gene dosage of its resistance gene tetB. In this study, we estimated the multiplication effect of tetB on tetracycline tolerance. Cells harbouring multiple copies of tetB were found to comprise approximately 30 % of the total tetracycline-resistant cell population when selected on medium containing 10 µg tetracycline ml−1. Disruption of recA resulted in a significant decrease in the frequency of tetB amplification. Although four direct repeats exist around tetB, the majority of tetB amplicons were found to be flanked by non-homologous sequences, indicating that the initial duplication of tetB can occur largely through RecA-independent recombination. The correlation between the tetB copy number and the MIC values for tetracycline indicated that more than three copies of tetB were required for tolerance to 10 µg tetracycline ml−1. Thus, the RecA-dependent expansion step appears to be necessary for developing significant tetracycline tolerance mediated by tetB amplification.
Aggregatibacter actinomycetemcomitans leukotoxin (LtxA; Leukothera) induces cofilin dephosphorylation and actin depolymerization during killing of malignant monocytes
Authors: Kaur Manpreet , Kachlany Scott C.
Published: August 2014

Leukotoxin (LtxA; Leukothera), a protein toxin secreted by the oral bacteriumAggregatibacter actinomycetemcomitans, specifically kills white blood cells (WBCs). LtxA binds to the receptor known as lymphocyte function associated antigen-1 (LFA-1), a β2integrin expressed only on the surface of WBCs. LtxA is being studied as a virulence factor that helpsA. actinomycetemcomitansevade host defences and as a potential therapeutic agent for the treatment of WBC diseases. LtxA-mediated cell death in monocytes involves both caspases and lysosomes; however, the signalling proteins that regulate and mediate cell death remain largely unknown. We used a 2D-gel proteomics approach to analyse the global protein expression changes that occur in response to LtxA. This approach identified the protein cofilin, which underwent dephosphorylation upon LtxA treatment. Cofilin is a ubiquitous actin-binding protein known to regulate actin dynamics and is regulated by LIM kinase (LIMK)-mediated phosphorylation. LtxA-mediated cofilin dephosphorylation was dependent on LFA-1 and cofilin dephosphorylation did not occur when LFA-1 bound to its natural ligand, ICAM-1. Treatment of cells with an inhibitor of LIMK (LIMKi) also led to cofilin dephosphorylation and enhanced killing by LtxA. This enhanced sensitivity to LtxA coincided with an increase in lysosomal disruption, and an increase in LFA-1 surface expression and clustering. Both LIMKi and LtxA treatment also induced actin depolymerization, which could play a role in trafficking and surface distribution of LFA-1. We propose a model in which LtxA-mediated cofilin dephosphorylation leads to actin depolymerization, LFA-1 overexpression/clustering, and enhanced lysosomal-mediated cell death.
Gene Expression: edgeRun: an R package for sensitive, functionally relevant differential expression discovery using an unconditional exact test
Authors: Dimont Emmanuel , Shi Jiantao , Kirchner Rory , Hide Winston
Published: August 2014

Next-generation sequencing platforms for measuring digital expression such as RNA-Seq are displacing traditional microarray-based methods in biological experiments. The detection of differentially expressed genes between groups of biological conditions has led to the development of numerous bioinformatics tools, but so far few, exploit the expanded dynamic range afforded by the new technologies. We present edgeRun, an R package that implements an unconditional exact test that is a more powerful version of the exact test in edgeR. This increase in power is especially pronounced for experiments with as few as 2 replicates per condition, for genes with low total expression and with large biological coefficient of variation. In comparison with a panel of other tools, edgeRun consistently captures functionally similar differentially expressed genes.

Availability

The package is freely available under the MIT license from CRAN (http://cran.r-project.org/web/packages/edgeRun)

Contact

edimont@mail.harvard.edu
Nutritional aspects of honey bee-collected pollen and constraints on colony development in the eastern Mediterranean
Authors: Avni Dorit , Hendriksma Harmen P. , Dag Arnon , Uni Zehava , Shafir Sharoni
Published: August 2014

*Graphical Abstract (for review)

*Highlights (for review)

Honey bee colonies in Israel collect a mean 7 kg protein and 0.7 kg fat per year

At maximal colony development, honey bee queens lay up to 3,300 eggs per day

Amount and content of collected pollen differ between sites and seasons

Pollen linoleic acid and protein levels best describe bee production cost

Colonies seem limited first by climate, then protein and finally specific nutrients

