Interstitial defects stability in FeCr alloys vs Cr concentration

Finding adequate materials to withstand the demanding conditions in the future fusion and fission reactors is a real challenge in the development of these technologies. Structural materials need to sustain high irradiation doses and temperatures that will change the microstructure over time. A better understanding of the changes produced by the irradiation will allow for a better choice of materials, ensuring a safer and reliable future power plants. High-Cr ferritic/martensitic steels head the list of structural materials due to their high resistance to swelling and corrosion. However, it is well known that these alloys present a problem of embrittlement, which could be caused by the presence of defects created by irradiation as these defects act as obstacles for dislocation motion. Therefore, the mechanical response of these materials will depend on the type of defects created during irradiation. In this work, we address a study of the effect Cr concentration has on single interstitial defect formation energies in FeCr alloys.

Resilience analysis of networked systems-of-systems based on structural and dynamic interdependencies

Critical infrastructures support everyday activities in modern societies, facilitating the exchange of services and quantities of various nature. Their functioning is the result of the integration of diverse technologies, systems and organizations into a complex network of interconnections. Benefits from networking are accompanied by new threats and risks. In particular, because of the increased interdependency, disturbances and failures may propagate and render unstable the whole infrastructure network. This paper presents a methodology of resilience analysis of networked systems of systems. Resilience generalizes the concept of stability of a system around a state of equilibrium, with respect to a disturbance and its ability of preventing, resisting and recovery. The methodology provides a tool for the analysis of off-equilibrium conditions that may occur in a single system and propagate through the network of dependencies. The analysis is conducted in two stages. The first stage of the analysis is qualitative. It identifies the resilience scenarios, i.e. the sequence of events, triggered by an initial disturbance, which include failures and the system response. The second stage is quantitative. The most critical scenarios can be simulated, for the desired parameter settings, in order to check if they are successfully handled, i.e recovered to nominal conditions, or they end into the network failure. The proposed methodology aims at providing an effective support to resilience-informed design.

Stability of vacancy clusters in FeCr alloys: A study of the Cr concentration dependence

Fe–Cr based alloys are the leading structural material candidates in the design of next generation reactors due to their high resistance to swelling and corrosion. Despite these good properties there are others, such as embrittlement, which require a higher level of understanding in order to improve aspects such as safety or lifetime of the reactors. The addition of Cr improves the behavior of the steels under irradiation, but not in a monotonic way. Therefore, understanding the changes in the Fe–Cr based alloys microstructure induced by irradiation and the role played by the alloying element (Cr) is needed in order to predict the response of these materials under the extreme conditions they are going to support. In this work we perform a study of the effect of Cr concentration in a bcc Fe–Cr matrix on formation and binding energies of vacancy clusters up to 5 units. The dependence of the calculated formation and binding energy is investigated with two empirical interatomic potentials specially developed to study radiation damage in Fe–Cr alloys. Results are very similar for both potentials showing an increase of the defect stability with the cluster size and no real dependence on Cr concentration for the binding energy.

Protein isolate and inulin: chemical, rheological and structural basis

Soy protein isolate is typical vegetable protein with health-enhancing activities. Inulin, a prebiotic no digestible carbohydrate, has functional properties. A mashed potato serving of 200 g with added soy protein isolate and inulin concentrations of 15?60 g kg provides from 3 to 12 g of soy protein isolate and/or inulin, respectively. Currently, no information is available about the possible texture-modifying effect of this non-ionizable polar carbohydrate in different soy-based food systems. In this study, the effect of the addition of soy protein isolate and inulin blends at different soy protein isolate: inulin ratios on the degree of inulin polymerization and the rheological and structural properties of fresh mashed and frozen/thawed mashed potatoes were evaluated. The inulin chemical structure remained intact throughout the various treatments, and soy protein isolate did not affect inulin composition being a protein compatible with this fructan. Small-strain rheology showed that both ingredients behaved like soft fillers. In the frozen/thawed mashed potatoes samples,0 addition of 30 : 30 and 15 : 60 blend ratios significantly increased elasticity (G value) compared with 0 : 0 control, consequently reducing the freeze/thaw stability conferred by the cryoprotectants. Inulin crystallites caused a significant strengthening effect on soy protein isolate gel. Micrographs revealed that soy protein isolate supports the inulin structure by building up a second fine-stranded network. Thereby, possibility of using soy protein isolate and inulin in combination with mashed potatoes to provide a highly nutritious and healthy product is promising.

