High-Pressure Behavior and Phase Stability of Al5BO9, a Mullite-Type Ceramic Material

Phase stability, elastic behavior, and pressure-induced structural evolution of synthetic boron-mullite Al5BO9 (a = 5.6780(7), b = 15.035(6), and c =7.698(3) Å, space group Cmc21, Z = 4) were investigated up to 25.6(1) GPa by in situ single-crystal synchrotron X-ray diffraction with a diamond anvil cell (DAC) under hydrostatic conditions. No evidence of phase transition was observed up to 21.7(1) GPa. At 25.6(1) GPa, the refined unit-cell parameters deviated significantly from the compressional trend, and the diffraction peaks appeared broader than at lower pressure. At 26.7(1) GPa, the diffraction pattern was not indexable, suggesting amorphization of the material or a phase transition to a high-pressure polymorph. Fitting the P–V data up to 21.7(1) GPa with a second-order Birch–Murnaghan Equation-of-State, we obtained a bulk modulus KT0 = 164(1) GPa. The axial compressibilities, here described as linearized bulk moduli, are as follows: KT0(a) = 244(9), KT0(b) = 120(4), and KT0(c) = 166(11) GPa (KT0(a):KT0(b):KT0(c) = 2.03:1:1.38). The structure refinements allowed a description of the main deformation mechanisms in response to the applied pressure. The stiffer crystallographic direction appears to be controlled by the infinite chains of edge-sharing octahedra running along [100], making the structure less compressible along the a-axis than along the b- and c-axis.

Does the (structural) context matter? Analysing individual and structural factors of members’ commitment to sports clubs

Introduction: Over the last decades, Swiss sports clubs have lost their "monopoly" in the market for sports-related services and increasingly are in competition with other sports providers. For many sport clubs long-term membership cannot be seen as a matter of course. Current research on sports clubs in Switzerland – as well as for other European countries – confirms the increasing difficulties in achieving long-term member commitment. Looking at recent findings of the Swiss sport clubs report (Lamprecht, Fischer & Stamm, 2012), it can be noted, that a decrease in memberships does not equally affect all clubs. There are sports clubs – because of their specific situational and structural conditions – that have few problems with member fluctuation, while other clubs show considerable declines in membership. Therefore, a clear understanding of individual and structural factors that trigger and sustain member commitment would help sports clubs to tackle this problem more effectively. This situation poses the question: What are the individual and structural determinants that influence the tendency to continue or to quit the membership? Methods: Existing research has extensively investigated the drivers of members’ commitment at an individual level. As commitment of members usually occurs within an organizational context, the characteristics of the organisation should be also considered. However, this context has been largely neglected in current research. This presentation addresses both the individual characteristics of members and the corresponding structural conditions of sports clubs resulting in a multi-level framework for the investigation of the factors of members’ commitment in sports clubs. The multilevel analysis grant a adequate handling of hierarchically structured data (e.g., Hox, 2002). The influences of both the individual and context level on the stability of memberships are estimated in multi-level models based on a sample of n = 1,434 sport club members from 36 sports clubs. Results: Results of these multi-level analyses indicate that commitment of members is not just an outcome of individual characteristics, such as strong identification with the club, positively perceived communication and cooperation, satisfaction with sports clubs’ offers, or voluntary engagement. It is also influenced by club-specific structural conditions: stable memberships are more probable in rural sports clubs, and in clubs that explicitly support sociability, whereas sporting-success oriented goals in clubs have a destabilizing effect. Discussion/Conclusion: The proposed multi-level framework and the multi-level analysis can open new perspectives for research concerning commitment of members to sports clubs and other topics and problems of sport organisation research, especially in assisting to understand individual behavior within organizational contexts. References: Hox, J. J. (2002). Multilevel analysis: Techniques and applications. Mahwah: Lawrence Erlbaum. Lamprecht, M., Fischer, A., & Stamm, H.-P. (2012). Die Schweizer Sportvereine – Strukturen, Leistungen, Herausforderungen. Zurich: Seismo.