Pollen is the main protein and lipid source for honey bees (Apis mellifera), and nutritionally impoverished landscapes pose a threat to colony development. To determine colony nutritional demands, we analyzed a yearly cycle of bee-collected pollen from colonies in the field and compared it to colony worker production and honey bee body composition, for the first time in social insects. We monitored monthly bee production in ten colonies at each of seven sites throughout Israel, and trapped pollen bi-monthly in five additional colonies at each of four of these sites. Pollen mixtures from each sampling date and site were analyzed for weight, total protein, total fatty acids (FAs), and FA composition. Compared to more temperate climates, the eastern Mediterranean allows a relatively high yearly colony growth of ca. 300,000 to 400,000 bees. Colonies at higher elevation above sea level showed lower growth rates. Queen egg-laying rate did not seem to limit growth, as peaks in capped brood areas showed that queens lay a prolific 2,000 eggs a day on average, with up to 3,300 eggs in individual cases. Pollen uptake varied significantly among sites and seasons, with an overall annual mean total 16.8 kg per colony, containing 7.14 kg protein and 677 g fat. Overall mean pollen protein content was high (39.8%), and mean total FA content was 3.8%. Production cost, as expressed by the amount of nutrient used per bee, was least variable for linoleic acid and protein, suggesting these as the best descriptive variables for total number of bees produced. Linolenic acid levels in pollen during the autumn were relatively low, and supplementing colonies with this essential FA may mitigate potential nutritional deficiency. The essentiality of linoleic and linolenic acids was consistent with these FAs’ tendency to be present at higher levels in collected pollen than in the expected nutrients in bee bodies, demonstrating a well-developed adjustment between pollinator nutritional demands and the nutritional value of food offered by pollinated plants.
Genome-wide DNA methylation patterns in LSH mutant reveals de-repression of repeat elements and redundant epigenetic silencing pathways
Authors: Yu Weishi , McIntosh Carl , Lister Ryan , Zhu Iris , Han Yixing , Ren Jianke , Landsman David , Lee Eunice , Briones Victorino , Terashima Minoru , Leighty Robert , Ecker Joseph R. , Muegge Kathrin
Published: August 2014

Cytosine methylation is critical in mammalian development and plays a role in diverse biologic processes such as genomic imprinting, X chromosome inactivation, and silencing of repeat elements. Several factors regulate DNA methylation in early embryogenesis, but their precise role in the establishment of DNA methylation at a given site remains unclear. We have generated a comprehensive methylation map in fibroblasts derived from the murine DNA methylation mutant Hells−/− (helicase, lymphoid specific, also known as LSH). It has been previously shown that HELLS can influence de novo methylation of retroviral sequences and endogenous genes. Here, we describe that HELLS controls cytosine methylation in a nuclear compartment that is in part defined by lamin B1 attachment regions. Despite widespread loss of cytosine methylation at regulatory sequences, including promoter regions of protein-coding genes and noncoding RNA genes, overall relative transcript abundance levels in the absence of HELLS are similar to those in wild-type cells. A subset of promoter regions shows increases of the histone modification H3K27me3, suggesting redundancy of epigenetic silencing mechanisms. Furthermore, HELLS modulates CG methylation at all classes of repeat elements and is critical for repression of a subset of repeat elements. Overall, we provide a detailed analysis of gene expression changes in relation to DNA methylation alterations, which contributes to our understanding of the biological role of cytosine methylation.
Structure of human endonuclease V as an inosine-specific ribonuclease
Authors: Zhang Zhemin , Hao Zhitai , Wang Zhong , Li Qing , Xie Wei
Published: August 2014

The 6-aminopurine ring of adenosine (A) can be deaminated to form the 6-oxopurine of inosine (I). Endonuclease Vs (EndoVs) are inosine-specific nucleases that cleave at the second phosphodiester bond 3′ to inosine. EndoV proteins are highly conserved in all domains of life, but the bacterial and human enzymes seem to display distinct substrate preferences. While the bacterial enzymes exhibit high cleavage efficiency on various nucleic acid substrates, human EndoV (hEndoV) is most active towards ssRNA but is much less active towards other substrates. However, the structural basis of substrate recognition by hEndoV is not well understood. In this study, the 2.3 Å resolution crystal structure of hEndoV was determined and its unusual RNA-cleaving properties were investigated. The enzyme preserves the general `RNase H-like' structure, especially in the wedge motif, the metal-binding site and the hypoxanthine-binding pocket. hEndoV also features several extra insertions and a characteristic four-cysteine motif, in which Cys227 and Cys228, two cysteines that are highly conserved in higher eukaryotes, play important roles in catalysis. The structure presented here helps in understanding the substrate preference of hEndoV catalysis.
Mutational and structural study of RipA, a key enzyme inMycobacterium tuberculosiscell division: evidence for theL-to-Dinversion of configuration of the catalytic cysteine
Authors: Squeglia Flavia , Ruggiero Alessia , Romano Maria , Vitagliano Luigi , Berisio Rita
Published: August 2014

RipA is a key cysteine protease ofMycobacterium tuberculosisas it is responsible for bacterial daughter-cell separation. Although it is an important target for antimicrobial development, its mechanism of action and its interaction pattern with its substrate are hitherto unknown. By combining crystallographic and mutational studies with functional assays and molecular modelling, it is shown that the catalytic activity of the enzyme relies on a Cys–His–Glu triad and the impact of the mutation of each residue of the triad on the structure and function of RipA is analysed. Unexpectedly, the crystallographic analyses reveal that mutation of the glutamic acid to alanine results in inversion of the configuration of the catalytic cysteine. The consequent burial of the catalytic cysteine side chain explains the enzyme inactivation upon mutation. These data point to a novel role of the acidic residue often present in the triad of cysteine proteases as a supervisor of cysteine configuration through preservation of the local structural integrity.
xMDFF: molecular dynamics flexible fitting of low-resolution X-ray structures
Authors: McGreevy Ryan , Singharoy Abhishek , Li Qufei , Zhang Jingfen , Xu Dong , Perozo Eduardo , Schulten Klaus
Published: August 2014