Global Stability Analysis of a Compressible Turbulent Flow around a High-Lift Configuration

The purpose of this work is to analyze a complex high lift configuration for which significant regions of separated flow are present. Current state of the art methods have some diffculty to predict the origin and the progression of this separated flow when increasing the angle of attack. The mechanisms responsible for the maximum lift limit on multi-element wing con?gurations are not clear; this stability analysis could help to understand the physics behind the phenomenon and to find a relation between the flow separation and the instability onset. The methodology presented herein consists in the computation of a steady base flow solution based on a finite volume discretization and a proposal of the solution for a generalized eigenvalue problem corresponding to the perturbed and linearized problem. The eigenvalue problem has been solved with the Arnoldi iterative method, one of the Krylov subspace projection methods. The described methodology was applied to the NACA0012 test case in subsonic and in transonic conditions and, finally, for the first time to the authors knowledge, on an industrial multi-component geometry, such as the A310 airfoil, in order to identify low frequency instabilities related to the separation. One important conclusion is that for all the analyzed geometries, one unstable mode related to flow separation appears for an angle of attack greater than the one correspondent to the maximum lift coe?cient condition. Finally, an adjoint study was carried out in order to evaluate the receptivity and the structural sensitivity of the geometries, giving an indication of the domain region that could be modified resulting in the biggest change of the flowfield.

Global Stability Analysis of Turbulent Transonic Flows on Airfoil geometries

La aparición de inestabilidades en un flujo es un problema importante que puede afectar a algunas aplicaciones aerodinámicas. De hecho existen diferentes tipos de fenómenos no-estacionarios que actualmente son tema de investigación; casos como la separación a altos ángulos de ataque o el buffet transónico son dos ejemplos de cierta relevancia. El análisis de estabilidad global permite identificar la aparición de dichas condiciones inestables, proporcionando información importante sobre la región donde la inestabilidad es dominante y sobre la frecuencia del fenómeno inestable. La metodología empleada es capaz de calcular un flujo base promediado mediante una discretización con volúmenes finitos y posteriormente la solución de un problema de autovalores asociado a la linealización que aparece al perturbar el flujo base. El cálculo numérico se puede dividir en tres pasos: primero se calcula una solución estacionaria para las ecuaciones RANS, luego se extrae la matriz del Jacobiano que representa el problema linealizado y finalmente se deriva y se resuelve el problema de autovalores generalizado mediante el método iterativo de Arnoldi. Como primer caso de validación, la técnica descrita ha sido aplicada a un cilindro circular en condiciones laminares para detectar el principio de las oscilaciones de los vórtices de von Karman, y se han comparado los resultados con experimentos y cálculos anteriores. La parte más importante del estudio se centra en el análisis de flujos compresibles en régimen turbulento. La predicción de la aparición y la progresión de flujo separado a altos ángulos de ataque se han estudiado en el perfil NACA0012 en condiciones tanto subsónicas como supersónicas y en una sección del ala del A310 en condiciones de despegue. Para todas las geometrías analizadas, se ha podido observar que la separación gradual genera la aparición de un modo inestable específico para altos ángulos de ataque siempre mayores que el ángulo asociado al máximo coeficiente de sustentación. Además, se ha estudiado el problema adjunto para obtener información sobre la zona donde una fuerza externa provoca el máximo cambio en el campo fluido. El estudio se ha completado calculando el mapa de sensibilidad estructural y localizando el centro de la inestabilidad. En el presente trabajo de tesis se ha analizado otro importante fenómeno: el buffet transónico. En condiciones transónicas, la interacción entre la onda de choque y la capa límite genera una oscilación de la posición de la onda de choque y, por consiguiente, de las fuerzas aerodinámicas. El conocimiento de las condiciones críticas y su origen puede ayudar a evitar la oscilación causada por estas fuerzas. Las condiciones para las cuales comienza la inestabilidad han sido calculadas y comparadas con trabajos anteriores. Por otra parte, los resultados del correspondiente problema adjunto y el mapa de sensibilidad se han obtenido por primera vez para el buffet, indicando la región del dominio