Stability and kinetics of nucleic acid triplexes with chimaeric DNA/RNA third strands

A series of chimaeric DNA/RNA triplex-forming oligonucleotides (TFOs) with identical base-sequence but varying sequential composition of the sugar residues were prepared. The structural, kinetic and thermodynamic properties of triplex formation with their corresponding double-helical DNA target were investigated by spectroscopic methods. Kinetic and thermodynamic data were obtained from analysis of non-equilibrium UV-melting- and annealing curves in the range of pH 5.1 to 6.7 in a 10 mM citrate/phosphate buffer containing 0.1M NaCl and 1 mM EDTA. It was found that already single substitutions of ribo- for deoxyribonucleotides in the TFOs greatly affect stability and kinetics of triplex formation in a strongly sequence dependent manner. Within the sequence context investigated, triplex stability was found to increase when deoxyribonucleotides were present at the 5'-side and ribonucleotides in the center of the TFO. Especially the substitution of thymidines for uridines in the TFO was found to accelerate both, the association and dissociation process, in a strongly position-dependent way. Differential structural information on triplexes and TFO single-strands was obtained from CD-spectroscopy and gel mobility experiments. Only minor changes were observed in the CD spectra of the triplexes at all pH values investigated, and the electrophoretic mobility was nearly identical in all cases, indicating a high degree of structural similarity. In contrast, the single-stranded TFOs showed high structural variability as determined in the same way. The results are discussed in the context of the design of TFOs for therapeutic or biochemical applications.

Synthesis and Structural Elucidation of Triazolylmolybdenum(VI) Oxide Hybrids and Their Behavior as Oxidation Catalysts

A large family of bifunctional 1,2,4-triazole molecular tectons (tr) has been explored for engineering molybdenum(VI) oxide hybrid solids. Specifically, tr ligands bearing auxiliary basic or acidic groups were of the type amine, pyrazole, 1H-tetrazole, and 1,2,4-triazole. The organically templated molybdenum(VI) oxide solids with the general compositions [MoO3(tr)], [Mo2O6(tr)], and [Mo2O6(tr)(H2O)2] were prepared under mild hydrothermal conditions or by refluxing in water. Their crystal structures consist of zigzag chains, ribbons, or helixes of alternating cis-{MoO4N2} or {MoO5N} polyhedra stapled by short [N–N]-tr bridges that for bitriazole ligands convert the motifs into 2D or 3D frameworks. The high thermal (235–350 °C) and chemical stability observed for the materials makes them promising for catalytic applications. The molybdenum(VI) oxide hybrids were successfully explored as versatile oxidation catalysts with tert-butyl hydroperoxide (TBHP) or aqueous H2O2 as an oxygen source, at 70 °C. Catalytic performances were influenced by the different acidic–basic properties and steric hindrances of coordinating organic ligands as well as the structural dimensionality of the hybrid.

Bridged bicyclic peptides as potential drug scaffolds: synthesis, structure, protein binding and stability

Double cyclization of short linear peptides obtained by solid phase peptide synthesis was used to prepare bridged bicyclic peptides (BBPs) corresponding to the topology of bridged bicyclic alkanes such as norbornane. Diastereomeric norbornapeptides were investigated by 1H-NMR, X-ray crystallography and CD spectroscopy and found to represent rigid globular scaffolds stabilized by intramolecular backbone hydrogen bonds with scaffold geometries determined by the chirality of amino acid residues and sharing structural features of β-turns and α-helices. Proteome profiling by capture compound mass spectrometry (CCMS) led to the discovery of the norbornapeptide 27c binding selectively to calmodulin as an example of a BBP protein binder. This and other BBPs showed high stability towards proteolytic degradation in serum.