X-ray crystallography remains the most dominant method for solving atomic structures. However, for relatively large systems, the availability of only medium-to-low-resolution diffraction data often limits the determination of all-atom details. A new molecular dynamics flexible fitting (MDFF)-based approach, xMDFF, for determining structures from such low-resolution crystallographic data is reported. xMDFF employs a real-space refinement scheme that flexibly fits atomic models into an iteratively updating electron-density map. It addresses significant large-scale deformations of the initial model to fit the low-resolution density, as tested with synthetic low-resolution maps of D-ribose-binding protein. xMDFF has been successfully applied to re-refine six low-resolution protein structures of varying sizes that had already been submitted to the Protein Data Bank. Finally,viasystematic refinement of a series of data from 3.6 to 7 Å resolution, xMDFF refinements together with electrophysiology experiments were used to validate the first all-atom structure of the voltage-sensing protein Ci-VSP.
Direct phase selection of initial phases from single-wavelength anomalous dispersion (SAD) for the improvement of electron density andab initiostructure determination
Authors: Chen Chung-De , Huang Yen-Chieh , Chiang Hsin-Lin , Hsieh Yin-Cheng , Guan Hong-Hsiang , Chuankhayan Phimonphan , Chen Chun-Jung
Published: August 2014

Optimization of the initial phasing has been a decisive factor in the success of the subsequent electron-density modification, model building and structure determination of biological macromolecules using the single-wavelength anomalous dispersion (SAD) method. Two possible phase solutions (ϕ1and ϕ2) generated from two symmetric phase triangles in the Harker construction for the SAD method cause the well known phase ambiguity. A novel direct phase-selection method utilizing the θDSlist as a criterion to select optimized phases ϕamfrom ϕ1or ϕ2of a subset of reflections with a high percentage of correct phases to replace the corresponding initial SAD phases ϕSADhas been developed. Based on this work, reflections with an angle θDSin the range 35–145° are selected for an optimized improvement, where θDSis the angle between the initial phase ϕSADand a preliminary density-modification (DM) phase ϕDMNHL. The results show that utilizing the additional direct phase-selection step prior to simple solvent flattening without phase combination using existing DM programs, such asRESOLVEorDMfromCCP4, significantly improves the final phases in terms of increased correlation coefficients of electron-density maps and diminished mean phase errors. With the improved phases and density maps from the direct phase-selection method, the completeness of residues of protein molecules built with main chains and side chains is enhanced for efficient structure determination.
Structural insight into arginine methylation by the mouse protein arginine methyltransferase 7: a zinc finger freezes the mimic of the dimeric state into a single active site
Authors: Cura Vincent , Troffer-Charlier Nathalie , Wurtz Jean-Marie , Bonnefond Luc , Cavarelli Jean
Published: August 2014

Protein arginine methyltransferase 7 (PRMT7) is a type III arginine methyltransferase which has been implicated in several biological processes such as transcriptional regulation, DNA damage repair, RNA splicing, cell differentiation and metastasis. PRMT7 is a unique but less characterized member of the family of PRMTs. The crystal structure of full-length PRMT7 fromMus musculusrefined at 1.7 Å resolution is described. The PRMT7 structure is composed of two catalytic modules in tandem forming a pseudo-dimer and contains only one AdoHcy molecule bound to the N-terminal module. The high-resolution crystal structure presented here revealed several structural features showing that the second active site is frozen in an inactive state by a conserved zinc finger located at the junction between the two PRMT modules and by the collapse of two degenerated AdoMet-binding loops.
An evaluation of adhesive sample holders for advanced crystallographic experiments
Authors: Mazzorana Marco , Sanchez-Weatherby Juan , Sandy James , Lobley Carina M. C. , Sorensen Thomas
Published: August 2014

The hydration state of macromolecular crystals often affects their overall order and, ultimately, the quality of the X-ray diffraction pattern that they produce. Post-crystallization techniques that alter the solvent content of a crystal may induce rearrangement within the three-dimensional array making up the crystal, possibly resulting in more ordered packing. The hydration state of a crystal can be manipulated by exposing it to a stream of air at controlled relative humidity in which the crystal can equilibrate. This approach provides a way of exploring crystal hydration space to assess the diffraction capabilities of existing crystals. A key requirement of these experiments is to expose the crystal directly to the dehydrating environment by having the minimum amount of residual mother liquor around it. This is usually achieved by placing the crystal on a flat porous support (Kapton mesh) and removing excess liquid by wicking. Here, an alternative approach is considered whereby crystals are harvested using adhesives that capture naked crystals directly from their crystallization drop, reducing the process to a one-step procedure. The impact of using adhesives to ease the harvesting of different types of crystals is presented together with their contribution to background scattering and their usefulness in dehydration experiments. It is concluded that adhesive supports represent a valuable tool for mounting macromolecular crystals to be used in humidity-controlled experiments and to improve signal-to-noise ratios in diffraction experiments, and how they can protect crystals from modifications in the sample environment is discussed.
Local dynamics of proteins and DNA evaluated from crystallographicBfactors
Authors: Schneider Bohdan , Gelly Jean-Christophe , de Brevern Alexandre G. , Černý Jiří
Published: August 2014

