Model-based Self-awareness Patterns for Autonomy

Los sistemas técnicos son cada vez más complejos, incorporan funciones más avanzadas, están más integrados con otros sistemas y trabajan en entornos menos controlados. Todo esto supone unas condiciones más exigentes y con mayor incertidumbre para los sistemas de control, a los que además se demanda un comportamiento más autónomo y fiable. La adaptabilidad de manera autónoma es un reto para tecnologías de control actualmente. El proyecto de investigación ASys propone abordarlo trasladando la responsabilidad de la capacidad de adaptación del sistema de los ingenieros en tiempo de diseño al propio sistema en operación. Esta tesis pretende avanzar en la formulación y materialización técnica de los principios de ASys de cognición y auto-consciencia basadas en modelos y autogestión de los sistemas en tiempo de operación para una autonomía robusta. Para ello el trabajo se ha centrado en la capacidad de auto-conciencia, inspirada en los sistemas biológicos, y se ha explorado la posibilidad de integrarla en la arquitectura de los sistemas de control. Además de la auto-consciencia, se han explorado otros temas relevantes: modelado funcional, modelado de software, tecnología de los patrones, tecnología de componentes, tolerancia a fallos. Se ha analizado el estado de la técnica en los ámbitos pertinentes para las cuestiones de la auto-consciencia y la adaptabilidad en sistemas técnicos: arquitecturas cognitivas, control tolerante a fallos, y arquitecturas software dinámicas y computación autonómica. El marco teórico de ASys existente de sistemas autónomos cognitivos ha sido adaptado para servir de base para este análisis de autoconsciencia y adaptación y para dar sustento conceptual al posterior desarrollo de la solución. La tesis propone una solución general de diseño para la construcción de sistemas autónomos auto-conscientes. La idea central es la integración de un meta-controlador en la arquitectura de control del sistema autónomo, capaz de percibir la estado funcional del sistema de control y, si es necesario, reconfigurarlo en tiempo de operación. Esta solución de metacontrol se ha formalizado en cuatro patrones de diseño: i) el Patrón Metacontrol, que define la integración de un subsistema de metacontrol, responsable de controlar al propio sistema de control a través de la interfaz proporcionada por su plataforma de componentes, ii) el patrón Bucle de Control Epistémico, que define un bucle de control cognitivo basado en el modelos y que se puede aplicar al diseño del metacontrol, iii) el patrón de Reflexión basada en Modelo Profundo propone una solución para construir el modelo ejecutable utilizado por el meta-controlador mediante una transformación de modelo a modelo a partir del modelo de ingeniería del sistema, y, finalmente, iv) el Patrón Metacontrol Funcional, que estructura el meta-controlador en dos bucles, uno para el control de la configuración de los componentes del sistema de control, y otro sobre éste, controlando las funciones que realiza dicha configuración de componentes; de esta manera las consideraciones funcionales y estructurales se desacoplan. La Arquitectura OM y el metamodelo TOMASys son las piezas centrales del marco arquitectónico desarrollado para materializar la solución compuesta de los patrones anteriores. El metamodelo TOMASys ha sido desarrollado para la representación de la estructura y su relación con los requisitos funcionales de cualquier sistema autónomo. La Arquitectura OM es un patrón de referencia para la construcción de una metacontrolador integrando los patrones de diseño propuestos. Este meta-controlador se puede integrar en la arquitectura de cualquier sistema control basado en componentes. El elemento clave de su funcionamiento es un modelo TOMASys del sistema decontrol, que el meta-controlador usa para monitorizarlo y calcular las acciones de reconfiguración necesarias para adaptarlo a las circunstancias en cada momento. Un proceso de ingeniería, complementado con otros recursos, ha sido elaborado para guiar la aplicación del marco arquitectónico OM. Dicho Proceso de Ingeniería OM define la metodología a seguir para construir el subsistema de metacontrol para un sistema autónomo a partir del modelo funcional del mismo. La librería OMJava proporciona una implementación del meta-controlador OM que se puede integrar en el control de cualquier sistema autónomo, independientemente del dominio de la aplicación o de su tecnología de implementación. Para concluir, la solución completa ha sido validada con el desarrollo de un robot móvil autónomo que incorpora un meta-controlador con la Arquitectura OM. Las propiedades de auto-consciencia y adaptación proporcionadas por el meta-controlador han sido validadas en diferentes escenarios de operación del robot, en los que el sistema era capaz de sobreponerse a fallos en el sistema de control mediante reconfiguraciones orquestadas por el metacontrolador. ABSTRACT Technical systems are becoming more complex, they incorporate more advanced functionalities, they are more integrated with other systems and they are deployed in less controlled environments. All this supposes a more demanding