Structural elucidation and antisense properties of a sugar-modified DNA analogue

Antisense oligonucleotides deserve great attention as potential drug candidates for the treatment of genetic disorders. For example, muscle dystrophy can be treated successfully in mice by antisense-induced exon skipping in the pre-mRNA coding for the structural protein dystrophin in muscle cells. For this purpose a sugar- and backbone-modified DNA analogue was designed, in which a tricyclic ring system substitutes the deoxyribose. These chemical modifications stabilize the dimers formed with the targeted RNA relative to native nucleic acid duplexes and increase the biostability of the antisense oligonucleotide. While evading enzymatic degradation constitutes an essential property of antisense oligonucleotides for therapeutic application, it renders the oligonucleotide inaccessible to biochemical sequencing techniques and requires the development of alternative methods based on mass spectrometry. The set of sequences studied includes tcDNA oligonucleotides ranging from 10 to 15 nucleotides in length as well as their hybrid duplexes with DNA and RNA complements. All samples were analyzed on a LTQ Orbitrap XL instrument equipped with a nano-electrospray source. For tandem mass spectrometric experiments collision-induced dissociation was performed, using helium as collision gas. Mass spectrometric sequencing of tcDNA oligomers manifests the applicability of the technique to substrates beyond the scope of enzyme-based methods. Sequencing requires the formation of characteristic backbone fragments, which take the form of a-B- and w-ions in the product ion spectra of tcDNA. These types of product ions are typically associated with unmodified DNA, which suggests a DNA-like fragmentation mechanism in tcDNA. The loss of nucleobases constitutes the second prevalent dissociation pathway observed in tcDNA. Comparison of partially and fully modified oligonucleotides indicates a pronounced impact of the sugar-moiety on the base loss. As this event initiates cleavage of the backbone, the presented results provide new mechanistic insights into the fragmentation of DNA in the gas-phase. The influence of the sugar-moiety on the dissociation extends to tcDNA:DNA and tcDNA:RNA hybrid duplexes, where base loss was found to be much more prominent from sugar-modified oligonucleotides than from their natural complements. Further prominent dissociation channels are strand separation and backbone cleavage of the single strands, as well as the ejection of backbone fragments from the intact duplex. The latter pathway depends noticeably on the base sequence. Moreover, it gives evidence of the high stability of the hybrid dimers, and thus directly reflects the affinity of tcDNA for its target in the cell. As the cellular target of tcDNA is a pre-mRNA, the structure was designed to discriminate RNA from DNA complements, which could be demonstrated by mass spectrometric experiments.

Murine coronavirus ubiquitin-like domain is important for papain-like protease stability and viral pathogenesis

Ubiquitin-like domains (Ubls) now are recognized as common elements adjacent to viral and cellular proteases; however, their function is unclear. Structural studies of the papain-like protease (PLP) domains of coronaviruses (CoVs) revealed an adjacent Ubl domain in severe acute respiratory syndrome CoV, Middle East respiratory syndrome CoV, and the murine CoV, mouse hepatitis virus (MHV). Here, we tested the effect of altering the Ubl adjacent to PLP2 of MHV on enzyme activity, viral replication, and pathogenesis. Using deletion and substitution approaches, we identified sites within the Ubl domain, residues 785 to 787 of nonstructural protein 3, which negatively affect protease activity, and valine residues 785 and 787, which negatively affect deubiquitinating activity. Using reverse genetics, we engineered Ubl mutant viruses and found that AM2 (V787S) and AM3 (V785S) viruses replicate efficiently at 37°C but generate smaller plaques than wild-type (WT) virus, and AM2 is defective for replication at higher temperatures. To evaluate the effect of the mutation on protease activity, we purified WT and Ubl mutant PLP2 and found that the proteases exhibit similar specific activities at 25°C. However, the thermal stability of the Ubl mutant PLP2 was significantly reduced at 30°C, thereby reducing the total enzymatic activity. To determine if the destabilizing mutation affects viral pathogenesis, we infected C57BL/6 mice with WT or AM2 virus and found that the mutant virus is highly attenuated, yet it replicates sufficiently to elicit protective immunity. These studies revealed that modulating the Ubl domain adjacent to the PLP reduces protease stability and viral pathogenesis, revealing a novel approach to coronavirus attenuation. IMPORTANCE Introducing mutations into a protein or virus can have either direct or indirect effects on function. We asked if changes in the Ubl domain, a conserved domain adjacent to the coronavirus papain-like protease, altered the viral protease activity or affected viral replication or pathogenesis. Our studies using purified wild-type and Ubl mutant proteases revealed that mutations in the viral Ubl domain destabilize and inactivate the adjacent viral protease. Furthermore, we show that a CoV encoding the mutant Ubl domain is unable to replicate at high temperature or cause lethal disease in mice. Our results identify the coronavirus Ubl domain as a novel modulator of viral protease stability and reveal manipulating the Ubl domain as a new approach for attenuating coronavirus replication and pathogenesis.