The dynamics of protein and nucleic acid structures is as important as their average static picture. The local molecular dynamics concealed in diffraction images is expressed as so-calledBfactors. To find out how the crystal-derivedBfactors represent the dynamic behaviour of atoms and residues of proteins and DNA in their complexes, the distributions of scaledBfactors from a carefully curated data set of over 700 protein–DNA crystal structures were analyzed [Schneideret al.(2014),Nucleic Acids Res.42, 3381–3394]. Amino acids and nucleotides were categorized based on their molecular neighbourhood as solvent-accessible, solvent-inaccessible (i.e.forming the protein core) or lying at protein–protein or protein–DNA interfaces; the backbone and side-chain atoms were analyzed separately. TheBfactors of two types of crystal-ordered water molecules were also analyzed. The analysis confirmed several expected features of protein and DNA dynamics, but also revealed surprising facts. Solvent-accessible amino acids haveBfactors that are larger than those of residues at the biomolecular interfaces, and core-forming amino acids are the most restricted in their movement. A unique feature of the latter group is that their side-chain and backbone atoms are restricted in their movement to the same extent; in all other amino-acid groups the side chains are more floppy than the backbone. The low values of theBfactors of water molecules bridging proteins with DNA and the very large fluctuations of DNA phosphates are surprising. The features discriminating different types of residues are less pronounced in structures with lower crystallographic resolution. Some of the observed trends are likely to be the consequence of improper refinement protocols that may need to be rectified.
Structure of the 2,4′-dihydroxyacetophenone dioxygenase fromAlcaligenessp. 4HAP
Authors: Keegan R. , Lebedev A. , Erskine P. , Guo J. , Wood S. P. , Hopper D. J. , Rigby S. E. J. , Cooper J. B.
Published: August 2014

The enzyme 2,4′-dihydroxyacetophenone dioxygenase (DAD) catalyses the conversion of 2,4′-dihydroxyacetophenone to 4-hydroxybenzoic acid and formic acid with the incorporation of molecular oxygen. Whilst the vast majority of dioxygenases cleave within the aromatic ring of the substrate, DAD is very unusual in that it is involved in C—C bond cleavage in a substituent of the aromatic ring. There is evidence that the enzyme is a homotetramer of 20.3 kDa subunits, each containing nonhaem iron, and its sequence suggests that it belongs to the cupin family of dioxygenases. In this paper, the first X-ray structure of a DAD enzyme from the Gram-negative bacteriumAlcaligenessp. 4HAP is reported, at a resolution of 2.2 Å. The structure establishes that the enzyme adopts a cupin fold, forming dimers with a pronounced hydrophobic interface between the monomers. The catalytic iron is coordinated by three histidine residues (76, 78 and 114) within a buried active-site cavity. The iron also appears to be tightly coordinated by an additional ligand which was putatively assigned as a carbonate dianion since this fits the electron density optimally, although it might also be the product formate. The modelled carbonate is located in a position which is highly likely to be occupied by the α-hydroxyketone group of the bound substrate during catalysis. Modelling of a substrate molecule in this position indicates that it will interact with many conserved amino acids in the predominantly hydrophobic active-site pocket where it undergoes peroxide radical-mediated heterolysis.
Crystallographic analysis and biochemical applications of a novel penicillin-binding protein/β-lactamase homologue from a metagenomic library
Authors: Ngo Tri Duc , Ryu Bum Han , Ju Hansol , Jang Eun Jin , Kim Kyeong Kyu , Kim T. Doohun
Published: August 2014

Interest in penicillin-binding proteins and β-lactamases (the PBP-βL family) is increasing owing to their biological and clinical significance. In this study, the crystal structure of Est-Y29, a metagenomic homologue of the PBP-βL family, was determined at 1.7 Å resolution. In addition, complex structures of Est-Y29 with 4-nitrophenyl phosphate (4NP) and with diethyl phosphonate (DEP) at 2.0 Å resolution were also elucidated. Structural analyses showed that Est-Y29 is composed of two domains: a β-lactamase fold and an insertion domain. A deep hydrophobic patch between these domains defines a wide active site, and a nucleophilic serine (Ser58) residue is located in a groove defined primarily by hydrophobic residues between the two domains. In addition, three hydrophobic motifs, which make up the substrate-binding site, allow this enzyme to hydrolyze a wide variety of hydrophobic compounds, including fish and olive oils. Furthermore, cross-linked Est-Y29 aggregates (CLEA-Est-Y29) significantly increase the stability of the enzyme as well as its potential for extensive reuse in various deactivating conditions. The structural features of Est-Y29, together with biochemical and biophysical studies, could provide a molecular basis for understanding the properties and regulatory mechanisms of the PBP-βL family and their potential for use in industrial biocatalysts.
Crystal structure of 2-ethyl-3-(4-fluorophenylsulfinyl)-5,7-dimethyl-1-benzofuran
Authors: Choi Hong Dae , Lee Uk
Published: August 2014