que sera necesario modificar para crear el mayor cambio en las propiedades del campo fluido. Dado el gran consumo de memoria requerido para los casos 3D, se ha realizado un estudio sobre la reducción del domino con la finalidad de reducirlo a la región donde está localizada la inestabilidad. La eficacia de dicha reducción de dominio ha sido evaluada investigando el cambio en la dimensión de la matriz del Jacobiano, no resultando muy eficiente en términos del consumo de memoria. Dado que el buffet es un problema en general tridimensional, el análisis TriGlobal de una geometría 3D podría considerarse el auténtico reto futuro. Como aproximación al problema, un primer estudio se ha realizado empleando una geometría tridimensional extruida del NACA00f2. El cálculo del flujo 3D y, por primera vez en casos tridimensionales compresibles y turbulentos, el análisis de estabilidad TriGlobal, se han llevado a cabo. La comparación de los resultados obtenidos con los resultados del anterior modelo 2D, ha permitido, primero, verificar la exactitud del cálculo 2D realizado anteriormente y también ha proporcionado una estimación del consumo de memoria requerido para el caso 3D. ABSTRACT Flow unsteadiness is an important problem in aerodynamic applications. In fact, there are several types of unsteady phenomena that are still at the cutting edge of research in the field; separation at high angles of attack and transonic buffet are two important examples. Global Stability Analysis can identify the unstable onset conditions, providing important information about the instability location in the domain and the frequency of the unstable phenomenon. The methodology computes a base flow averaged state based on a finite volume discretization and a solution for a generalized eigenvalue problem corresponding to the perturbed linearized equations. The numerical computation is then performed in three steps: first, a steady solution for the RANS equation is computed; second, the Jacobian matrix that represents the linearized problem is obtained; and finally, the generalized eigenvalue problem is derived and solved with an Arnoldi iterative method. As a first validation test, the technique has been applied on a laminar circular cylinder in order to detect the von Karman vortex shedding onset, comparing the results with experiments and with previous calculations. The main part of the study focusses on turbulent and compressible cases. The prediction of the origin and progression of separated flows at high angles of attack has been studied on the NACA0012 airfoil at subsonic and transonic conditions and for the A310 airfoil in take-off configuration. For all the analyzed geometries, it has been found that gradual separation generates the appearance of one specific unstable mode for angles of attack always greater than the ones related to the maximum lift coefficient. In addition, the adjoint problem has been studied to suggest the location of an external force that results in the largest change to the flow field. From the direct and the adjoint analysis the structural sensitivity map has been computed and the core of the instability has been located. The other important phenomenon analyzed in this work is the transonic buffet. In transonic conditions, the interaction between the shock wave and the boundary layer leads to an oscillation of the shock location and, consequently, of the aerodynamic forces. Knowing the critical operational conditions and its origin can be helpful in preventing such fluctuating forces. The instability onset has then been computed and compared with the literature. Moreover, results of the corresponding adjoint problem and a sensitivity map have been provided for the first time for the buffet problem, indicating the region that must be modified to create the biggest change in flow field properties. Because of the large memory consumption required when a 3D case is approached, a domain reduction study has been carried out with the aim of limiting the domain size to the region where the instability is located. The effectiveness of the domain reduction has been evaluated by investigating the change in the Jacobian matrix size, not being very efficient in terms of memory consumption. Since buffet is a three-dimensional problem, TriGlobal stability analysis can be seen as a future challenge. To approximate the problem, a first study has been carried out on an extruded three-dimensional geometry of the NACA0012 airfoil. The 3D flow computation and the TriGlobal stability analysis have been performed for the first time on a compressible and turbulent 3D case. The results have been compared with a 2D model, confirming that the buffet onset evaluated in the 2D case is well detected. Moreover, the computation has given an indication about the memory consumption for a 3D case.

