<p>Flaviviruses are enveloped, positive-strand RNA viruses that are spread by mosquitoes and ticks and can cause serious disease in humans. Flavivirus virions undergo extensive structural changes during their life cycle, including during maturation and fusion. Flaviviruses are initially assembled at the endoplasmic reticulum in a non-infectious, immature state, and then traffic to the trans-Golgi network, where a pH drop triggers a structural rearrangement of glycoproteins prM and E on the virus surface from 60 trimers to 90 dimers. A host protease, furin, then cleaves prM which makes the transition irreversible. Upon exiting the host cell, pr disassociates from the virus and the infectious, mature virus is able to enter a new cell. <br></p><p><br></p> <p> </p> <p>In Chapter 1, an overview of flaviviruses is presented, including a brief history of their discovery and interaction with humans, followed by what is known about their life cycle and the maturation process. The structure of a mature flavivirus is then described, including the symmetrical arrangement of glycoproteins on the virion surface, the lipid membrane, and the nucleocapsid core, followed by an introduction of the structural proteins that assemble into the virion. The structure of the immature flavivirus is then described. The chapter concludes with a description of the dynamics and heterogeneity observed for flaviviruses.</p><p><br></p> <p> </p> <p>The conformational rearrangements that occur during flavivirus maturation remain unclear. The structures of immature and mature flaviviruses determined with cryo-electron microscopy (cryo-EM) demonstrated that flaviviruses are icosahedral particles with 180 copies of glycoproteins on their surface. Icosahedral viruses typically have a quasi-equivalent arrangement of glycoproteins, but flaviviruses lack quasi-equivalence and instead the three subunits within an asymmetric unit occupy different chemical environments. Although the subunits are the same proteins, the unique environment of each subunit can be exploited for tracking subunits during conformational rearrangements. For example, the unique labeling of a subunit can be used to identify it in the immature and mature virion.</p><p><br></p> <p> </p> <p>In Chapter 2, the maturation process was studied by developing tools to differentially label protein subunits and trap potential intermediates of maturation. The tools included heavy-atom compounds and antibody Fabs, which were used to probe Kunjin virus (KUNV), an Australian subtype of West Nile virus (WNV). One heavy-atom compound, potassium tetranitroplatinate(II), was found to derivatize immature KUNV, likely at sites on both E and prM. Higher-resolution studies will be required to determine if the compound differentially labeled the three subunits. The other tool developed was the E16 Fab. E16 Fab, originally isolated from a mouse immunized with WNV E and found to bind to two out of three subunits on mature WNV, was used to differentially label subunits in immature KUNV. Based on poor epitope accessibility on immature KUNV, E16 Fab was hypothesized to trap an intermediate state of maturation. In the cryo-EM reconstruction of E16 Fab bound to immature KUNV it was found that the virion had localized distorted density and apparent non-uniform binding of the E16 Fab. Based on this result it was proposed that flaviviruses had imperfect icosahedral symmetry. <br></p><p><br></p> <p> </p> <p>The structural asymmetry of immature and mature flaviviruses was investigated in Chapter 3. Icosahedral symmetry has always been imposed during cryo-EM reconstructions of flaviviruses, as it led to stable convergence of orientations. When reconstructions of immature KUNV and ZIKV were performed without imposing symmetry, the reconstructions showed that the flaviviruses had an eccentric nucleocapsid core, which was positioned closer to the membrane at one pole. At the opposite pole, the glycoprotein and inner leaflet densities were weak and distorted. Furthermore, there were protrusions from the core that contacted the transmembrane helices of the glycoproteins. In the asymmetric reconstruction of mature KUNV, the core was positioned concentric with the glycoprotein shell, in contrast to the immature virion, indicating that maturation alters the interactions between the core and the glycoproteins. The asymmetric reconstructions suggested that there is variable contact between the core and glycoproteins during assembly, which may be due to membrane curvature restrictions in the budding process. </p> <p> </p> <p><br></p><p>In Chapter 4, extracellular vesicles (EVs) that were released during dengue virus (DENV) infection were characterized by mass spectrometry. EVs may play a significant role in flavivirus infection, as they have been shown to transport both viral proteins and infectious RNA. EVs likely represent alternative modes of virus transmission and aid in immune evasion. However, previous studies on EVs are controversial because EVs are potential contaminated during assays by co-purifying virions and other particulates. The identification of EV biomarkers would greatly reduce contamination because biomarkers would enable isolation of pure EVs by affinity purification. Therefore, a strategy was developed to isolate EVs and profile them with proteomics. The four proteins cystatin-A, filamin B, fibrinogen beta chain, and endothelin converting enzyme 1 were found to be statistically enriched in the DENV sample and represent potential EV biomarkers. </p> <p> </p>