In the title compound, C18H17FO2S, the dihedral angle between the planes of the benzofuran ring system (r.m.s. deviation = 0.004 Å) and the 4-fluorophenyl ring is 86.38 (6)°. The terminal C atom of the ethyl substituent is displaced by 1.444 (3) Å from the benzofuran ring system to the same side of the molecule as the 4-fluorophenyl ring. In the crystal, molecules are linkedviapairs of C—H...π interactions into inversion-related dimers. These dimers are further linked by π–π interactions between the benzene rings of neighbouring molecules [centroid–centroid distance = 3.715 (3) Å] and between the furan rings of neighbouring molecules [centroid–centroid distance = 3.598 (3) Å]. The molecules are stacked along thea-axis direction. In the sulfinyl group, the S and O atoms are disordered over two sets of sites, with site-occupancy factors of 0.797 (3) and 0.213 (3).
Crystal structure of 4-(2,2-dimethylpropanamido)pyridin-3-ylN,N-diisopropyldithiocarbamate
Authors: El-Hiti Gamal A. , Smith Keith , Hegazy Amany S. , Baashen Mohammed , Kariuki Benson M.
Published: August 2014

In the title compound, C17H27N3OS2, the amide group is approximately coplanar with the pyridine ring [dihedral angle = 1.6 (1)°], whereas the dithiocarbamate group is nearly perpendicular to the pyridine ring [dihedral angle = 76.7 (1)°]. In the crystal, pairs of weak C—H...O hydrogen bonds link the molecules into inversion dimers.
Hirshfeld atom refinement
Authors: Capelli Silvia C. , Bürgi Hans-Beat , Dittrich Birger , Grabowsky Simon , Jayatilaka Dylan
Published: August 2014

Hirshfeld atom refinement (HAR) is a method which determines structural parameters from single-crystal X-ray diffraction data by using an aspherical atom partitioning of tailor-madeab initioquantum mechanical molecular electron densities without any further approximation. Here the original HAR method is extended by implementing an iterative procedure of successive cycles of electron density calculations, Hirshfeld atom scattering factor calculations and structural least-squares refinements, repeated until convergence. The importance of this iterative procedure is illustratedviathe example of crystalline ammonia. The new HAR method is then applied to X-ray diffraction data of the dipeptide Gly–L-Ala measured at 12, 50, 100, 150, 220 and 295 K, using Hartree–Fock and BLYP density functional theory electron densities and three different basis sets. All positions and anisotropic displacement parameters (ADPs) are freely refined without constraints or restraints – even those for hydrogen atoms. The results are systematically compared with those from neutron diffraction experiments at the temperatures 12, 50, 150 and 295 K. Although non-hydrogen-atom ADPs differ by up to three combined standard uncertainties (csu's), all other structural parameters agree within less than 2 csu's. Using our best calculations (BLYP/cc-pVTZ, recommended for organic molecules), the accuracy of determining bond lengths involving hydrogen atoms from HAR is better than 0.009 Å for temperatures of 150 K or below; for hydrogen-atom ADPs it is better than 0.006 Å2as judged from the mean absolute X-ray minus neutron differences. These results are among the best ever obtained. Remarkably, the precision of determining bond lengths and ADPs for the hydrogen atoms from the HAR procedure is comparable with that from the neutron measurements – an outcome which is obtained with a routinely achievable resolution of the X-ray data of 0.65 Å.
Electronic materials with a wide band gap: recent developments
Authors: Klimm Detlef
Published: August 2014

The development of semiconductor electronics is reviewed briefly, beginning with the development of germanium devices (band gapEg= 0.66 eV) after World War II. A tendency towards alternative materials with wider band gaps quickly became apparent, starting with silicon (Eg= 1.12 eV). This improved the signal-to-noise ratio for classical electronic applications. Both semiconductors have a tetrahedral coordination, and by isoelectronic alternative replacement of Ge or Si with carbon or various anions and cations, other semiconductors with widerEgwere obtained. These are transparent to visible light and belong to the group of wide band gap semiconductors. Nowadays, some nitrides, especially GaN and AlN, are the most important materials for optical emission in the ultraviolet and blue regions. Oxide crystals, such as ZnO and β-Ga2O3, offer similarly good electronic properties but still suffer from significant difficulties in obtaining stable and technologically adequatep-type conductivity.
The binding of platinum hexahalides (Cl, Br and I) to hen egg-white lysozyme and the chemical transformation of the PtI6octahedral complex to a PtI3moiety bound to His15
Authors: Tanley Simon W. M. , Starkey Laurina-Victoria , Lamplough Lucinda , Kaenket Surasek , Helliwell John R.
Published: August 2014