Two-Dimensional Crystallisation of Membrane Proteins and Structural Assessment

Two-dimensional (2D) crystallisation of Membrane proteins reconstitutes them into their native environment, the lipid bilayer. Electron crystallography allows the structural analysis of these regular protein–lipid arrays up to atomic resolution. The crystal quality depends on the protein purity, ist stability and on the crystallisation conditions. The basics of 2D crystallisation and different recent advances are reviewed and electron crystallography approaches summarised. Progress in 2D crystallisation, sample preparation, image detectors and automation of the data acquisition and processing pipeline makes 2D electron crystallography particularly attractive for the structural analysis of membrane proteins that are too small for single-particle analyses and too unstable to form three-dimensional (3D) crystals.

IGHD II: A Novel GH-1 Gene Mutation (GH-L76P) Severely Affects GH Folding, Stability, and Secretion

CONTEXT The autosomal dominant form of GH deficiency (IGHD II) is characterized by markedly reduced GH secretion combined with low concentrations of IGF-1 leading to short stature. OBJECTIVE Structure-function analysis of a missense mutation in the GH-1 gene converting codon 76 from leucine (L) to proline (P) yielding a mutant GH-L76P peptide. DESIGN, SETTINGS, AND PATIENTS Heterozygosity for GH-L76P/wt-GH was identified in a nonconsanguineous Spanish family. The index patients, two siblings, a boy and a girl, were referred for assessment of their short stature (-3.2 and -3.8 SD). Their grandmother, father, and aunt were also carrying the same mutation and showed severe short stature; therefore, IGHD II was diagnosed. INTERVENTIONS AND RESULTS AtT-20 cells coexpressing both wt-GH and GH-L76P showed a reduced GH secretion (P < .001) after forskolin stimulation when compared with the cells expressing only wt-GH. In silico mutagenesis and molecular dynamics simulations presented alterations of correct folding and mutant stability compared with wt-GH. Therefore, further structural analysis of the GH-L76P mutant was performed using expressed and purified proteins in Escherichia coli by thermofluor assay and fast degradation proteolysis assay. Both assays revealed that the GH-L76P mutant is unstable and misfolded compared to wt-GH confirming the bioinformatic model prediction. CONCLUSIONS This is the first report of a family suffering from short stature caused by IGHD II, which severely affects intracellular GH folding and stability as well as secretion, highlighting the necessity of functional analysis of any GH variant for defining new mechanisms as a cause for IGHD II.