Structural mimicry of a native protein by a minimized binding domain

The affinity between molecules depends both on the nature and presentation of the contacts. Here, we observe coupling of functional and structural elements when a protein binding domain is evolved to a smaller functional mimic. Previously, a 38-residue form of the 59-residue B-domain of protein A, termed Z38, was selected by phage display. Z38 contains 13 mutations and binds IgG only 10-fold weaker than the native B-domain. We present the solution structure of Z38 and show that it adopts a tertiary structure remarkably similar to that observed for the first two helices of B-domain in the B-domain/Fc complex [Deisenhofer, J. (1981) Biochemistry 20, 2361–2370], although it is significantly less stable. Based on this structure, we have improved on Z38 by designing a 34-residue disulfide-bonded variant (Z34C) that has dramatically enhanced stability and binds IgG with 9-fold higher affinity. The improved stability of Z34C led to NMR spectra with much greater chemical shift dispersion, resulting in a more precisely determined structure. Z34C, like Z38, has a structure virtually identical to the equivalent region from native protein A domains. The well-defined hydrophobic core of Z34C reveals key structural features that have evolved in this small, functional domain. Thus, the stabilized two-helix peptide, about half the size and having one-third of the remaining residues altered, accurately mimics both the structure and function of the native domain.

Structural characterization of an engineered tandem repeat contrasts the importance of context and sequence in protein folding

To test a different approach to understanding the relationship between the sequence of part of a protein and its conformation in the overall folded structure, the amino acid sequence corresponding to an α-helix of T4 lysozyme was duplicated in tandem. The presence of such a sequence repeat provides the protein with “choices” during folding. The mutant protein folds with almost wild-type stability, is active, and crystallizes in two different space groups, one isomorphous with wild type and the other with two molecules in the asymmetric unit. The fold of the mutant is essentially the same in all cases, showing that the inserted segment has a well-defined structure. More than half of the inserted residues are themselves helical and extend the helix present in the wild-type protein. Participation of additional duplicated residues in this helix would have required major disruption of the parent structure. The results clearly show that the residues within the duplicated sequence tend to maintain a helical conformation even though the packing interactions with the remainder of the protein are different from those of the original helix. It supports the hypothesis that the structures of individual α-helices are determined predominantly by the nature of the amino acids within the helix, rather than the structural environment provided by the rest of the protein.

Conserved structural features of the synaptic fusion complex: SNARE proteins reclassified as Q- and R-SNAREs

SNARE [soluble NSF (N-ethylmaleimide-sensitive fusion protein) attachment protein receptor] proteins are essential for membrane fusion and are conserved from yeast to humans. Sequence alignments of the most conserved regions were mapped onto the recently solved crystal structure of the heterotrimeric synaptic fusion complex. The association of the four α-helices in the synaptic fusion complex structure produces highly conserved layers of interacting amino acid side chains in the center of the four-helix bundle. Mutations in these layers reduce complex stability and cause defects in membrane traffic even in distantly related SNAREs. When syntaxin-4 is modeled into the synaptic fusion complex as a replacement of syntaxin-1A, no major steric clashes arise and the most variable amino acids localize to the outer surface of the complex. We conclude that the main structural features of the neuronal complex are highly conserved during evolution. On the basis of these features we have reclassified SNARE proteins into Q-SNAREs and R-SNAREs, and we propose that fusion-competent SNARE complexes generally consist of four-helix bundles composed of three Q-SNAREs and one R-SNARE.

Structural features of the kringle domain determine the intracellular degradation of under-γ-carboxylated prothrombin: Studies of chimeric rat/human prothrombin

Vitamin K antagonists such as warfarin inhibit the vitamin K-dependent γ-glutamyl carboxylation during protein processing and block the secretion of under-γ-carboxylated prothrombin (FII) in the rat but not in the human or bovine. Under-γ-carboxylated prothrombin is also secreted from warfarin-treated human (HepG2) cell cultures but is degraded in the endoplasmic reticulum in warfarin-treated rat (H-35) cell cultures. This differential response to warfarin has been shown to be determined by the structural difference in the proteins rather than by the origin of the cell line. When recombinant rat prothrombin (rFII) and human prothrombin (hFII) were expressed in a transformed human kidney cell line (HEK293), secretion of rFII but not hFII was drastically decreased in response to warfarin. To determine the structural signal required for this differential response, chimeric cDNAs with the propeptide/Gla domains, kringle domain, and serine protease domain exchanged between rFII and hFII were generated (FIIRHH and FIIHRR, FIIRRH and FIIHHR, FIIRHR and FIIHRH) and expressed in both warfarin-treated HEK293 cells and HepG2 cells. The presence of the hFII kringle domain changed the stability of rFII to that of hFII, and the rFII kringle domain changed the stability of hFII to that of rFII. The kringle domain therefore is critical in determining the metabolic fate of under-γ-carboxylated prothrombin precursors during processing. Prothrombin contains two kringle structures, and expression of additional rFII/hFII chimeras (FIIHrhH and FIIHhrH, FIIRrhR, and FIIRhrR) was used to determine that the first of the two kringles plays a more important role in the recognition process.

The nitrite reductase from Pseudomonas aeruginosa: Essential role of two active-site histidines in the catalytic and structural properties

Cd1 nitrite reductase catalyzes the conversion of nitrite to NO in denitrifying bacteria. Reduction of the substrate occurs at the d1-heme site, which faces on the distal side some residues thought to be essential for substrate binding and catalysis. We report the results obtained by mutating to Ala the two invariant active site histidines, His-327 and His-369, of the enzyme from Pseudomonas aeruginosa. Both mutants have lost nitrite reductase activity but maintain the ability to reduce O2 to water. Nitrite reductase activity is impaired because of the accumulation of a catalytically inactive form, possibly because the productive displacement of NO from the ferric d1-heme iron is impaired. Moreover, the two distal His play different roles in catalysis; His-369 is absolutely essential for the stability of the Michaelis complex. The structures of both mutants show (i) the new side chain in the active site, (ii) a loss of density of Tyr-10, which slipped away with the N-terminal arm, and (iii) a large topological change in the whole c-heme domain, which is displaced 20 Å from the position occupied in the wild-type enzyme. We conclude that the two invariant His play a crucial role in the activity and the structural organization of cd1 nitrite reductase from P. aeruginosa.