This study examines the binding and chemical stability of the platinum hexahalides K2PtCl6, K2PtBr6and K2PtI6when soaked into pre-grown hen egg-white lysozyme (HEWL) crystals as the protein host. Direct comparison of the iodo complex with the chloro and bromo complexes shows that the iodo complex is partly chemically transformed to a square-planar PtI3complex bound to the Nδatom of His15, a chemical behaviour that is not exhibited by the chloro or bromo complexes. Each complex does, however, bind to HEWL in its octahedral form either at one site (PtI6) or at two sites (PtBr6and PtCl6). As heavy-atom derivatives of a protein, the octahedral shape of the hexahalides could be helpful in cases of difficult-to-interpret electron-density maps as they would be recognisable `objects'.
Structural analysis of the GH43 enzyme Xsa43E fromButyrivibrio proteoclasticus
Authors: Till M. , Goldstone D. , Card G. , Attwood G. T. , Moon C. D. , Arcus V. L.
Published: August 2014

The rumen of dairy cattle can be thought of as a large, stable fermentation vat and as such it houses a large and diverse community of microorganisms. The bacteriumButyrivibrio proteoclasticusis a representative of a significant component of this microbial community. It is a xylan-degrading organism whose genome encodes a large number of open reading frames annotated as fibre-degrading enzymes. This suite of enzymes is essential for the organism to utilize the plant material within the rumen as a fuel source, facilitating its survival in this competitive environment. Xsa43E, a GH43 enzyme fromB. proteoclasticus, has been structurally and functionally characterized. Here, the structure of selenomethionine-derived Xsa43E determined to 1.3 Å resolution using single-wavelength anomalous diffraction is reported. Xsa43E possesses the characteristic five-bladed β-propeller domain seen in all GH43 enzymes. GH43 enzymes can have a range of functions, and the functional characterization of Xsa43E shows it to be an arabinofuranosidase capable of cleaving arabinose side chains from short segments of xylan. Full functional and structural characterization of xylan-degrading enzymes will aid in creating an enzyme cocktail that can be used to completely degrade plant material into simple sugars. These molecules have a range of applications as starting materials for many industrial processes, including renewable alternatives to fossil fuels.
Microseed matrix screening for optimization in protein crystallization: what have we learned?
Authors: D'Arcy Allan , Bergfors Terese , Cowan-Jacob Sandra W. , Marsh May
Published: August 2014

Protein crystals obtained in initial screens typically require optimization before they are of X-ray diffraction quality. Seeding is one such optimization method. In classical seeding experiments, the seed crystals are put into new, albeit similar, conditions. The past decade has seen the emergence of an alternative seeding strategy: microseed matrix screening (MMS). In this strategy, the seed crystals are transferred into conditions unrelated to the seed source. Examples of MMS applications from in-house projects and the literature include the generation of multiple crystal forms and different space groups, better diffracting crystals and crystallization of previously uncrystallizable targets. MMS can be implemented robotically, making it a viable option for drug-discovery programs. In conclusion, MMS is a simple, time- and cost-efficient optimization method that is applicable to many recalcitrant crystallization problems.
X-ray structures of Nfs2, the plastidial cysteine desulfurase fromArabidopsis thaliana
Authors: Roret Thomas , Pégeot Henri , Couturier Jérémy , Mulliert Guillermo , Rouhier Nicolas , Didierjean Claude
Published: August 2014

The chloroplasticArabidopsis thalianaNfs2 (AtNfs2) is a group II pyridoxal 5′-phosphate-dependent cysteine desulfurase that is involved in the initial steps of iron–sulfur cluster biogenesis. The group II cysteine desulfurases require the presence of sulfurtransferases such as SufE proteins for optimal activity. Compared with group I cysteine desulfurases, proteins of this group contains a smaller extended lobe harbouring the catalytic cysteine and have a β-hairpin constraining the active site. Here, two crystal structures of AtNfs2 are reported: a wild-type form with the catalytic cysteine in a persulfide-intermediate state and a C384S variant mimicking the resting state of the enzyme. In both structures the well conserved Lys241 covalently binds pyridoxal 5′-phosphate, forming an internal aldimine. Based on available homologous bacterial complexes, a model of a complex between AtNfs2 and the SufE domain of its biological partner AtSufE1 is proposed, revealing the nature of the binding sites.
Radial spacing distributions from planar point sets
Authors: Baake M. , Götze F. , Huck C. , Jakobi T.
Published: August 2014

This paper explores the radial projection method for locally finite planar point sets and provides numerical examples for different types of order. The main question is whether the method is suitable to analyse order in a quantitative way. The findings indicate that the answer is affirmative. In this context, local visibility conditions are also studied for certain types of aperiodic point sets.
Hirshfeld atom refinement for modelling strong hydrogen bonds
Authors: Woińska Magdalena , Jayatilaka Dylan , Spackman Mark A. , Edwards Alison J. , Dominiak Paulina M. , Woźniak Krzysztof , Nishibori Eiji , Sugimoto Kunihisa , Grabowsky Simon
Published: August 2014