Structural conservations and diversities of dsRNA viruses: Structural studies of the cytoplasmic polyhedrosis virus by electron cryomicroscopy and 3D reconstruction

The Reoviridae virus family is a group of economically and pathologically important viruses that have either single-, double-, or triple-shelled protein layers enclosing a segmented double stranded RNA genome. Each virus particle in this family has its own viral RNA dependent RNA polymerase and the enzymatic activities necessary for the mature RNA synthesis. Based on the structure of the inner most cores of the viruses, the Reoviridae viruses can be divided into two major groups. One group of viruses has a smooth surfaced inner core, surrounded by complete outer shells of one or two protein layers. The other group has an inner core decorated with turrets on the five-fold vertices, and could either completely lack or have incomplete outer protein layers. The structural difference is one of the determinant factors for their biological differences during the infection. ^ Cytoplasmic polyhedrosis virus (CPV) is a single-shelled, turreted virus and the structurally simplest member in Reoviridae. It causes specific chronic infections in the insect gut epithelial cells. Due to its wide range of insect hosts, CPV has been engineered as a potential insecticide for use in fruit and vegetable farming. Its unique structural simplicity, unparalleled capsid stability and ease of purification make CPV an ideal model system for studying the structural basis of dsRNA virus assembly at the highest possible resolution by electron cryomicroscopy (cryoEM) and three-dimensional (3D) reconstruction. ^ In this thesis work, I determined the first 3D structure of CPV capsids using 100 kV cryoEM. At an effective resolution of 17 Å, the full capsid reveals a 600-Å diameter, T = 1 icosahedral shell decorated with A and B spikes at the 5-fold vertices. The internal space of the empty CPV is unoccupied except for 12 mushroom-shaped densities that are attributed to the transcriptional enzyme complexes. The inside of the full capsid is packed with icosahedrally-ordered viral genomic RNA. The interactions of viral RNA with the transcriptional enzyme complexes and other capsid proteins suggest a mechanism for RNA transcription and subsequent release. ^ Second, the interactions between the turret proteins (TPs) and the major capsid shell protein (CSPs) have been identified through 3D structural comparisons of the intact CPV capsids with the spikeless CPV capsids, which were generated by chemical treatments. The differential effects of these chemical treatment experiments also indicated that CPV has a significantly stronger structural integrity than other dsRNA viruses, such as the orthoreovirus subcores, which are normally enclosed within outer protein shells. ^ Finally, we have reconstructed the intact CPV to an unprecendented 8 Å resolution from several thousand of 400kV cryoEM images. The 8 Å structure reveals interactions among the 120 molecules of each of the capsid shell protein (CSP), the large protrusion protein (LPP), and 60 molecules of the turret protein (TP). A total of 1980 α-helices and 720 β-sheets have been identified in these capsid proteins. The CSP structure is largely conserved, with the majority of the secondary structures homologous to those observed in the x-ray structures of corresponding proteins of other reoviruses, such as orthoreovirus and bluetongue virus. The three domains of TP are well positioned to play multifunctional roles during viral transcription. The completely non-equivalent interactions between LPP and CSP and those between the anchoring domain of TP and CSP account for the unparalleled stability of this structurally simplest member of the Reoviridae. ^

Mineralogy and structural specification of ODP Leg 125 peridotite samples

Abundant serpentinite seamounts are found along the outer high of the Mariana forearc at the top of the inner slope of the trench. One of them, Conical Seamount, was drilled at Sites 778, 779, and 780 during Leg 125. The rocks recovered at Holes 779A and 780C, respectively, on the flanks and at the summit of the seamount, include moderately serpentinized depleted harzburgites and some dunites. These rocks exhibit evidence of resorption of the orthopyroxene, when present, and the local presence of very calcic-rich diopside in veins oblique to the main high-temperature foliation of the rock. The peridotites, initially well-foliated with locally poikiloblastic textures, show overprints of a two-stage deformation history: (1) a high-temperature (>1000°C), low-stress (0.02 GPa), homogeneous deformation that has led to the present Porphyroclastic textures displayed by the rocks and (2) heterogeneous ductile shearing at a much higher stress (0.05 GPa). This heterogeneous shearing probably describes a single tectonic event because it began at high temperatures, producing dynamic recrystallization of olivine in the shear zone, and ended at low temperatures in the stability field of chlorite and serpentine. In a few samples, olivine shows evidence of quasi-hydrostatic recrystallization at a very high temperature. Here, we propose that this recrystallization was related to fluid/magma percolation, a process that can also account for the resorption of the orthopyroxene and for the late crystallization of diopside veins in the rock. The impregnation by fluid or magma, development of the main high-temperature, low-stress deformation, and subsequent migration recrystallization of olivine probably occurred in a mantle fragment involved in the arc formation. In addition, this mantle has preserved structures that may have formed earlier in the oceanic lithosphere upon which the arc formed. Heterogeneous ductile shear zones in the peridotites may have developed during uplift. The "cold" deformation may have taken place during diapiric rise of hot mantle that underwent subsequent serpentinization or gliding along normal faults associated with the extension of the eastern margin of the forearc.