The fully halogenated or hydrogenated B12X12²- (X = H, F, Cl, Br and I) clusters are confirmed to be icosahedral.Onthe other hand, the bare B12 cluster is shown to have a planar structure.Aprevious study showed that a transformation from an icosahedron to a plane happens when 5 to 7 iodine atoms are remained [P. Farràs et al., Chem. - Eur. J. 18, 13208–13212 (2012)]. Later, the transition was confirmed to be seven iodine atoms based on an infrared spectroscopy study [M. R. Fagiania et al., Chem. Phys. Lett. 625, 48–52 (2015)]. In this study, we investigated the effects of different halogen atoms on the opening of the B12 icosahedral cage by means of density functional theory calculations.We found that the halogen elements do not have significant effects on the geometries of the clusters. The computed infrared (IR) spectra show similar representative peaks for all halogen doped clusters. Interestingly, we found a blue-shift in the IR spectra with the increase in the mass of the halogen atoms. Further, we compared the Gibbs free energies at different temperatures for different halogen atoms. The results show that the Gibbs free energy differences between open and close structures of B12X7- become larger when heavier halogen atoms are presented. This interesting finding was subsequently investigated by the energy decomposition analysis.

京都大学 / 0048 / 新制・課程博士 / 博士(工学) / 甲第19710号 / 工博第4165号 / 新制||工||1642(附属図書館) / 32746 / 京都大学大学院工学研究科材料工学専攻 / (主査)教授 酒井 明, 教授 河合 潤, 教授 中村 裕之 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DGAM

Master of Science / Department of Chemical Engineering / James H. Edgar / Pure boron exists in two main polymorphs, the common β-rhombohedral boron and the relatively rare α-rhombohedral boron. α-rhombohedral boron (α-B) possesses several extraordinary properties: self-healing from radiation damage and a high hole mobility. In addition, the [superscript]10B isotope has a large thermal neutron capture cross section. Such properties make it an excellent candidate for novel electronic device, such as direct energy conversion devices (alphacells and betacells) and neutron detectors. However, research on the properties and applications of α-B has been limited due to the difficulty to produce high quality α-B crystals of significant size. The preparation of α-rhombohedral boron is challenging for several reasons: first, α-rhombohedral boron has a low thermodynamic stability; it is only stable below 1100°C, at higher temperature β-rhombohedral boron is the stable polymorph. In addition, at elevated temperatures, boron is highly reactive, which make it is difficult to produce pure boron crystals. The primary goal of this research was to produce high quality α-B crystals of significant size. The main focus of this study was to explore the feasibility of producing α-B from a copper flux. Copper is a promising solvent for α-B crystal growth: the eutectic temperature of copper-boron is low, 996°C, and the phase diagram of copper-boron is relatively simple, and there are not many intermediate boride-copper compounds. In addition, copper is easily removed from crystals by etching with concentrated nitric acid. Last but not least, copper is less expensive than other metal solvents such as platinum. Boron crystal growth from a platinum solvent and vapor-liquid-solid growth by chemical vapor deposition were also performed for comparison. A series of crystals were grown over a range of initial boron concentrations (9.9 to 27.7 mole %) and cooling rates. Small irregular-shaped black crystals (>100μm) and well-faceted red crystals in various shapes, as large as 500 microns were produced. The crystals were characterized by optical microscopy, scanning electron microscopy, energy dispersive spectroscopy, x-ray diffraction analysis, and Raman spectroscopy. The correlation between experiment results and experimental parameters (source materials, the purity of growth atmosphere, and crucible materials, etc.) are reported. Suggestions about further investigation for α-B crystal growth are proposed.