High-resolution low-temperature synchrotron X-ray diffraction data of the salt L-phenylalaninium hydrogen maleate are used to test the new automated iterative Hirshfeld atom refinement (HAR) procedure for the modelling of strong hydrogen bonds. The HAR models used present the first examples ofZ′ > 1 treatments in the framework of wavefunction-based refinement methods. L-Phenylalaninium hydrogen maleate exhibits several hydrogen bonds in its crystal structure, of which the shortest and the most challenging to model is the O—H...O intramolecular hydrogen bond present in the hydrogen maleate anion (O...O distance is about 2.41 Å). In particular, the reconstruction of the electron density in the hydrogen maleate moiety and the determination of hydrogen-atom properties [positions, bond distances and anisotropic displacement parameters (ADPs)] are the focus of the study. For comparison to the HAR results, different spherical (independent atom model, IAM) and aspherical (free multipole model, MM; transferable aspherical atom model, TAAM) X-ray refinement techniques as well as results from a low-temperature neutron-diffraction experiment are employed. Hydrogen-atom ADPs are furthermore compared to those derived from a TLS/rigid-body (SHADE) treatment of the X-ray structures. The reference neutron-diffraction experiment reveals a truly symmetric hydrogen bond in the hydrogen maleate anion. Only with HAR is it possible to freely refine hydrogen-atom positions and ADPs from the X-ray data, which leads to the best electron-density model and the closest agreement with the structural parameters derived from the neutron-diffraction experiment,e.g.the symmetric hydrogen position can be reproduced. The multipole-based refinement techniques (MM and TAAM) yield slightly asymmetric positions, whereas the IAM yields a significantly asymmetric position.
Analysis of multicrystal pump–probe data sets. I. Expressions for the RATIO model
Authors: Fournier Bertrand , Coppens Philip
Published: August 2014

The RATIO method in time-resolved crystallography [Coppenset al.(2009).J. Synchrotron Rad.16, 226–230] was developed for use with Laue pump–probe diffraction data to avoid complex corrections due to wavelength dependence of the intensities. The application of the RATIO method in processing/analysis prior to structure refinement requires an appropriate ratio model for modeling the light response. The assessment of the accuracy of pump–probe time-resolved structure refinements based on the observed ratios was discussed in a previous paper. In the current paper, a detailed ratio model is discussed, taking into account both geometric and thermal light-induced changes.
Alianza: pricing to enter the pension industry
Authors: Farías Pablo
Published: August 2014

Subject area

The focus of the case is on the concepts of customer lifetime value (CLV) and customer equity (CE). Monitoring, measuring and maximizing CLV and CE have become a key priority for all marketers. Instructors can introduce these concepts and its key components.

The main focus of the case is a quantitative assignment that asks students to analyze the convenience for the existing five AFPs (Administradora de Fondos de Pensiones, Pension Fund Administrator) of winning the tender. The use of CLV and CE measurements is particularly relevant. Students need to estimate the impact of pricing on the CLV and CE of the existing five AFPs.

Study level/applicability

BA, MSc, MBA Courses: CE, Marketing Metrics, Pricing. The case can also be used in courses that focus on Marketing Plan, Marketing Research or Services Marketing.

Case overview

In early 2009, Valentina Vial was given the assignment to develop the pricing strategy of Alianza to enter the pension industry. The company will propose a commission fee to compete with the country's existing five AFPs. Whichever AFP presents the lowest commission will be awarded the tender. When there are several competitors, the company must guess each competitor's likely pricing decision. In the analysis of the convenience for the existing five AFPs of winning the tender, the use of CLV and CE measurements is particularly relevant. Valentina Vial needed to estimate the impact of pricing on the CLV and CE of the existing five AFPs.

Expected learning outcomes

Understand the concepts of CLV and CE and the importance of maximizing a customer's lifetime value for the firm by calculating the CLV and the CE based on a combination of financial and non-financial data.

Illustrate the importance of adopting a long-term strategic perspective (using CLV and CE) in choosing a pricing strategy. Once a firm commits to a pricing strategy, it is difficult to shift course. Given this, the choice of pricing levels should be informed by long-term strategic thinking, including consideration of potential competitive pricing decisions.

Supplementary materials

Teaching Notes are available for educators only. Please contact your library to gain login details or email support@emeraldinsight.com to request teaching notes.
A benchmarking framework for eTourism capability of destinations’ industries
Authors: Reino Sofia , J. Frew Andrew , Mitsche Nicole
Published: August 2014

Purpose

– This paper aims to provide a framework for benchmarking the eTourism capability of a destination’s tourism industry, understanding the eTourism capability of a destination’s industry as the contribution that the information and communication technologies (ICT) uptake of that industry makes to its own performance. The impact of ICT in tourism has been suggested through extensive research. Previous work has urged the development of wide-angle studies to enable benchmarking of destinations and their industries. However, relevant research is limited. Macro-level studies in the area tend to focus on a single aspect of technology to evaluate adoption, are not sector-specific nor take into consideration the different levels of contribution that systems may bring to performance.

Design/methodology/approach

– A review of the literature on tourism, eTourism, benchmarking and technology adoption provided the baseline for developing this benchmarking tool.