The Islamic Crossed-Arch Domes in Cordoba: Geometry and Structural Analysis of the “Capilla de Villaviciosa”

Crossed-arch domes are a singular type of ribbed vaults. Their characteristic feature is that the ribs that form the vault are intertwined, forming polygons or stars, leaving an empty space in the centre. The earliest known vaults of this type are found in the Great Mosque of Córdoba, built ca. 960 a.C. The type spread through Spain, and the north of Africa in the 10th to the 16th Centuries, and was used by Guarini and Vittone in the 17th and 18th Centuries in Italy. However, it was used only in a few buildings. Though the literature about the structural behaviour of ribbed Gothic vaults is extensive, so far no structural analysis of crossed arch domes has been made. The purpose of this work is, first to show the way to attack such an analysis within the frame of Modern Limit Analysis of Masonry Structures (Heyman 1995), and then to apply the approach to study the stability of the dome of the Capilla de Villaviciosa. The work may give some clues to art and architectural historians to understand better the origin and development of Islamic dome architecture.

The Ciborium or Lantern Tower of Valencia Cathedral: Geometry, Construction and Stability

Early 18th century treatise writer Tomas Vicente Tosca1 includes in his Tratado de la montea y cortes de Canteria [On Masonry Design and Stone Cutting], what is an important documentary source about the lantern of Valencia Cathedral. Tosca writes about this lantern as an example of vaulting over cross arches without the need of buttresses. A geometrical description is followed by an explanation of the structural behavior which manifests his deep understanding of the mechanics of masonry structures. He tries to demonstrate the absence of buttresses supporting his thesis on the appropriate distribution of loads which will reduce the "empujos" [horizontal thrusts] to the point of not requiring more than the thickness of the walls to stand (Tosca [1727] 1992, 227-230). The present article2 assesses T osca' s appreciation studying how loads and the thrusts they generate are transmitted through the different masonry elements that constitute this ciborium. In order to do so, we first present a geometrical analysis and make considerations regarding its materials and construction methods to, subsequently, analyze its stability adopting an equilibrium approach within the theoretical framework of the lower bound limit analysis.

The Ciborium or Lantern Tower of Valencia Cathedral: Geometry, Construction and Stability

Early 18th century treatise writer Tomas Vicente Tosca1 includes in his Tratado de la montea y cortes de Canteria [On Masonry Design and Stone Cutting], what is an important documentary source about the lantern of Valencia Cathedral. Tosca writes about this lantern as an example of vaulting over cross arches without the need of buttresses. A geometrical description is followed by an explanation of the structural behavior which manifests his deep understanding of the mechanics of masonry structures. He tries to demonstrate the absence of buttresses supporting his thesis on the appropriate distribution of loads which will reduce the "empujos" [horizontal thrusts] to the point of not requiring more than the thickness of the walls to stand (Tosca [1727] 1992, 227-230). The present article2 assesses T osca' s appreciation studying how loads and the thrusts they generate are transmitted through the different masonry elements that constitute this ciborium. In order to do so, we first present a geometrical analysis and make considerations regarding its materials and construction methods to, subsequently, analyze its stability adopting an equilibrium approach within the theoretical framework of the lower bound limit analysis.