This thesis is a theoretical study of configurational disorder in icosahedral boron-rich solids, in particular boron carbide, including also the development of a methodological framework for treating configurational disorder in such materials, namely superatom-special quasirandom structure (SA-SQS). In terms of its practical implementations, the SA-SQS method is demonstrated to be capable of efficiently modeling configurational disorder in icosahedral boron-rich solids, whiles the thermodynamic stability as well as the properties of the configurationally disordered icosahedral boron-rich solids, modeled from the SA-SQS method, can be directly investigated, using the density functional theory (DFT). In case of boron carbide, especially B4C and B13C2 compositions, the SA-SQS method is used for modeling configurational disorder, arising from a high concentration of low-energy B/C substitutional defects. The results, obtained from the DFT-based calculations, demonstrate that configurational disorder of B and C atoms in boron carbide is not only thermodynamically favored at high temperature, but it also plays an important role in altering the properties of boron carbide − for example, restoration of higher rhombohedral symmetry of B4C, a metal-to-nonmetal transition and a drastic increase in the elastic moduli of B13C2. The configurational disorder can also explain large discrepancies, regarding the proper- ties of boron carbide, between experiments and previous theoretical calculations, having been a long standing controversial issue in the field of icosahedral boron- rich solids, as the calculated properties of the disordered boron carbides are found to be in qualitatively good agreement with those, observed in experiments. In order to investigate the configurational evolution of B4C as a function of temperature, beyond the SA-SQS level, a brute-force cluster-expansion method in combination with Monte Carlo simulations is implemented. The results demonstrate that configurational disorder in B4C indeed essentially takes place within the icosahedra in a way that justifies the focus on lowenergy defect patterns of the superatom picture. The investigation of the thermodynamic stability of icosahedral carbon-rich boron carbides beyond the believed solubility limit of carbon (20 at.% C) demonstrates that, apart from B4C generally addressed in the literature, B2.5C represented by B10Cp2(CC) is predicted to be thermodynamically stable with respect to B4C as well as pure boron and carbon under high pressure, ranging between 40 and 67 GPa, and also at elevated temperature. B2.5C is expected to be metastable at ambient pressure, as indicated by its dynamical and mechanical stabilities at 0 GPa. A possible synthesis route of B2.5C and a fingerprint for its characterization from the simulations of x-ray powder diffraction pattern are suggested. Besides modeling configurational disorder in boron carbide, the SA-SQS method also opens up for theoretical studies of new alloys between different icosahedral boron-rich solids − for example, (B6O)1−x(B13C2)x and B12(As1−xPx)2. As for the pseudo-binary (B6O)1−x(B13C2)x alloy, it is predicted to display a miscibility gap resulting in B6O-rich and either ordered or disordered B13C2-rich domains for intermediate global compositions at all temperatures up to melting points of the materials. However, some intermixing of B6O and B13C2 to form solid solutions is also predicted at high temperature. A noticeable mutual solubility of icosahedral B12As2 and B12P2 in each other to form B12(As1−xPx)2 disordered alloy is predicted even at room temperature, and a complete closure of a pseudo-binary miscibility gap is achieved at around 900 K. Apart from B12(As1−xPx)2, the thermodynamic stability of other compounds and alloys in the ternary B-As-P system is also investigated. For the binary B-As system, zincblende BAs is found to be thermodynamically unstable with respect to icosahedral B12As2 and gray arsenic at 0 K and increasingly so at higher temperature, indicating that BAs may merely exist as a metastable phase. This is in contrast to the binary B-P system, in which zinc-blende BP and icosahedral B12P2 are both predicted to be stable. Owing to the instability of BAs with respect to B12As2 and gray arsenic, only a tiny amount of BAs is predicted to be able to dissolve in BP to form BAs1−xPx disordered alloy at elevated temperature. For example, less than 5% BAs can dissolve in BP at 1000 K. As for the binary As-P system, As1−xPx disordered alloys are predicted at elevated temperature − for example, a disordered solid solution of up to ∼75% As in black phosphorus as well as a small solubility of ∼1% P in gray arsenic at 750 K, together with the presence of miscibility gaps. The thermodynamic stability of three different compositions of α-rhombohedral boron-like boron subnitride, having been proposed so far in the literature, is investigated. Those are, B6N, B13N2, and B38N6, represented respectively by B12(N-N), B12(NBN), and [B12(N-N)]0.33[B12(NBN)]0.67. It is found that, out of these sub- nitrides, only B38N6 is thermodynamically stable from 0 GPa up to ∼7.5 GPa, depending on the temperature, and is thus concluded as a stable composition of α-rhombohedral boron-like boron subnitride.