Findings

– The literature supported the selection of key tourism industry sectors, i.e. accommodation establishments, visitor attractions and food and beverage; the most suitable methodology, i.e. intermediary performance measures; and the business characteristics that need being taken into consideration when assessing ICT adoption by tourism businesses, e.g. size, type of establishment and area of location.

Research limitations/implications

– The framework has not been tested yet. It is based on a review of the literature and needs to be validated through primary research. The framework was developed based on the context of Scotland. Further work should be done to adjust the framework to other destinations worldwide.

Practical implications

– The framework enables destinations to benchmark the eTourism capability of their industries.

Originality/value

– It provides a comprehensive framework for benchmarking tourism destinations’ industries, which takes into consideration elements of technology adoption, the characteristics of the tourism industry and the particularities of the different ICT elements.
Exploring the impact of franchise support on franchisee acceptance of intranet in quick service restaurant (QSR) franchise system
Authors: Park Kunsoon , “Shawn” Lee Seungwon , A. Khan Mahmood
Published: August 2014

Purpose

– The purpose of this paper is twofold: to investigate the impact of franchisor support on quick service restaurant (QSR) users’ intranet acceptance; and to examine the technology acceptance model (TAM) to explain QSR users’ attitude toward the franchise intranet usage. Franchisors are building intranet systems as a communication tool to assist their franchisees in performing their jobs better. However, there is a little available literature that discusses in detail the use of the intranet by franchise systems.

Design/methodology/approach

– This study adopted the TAM to investigate the acceptance of the intranet in the context of QSR franchise systems. The original TAM was modified to include an external variable, franchise support. A total of 161 returned and completed responses were examined. Descriptive analysis, validity, principal component factor analysis and regression analysis were used to estimate the relationships between constructs.

Findings

– The key finding of this study is that franchise support is a key in the decision for users to use intranet systems in the QSR franchise systems. This study also confirmed that there are positive and significant relationships among key variables: franchise support, perceived usefulness of intranet, perceived ease of use of intranet, attitude toward using intranet and behavioral intentions to use intranet. Regression analysis revealed that TAM is a valid model in predicting intranet adoption in restaurant franchise systems.

Originality/value

– This study pursued franchise support, which was not included in the original TAM, to see how it affects perceived ease of use, perceived usefulness and intention to use. In addition, a list of recommendations for improving the situation in that franchise organization is provided.
Hotel and IT decision-maker characteristics and information technology adoption relationship in the hotel industry
Authors: Bulent Ozturk Ahmet , Hancer Murat
Published: August 2014

Purpose

– The purpose of this paper is to investigate the impact of hotel property characteristics and information technology (IT) decision-maker characteristics on radio frequency identification (RFID) technology adoption in the hotel industry.

Design/methodology/approach

– The data of the study were collected using the subscription list of Hospitality Financial and Technology Professionals (HFTP). A Web survey was used to collect the data of the study. An email invitation was sent out to invite the entire list of HFTP subscribers (3,080) to participate in the survey. Approximately 3,000 emails were delivered and 154 questionnaires were returned. Of 154 questionnaires, 125 were used in the study for further analyses. Analysis of variance (ANOVA) was used to determine whether hotel property characteristics (property size and chain affiliation) and IT decision-maker characteristics (age, education level and job tenure) differed on intention to adopt RFID technology.

Findings

– ANOVA results indicated that there were significant differences on intention to adopt RFID technology by property size and chain affiliation, IT decision-makers’ age, education level and job tenure. The results indicated that larger hotels and hotels that are part of a franchise are more likely to adopt RFID technology. In addition, IT decision-makers who are young, with high level of education and with shorter job tenure are more likely to adopt RFID technology.

Originality/value

– RFID technology is one of the recent technologies that gained great attention from the hotel operators in recent years. This study is one of the first studies in the hotel industry that provides valuable information to technology vendors for identifying potential RFID technology adopters in the hotel industry.
An empirical investigation of repeat buying behavior of customers of two brands in India
Authors: Baidya Mehir , Ghosh Gautam
Published: August 2014

Purpose

– The purpose of this paper is to attempt to test whether price and quality have any influence on customer repeat purchase probability. Managers should know the strategic determinants of repeat purchase probability to retain customers.

Design/methodology/approach

– Primary data were gathered on repeat purchase, price and quality via a survey of 400 customers of two brands in the Business-to-customer (B2C) sector. Then 380 × 4 data points to a logit model were fitted to estimate the effects of price and quality on customer repeat purchase probability.

Findings

– It was found that price, quality and price × quality influence customer repeat purchase probability substantially. Furthermore, the elasticities of repeat purchase probability to price and quality are quite high in both cases.

Practical implications

– Managers should use elasticities to determine how to allocate funds between price and quality to maximize repeat purchase probability.

Originality/value

– In view of the complexity of customer repeat buying behavior, this study deals with one aspect or feature of the total process, which does not mean that it has no value. By confirming two strategic determinants of customer repeat purchase probability and suggesting a simple rule of fund allocation, this work definitely adds some research of value to the relevant marketing literature.