Kinetic stability as a mechanism for protease longevity

The folding of the extracellular serine protease, α-lytic protease (αLP; EC 3.4.21.12) reveals a novel mechanism for stability that appears to lead to a longer functional lifetime for the protease. For αLP, stability is based not on thermodynamics, but on kinetics. Whereas this has required the coevolution of a pro region to facilitate folding, the result has been the optimization of native-state properties independent of their consequences on thermodynamic stability. Structural and mutational data lead to a model for catalysis of folding in which the pro region binds to a conserved β-hairpin in the αLP C-terminal domain, stabilizing the folding transition state and the native state. The pro region is then proteolytically degraded, leaving the active αLP trapped in a metastable conformation. This metastability appears to be a consequence of pressure to evolve properties of the native state, including a large, highly cooperative barrier to unfolding, and extreme rigidity, that reduce susceptibility to proteolytic degradation. In a test of survival under highly proteolytic conditions, homologous mammalian proteases that have not evolved kinetic stability are much more rapidly degraded than αLP. Kinetic stability as a means to longevity is likely to be a mechanism conserved among the majority of extracellular bacterial pro-proteases and may emerge as a general strategy for intracellular eukaryotic proteases subject to harsh conditions as well.

Symmetry, stability, and dynamics of multidomain and multicomponent protein systems

Symmetry is commonly observed in many biological systems. Here we discuss representative examples of the role of symmetry in structural molecular biology. Point group symmetries are observed in many protein oligomers whose three-dimensional atomic structures have been elucidated by x-ray crystallography. Approximate symmetry also occurs in multidomain proteins. Symmetry often confers stability on the molecular system and results in economical usage of basic components to build the macromolecular structure. Symmetry is also associated with cooperativity. Mild perturbation from perfect symmetry may be essential in some systems for dynamic functions.

Structural Requirements of Tom40 for Assembly into Preexisting TOM Complexes of Mitochondria

Tom40 is the major subunit of the translocase of the outer mitochondrial membrane (the TOM complex). To study the assembly pathway of Tom40, we have followed the integration of the protein into the TOM complex in vitro and in vivo using wild-type and altered versions of the Neurospora crassa Tom40 protein. Upon import into isolated mitochondria, Tom40 precursor proteins lacking the first 20 or the first 40 amino acid residues were assembled as the wild-type protein. In contrast, a Tom40 precursor lacking residues 41 to 60, which contains a highly conserved region of the protein, was arrested at an intermediate stage of assembly. We constructed mutant versions of Tom40 affecting this region and transformed the genes into a sheltered heterokaryon containing a tom40 null nucleus. Homokaryotic strains expressing the mutant Tom40 proteins had growth rate defects and were deficient in their ability to form conidia. Analysis of the TOM complex in these strains by blue native gel electrophoresis revealed alterations in electrophoretic mobility and a tendency to lose Tom40 subunits from the complex. Thus, both in vitro and in vivo studies implicate residues 41 to 60 as containing a sequence required for proper assembly/stability of Tom40 into the TOM complex. Finally, we found that TOM complexes in the mitochondrial outer membrane were capable of exchanging subunits in vitro. A model is proposed for the integration of Tom40 subunits into the TOM complex.

Influence of cisplatin intrastrand crosslinking on the conformation, thermal stability, and energetics of a 20-mer DNA duplex.

cis-Diamminedichloroplatinum(II) (cisplatin) is a widely used anticancer drug that binds to and crosslinks DNA. The major DNA adduct of the drug results from coordination of two adjacent guanine bases to platinum to form the intrastrand crosslink cis-[Pt(NH3)2[d(GpG)-N7(1), -N7(2)]] (cis-Pt-GG). In the present study, spectroscopic and calorimetric techniques were employed to characterize the influence of this crosslink on the conformation, thermal stability, and energetics of a site-specifically platinated 20-mer DNA duplex. CD spectroscopic and thermal denaturation data revealed that the crosslink alters the structure of the host duplex, consistent with a shift from a B-like to an A-like conformation; lowers its thermal stability by approximately 9 degrees C; and reduces its thermodynamic stability by 6.3 kcal/mol at 25 degrees C, most of which is enthalpic in origin; but it does not alter the two-state melting behavior exhibited by the parent, unmodified duplex, despite the significant crosslink-induced changes noted above. The energetic consequences of the cis-Pt-GG crosslink are discussed in relation to the structural perturbations it induces in DNA and to how these crosslink-induced perturbations might modulate protein binding.