A esfera é comumente usada como domínio computacional para representar o planeta Terra. Dessa forma, é possível modelar diversos fenômenos físicos, como a previsão numérica do tempo. A discretização pode ser feita de formas distintas, mas devido a uma crescente necessidade de eficiência computacional, as malhas geodésicas têm ganhado a atenção da comunidade científica. Dentre as quais, por serem mais isotrópicas em relação às malhas latitude/longitude, destacam-se as malhas icosaédricas. A qualidade dos modelos de previsão do tempo é fortemente influenciada pela precisão da solução da equação de advecção (ou transporte), pois, é necessário avaliar o transporte de diversas substâncias presentes na atmosfera. Nesse contexto, pesquisadores têm se interessado em desenvolver métodos de alta ordem na esfera para melhorar a qualidade da solução do transporte escalar. Apesar de existirem alguns modelos numéricos de alta ordem que usam malhas icosaédricas, não há consenso sobre as metodologias e os tipos de malhas a serem utilizadas. O objetivo deste trabalho foi estudar os métodos disponíveis na literatura e propor um novo método de alta ordem na esfera, baseado nos trabalhos de Ollivier-Gooch e colaboradores. O método de volumes finitos de alta ordem foi validado com testes de interpolação, integração e discretização do divergente. Por fim, foram utilizadas várias funções testes para a advecção. Os resultados foram comparados com os da literatura para malhas icosaédricas com distintas otimizações. Os testes incluem funções suaves, com descontinuidades e testes de deformações na distribuição do campo transportado, que são fundamentais no desenvolvimento de modelos atmosféricos globais. Os resultados numéricos mostram que o método proposto, que será denominado por FV-OLG, foi capaz de obter alta ordem de precisão e verificou-se que as taxas de erro são pouco influenciadas por distorções de malha. Foi feito um teste adicional para avaliar o transporte de uma colina de gaussiana na malha icosaédrica com refinamento local. Os resultados obtidos demonstram que as taxas de convergências são as mesmas obtidas em malhas com distintas otimizações, demonstrando ser um método robusto a ser explorado em modelos atmosféricos globais. / The sphere is commonly used as a computational domain to represent the planet Earth. In this way, it is possible to model several physical phenomena, such as the numerical weather forecast. Discretization can be done in different ways, but due to an increasing need for computational efficiency, geodesic meshes have gained the attention of the scientific community. These are more isotropic in relation to the latitude / longitude meshes, among which, the icosahedral meshes stand out. The quality of weather forecast models is strongly influenced by the accuracy of the solution of the advection (or transport) equation, since it is necessary to evaluate the transport of various substances present in the atmosphere. In this context, researchers have been interested in developing high-order methods on the sphere to improve the quality of the scalar transport solution. Although there are some high order numerical models that use icosahedral meshes, there is no consensus on the methodologies and types of meshes to be used. The objective of this work was to study the methods available in the literature and to propose a new high order method in the sphere, based on the works of Ollivier-Gooch et al. The finite-order finite-volume method was validated with inter- polation, integration and discretization tests of the divergent. For this purpose, several tests were used for the advection and the results were compared with those from the literature for icosahedral meshes with different optimizations. The tests include smooth functions, with discontinuities and tests of deformations in the distribution of the transported field, which are fundamental in the development of global atmospheric models. The numerical results show that the proposed method, which will be called FV-OLG, was able to obtain a high order of accuracy and verified that the error rates are little influenced by mesh distortion. An additional test was carried out to evaluate the transport of a Gaussian hill in the icosahedral grid with local refinement. The results show that the convergence rates are the same as those obtained in meshes with different optimizations, demonstrating that it is a robust method to be used in global atmospheric models.

No description available.

Viruses are the most abundant biological entity in the biosphere and are known to infect hosts from all domains of life. The aim of my work is to identify conserved and non-conserved features among the capsid structures of related and divergent icosahedral viruses via cryogenic electron microscopy, sequence analysis, molecular modeling, and other techniques. Bird polyomaviruses often cause severe disease in their hosts whereas mammalian polyomaviruses generally do not. Avian polyomavirus is a type of bird polyomavirus with an unusually broad host range compared to the restricted tropism of other polyomaviruses. Although most polyomaviruses have a conserved, rigid capsid protein structure, avian polyomavirus has a flexible capsid shell and a non-conserved C-terminus in its major capsid protein. A β-hairpin motif appears to stabilize other polyomaviruses but is missing in avian polyomavirus. The lack of this structure in avian polyomavirus may account for its capsid flexibility and broad host range. A minor capsid protein unique to bird polyomaviruses may be located on the inner capsid surface. This protein may have a role in the acute disease caused by bird polyomaviruses. The solution-state capsid structure of satellite tobacco mosaic virus was unexpectedly different than the previously solved crystalline structure. The conformational differences were accounted for by a shift of the capsid protein about the icosahedral fivefold axis. Conversely, the RNA core was consistent between solution and crystalline structures. The stable RNA core supports previous observations that the viral genome stabilizes the flexible capsid. Halophage CW02 infects Salinivibrio bacteria in the Great Salt Lake. The three-dimensional structure of CW02 revealed a conserved HK97-like fold that is found in all tailed, double-stranded DNA viruses. The capsid sequence of CW02 shares less than 20% identity with HK97-like viruses, demonstrating that structure is more conserved than sequence. A conserved module of genes places CW02 in the viral T7 supergroup, members of which are found in diverse aquatic environments. No tail structure was observed in reconstructions of CW02, but turret-like densities were found on each icosahedral vertex, which may represent unique adaptations similar to those seen in other extremophilic viruses.

Emerging arthropod-borne viruses, such as alphaviruses and bunyaviruses, represent a serious threat to human and animal health worldwide, and for most of them, vaccines and specific treatments are unavailable. Viral host cell entry can be divided into several entry checkpoints, and the most important checkpoints for low pH-dependent enveloped viruses, such as bunyaviruses and alphaviruses, include receptor binding at the cell surface and, followed by endocytosis, low pH dependent membrane fusion from within intracellular compartments. A more thorough understanding of the detailed mechanisms allowing the viruses to pass these checkpoints is a pre-requisite for the design of viral entry inhibitors. This thesis reports the in vitro analysis of native alphavirus-receptor interactions, with the help of electron cryo-microscopy and icosahedral reconstruction of virus-recaptor complexes, using the prototypic alphavirus Semliki Forest virus (SFV) and the C-type lectin DC-SIGN. Together with results from collaborative work on SFV glycosylation, this study provides progress in defining the binding sites of DC-SIGN at the surface of SFV. Second, an in vitro system for phlebovirus fusion was developed using standard fluorometry, and has been characterized with the help of electron cryo-microscopy. It was discovered that negatively charged phospholipids with a conical shape, including the late endosomal phospholipid BMP, allow efficient phlebovirus fusion in vitro, thereby providing a possible rationale for phlebovirus fusion in late endosomes. Furthermore, electron cryo-microscopy of phlebovirus-liposome complexes allowed the capture of early stage fusion intermediates and laid the basis for possible future higher resolution studies of these fusion intermediates.

Le nombre croissant de nanostructures physiques et biologiques sont caractérisées par l'ordre non-cristallin et par les propriétés physiques et biologiques non-conventionnels. Parmi ses systèmes il faut distinguer les capsides virales. Ces coquilles solides qui sont formées par un certain nombre dec opies de la même protéine protègent le virus des agressions et facilitent le processus d'infection de la cellule hôte. La distribution des positions de protéines dans une capside est très régulière et montre un degré très élevé d'ordre, aussi bien orientationnel que positionnel. Les capsides virales de topologie sphérique possèdent la symétrie icosaédrique compatible avec l'ordre cristallin local, mais incompatible avec la symétrie cristalline globale et interdite dans les structures périodiques.Ici, sur l'exemple des Papovavirus, nous montrons l'existence d'un nouveau type d'organisation qui résulte dans l'ordre quasicristallin pentagonal chiral de protéines dans des capsides de topologie sphérique et géométrie dodécaédrique. La formation de cet ordre est décrite dans le cadre de la théorie de Landau de cristallisation. Les particularités de la structure sont élucidées grâce à la théorie d'élasticité des quasicristaux comme le résultat de la déformation phason nonlinéaire.La généralisation de la théorie de Landau de cristallisation que nous proposons permet également de décrire des structures quasicristallines octogonales et décagonales grâce à la minimisation contrainte de l'énergie libre, et donne un nouveau sens physique à la notion de « fenêtre de projection » utilisée dans la cristallographie multidimensionnelle. / A growing number of physical and biological nanostructures are characterized by non-crystallineorder and by unconventional physical and biological properties. Among these systems one can distinguish viral capsids. These solid shells formed by a certain number of copies of the same protein protect viruses from aggressions and facilitate infection of the host cell. Protein distributionin a capsid is quite regular and shows high degree of order, both orientational and positional. Viral capsids with spherical topology have icosahedral symmetry compatible with local crystalline orderbut incompatible with the global one and forbidden in periodic structures.Here, on the example of Papovaviruses we show the existence of a new type of organization whichresults in the chiral pentagonal quasicrystalline order of proteins in capsids with spherical topology and dodecahedral geometry. The formation of this order is described in the frame of the Landau theory of crystallization. The theory of elasticity of quasicrystals is used to show that the structure peculiarities result from the non-linear phason strain.Generalization of the proposed Landau theory of crystallization allows us to describe octagonal and decagonal quasicrystalline structures using constrained minimization of the free energy, thus giving a new physical sense to the « projection window » notion used in multi-dimensionalcrystallography.