and uncertain scenario for control systems, which are also required to be more autonomous and dependable. Autonomous adaptivity is a current challenge for extant control technologies. The ASys research project proposes to address it by moving the responsibility for adaptivity from the engineers at design time to the system at run-time. This thesis has intended to advance in the formulation and technical reification of ASys principles of model-based self-cognition and having systems self-handle at runtime for robust autonomy. For that it has focused on the biologically inspired capability of self-awareness, and explored the possibilities to embed it into the very architecture of control systems. Besides self-awareness, other themes related to the envisioned solution have been explored: functional modeling, software modeling, patterns technology, components technology, fault tolerance. The state of the art in fields relevant for the issues of self-awareness and adaptivity has been analysed: cognitive architectures, fault-tolerant control, and software architectural reflection and autonomic computing. The extant and evolving ASys Theoretical Framework for cognitive autonomous systems has been adapted to provide a basement for this selfhood-centred analysis and to conceptually support the subsequent development of our solution. The thesis proposes a general design solution for building self-aware autonomous systems. Its central idea is the integration of a metacontroller in the control architecture of the autonomous system, capable of perceiving the functional state of the control system and reconfiguring it if necessary at run-time. This metacontrol solution has been formalised into four design patterns: i) the Metacontrol Pattern, which defines the integration of a metacontrol subsystem, controlling the domain control system through an interface provided by its implementation component platform, ii) the Epistemic Control Loop pattern, which defines a modelbased cognitive control loop that can be applied to the design of such a metacontroller, iii) the Deep Model Reflection pattern proposes a solution to produce the online executable model used by the metacontroller by model-to-model transformation from the engineering model, and, finally, iv) the Functional Metacontrol pattern, which proposes to structure the metacontroller in two loops, one for controlling the configuration of components of the controller, and another one on top of the former, controlling the functions being realised by that configuration; this way the functional and structural concerns become decoupled. The OM Architecture and the TOMASys metamodel are the core pieces of the architectural framework developed to reify this patterned solution. The TOMASys metamodel has been developed for representing the structure and its relation to the functional requirements of any autonomous system. The OM architecture is a blueprint for building a metacontroller according to the patterns. This metacontroller can be integrated on top of any component-based control architecture. At the core of its operation lies a TOMASys model of the control system. An engineering process and accompanying assets have been constructed to complete and exploit the architectural framework. The OM Engineering Process defines the process to follow to develop the metacontrol subsystem from the functional model of the controller of the autonomous system. The OMJava library provides a domain and application-independent implementation of an OM Metacontroller than can be used in the implementation phase of OMEP. Finally, the complete solution has been validated in the development of an autonomous mobile robot that incorporates an OM metacontroller. The functional selfawareness and adaptivity properties achieved thanks to the metacontrol system have been validated in different scenarios. In these scenarios the robot was able to overcome failures in the control system thanks to reconfigurations performed by the metacontroller.

Variable structure control with chattering elimination and guaranteed stability for a generalized T-S model

In this paper, a fuzzy based Variable Structure Control (VSC) with guaranteed stability is presented. The main objective is to obtain an improved performance of highly non-linear unstable systems. The main contribution of this work is that, firstly, new functions for chattering reduction and error convergence without sacrificing invariant properties are proposed, which is considered the main drawback of the VSC control. Secondly, the global stability of the controlled system is guaranteed.The well known weighting parameters approach, is used in this paper to optimize local and global approximation and modeling capability of T-S fuzzy model.A one link robot is chosen as a nonlinear unstable system to evaluate the robustness, effectiveness and remarkable performance of optimization approach and the high accuracy obtained in approximating nonlinear systems in comparison with the original T-S model. Simulation results indicate the potential and generality of the algorithm. The application of the proposed FLC-VSC shows that both alleviation of chattering and robust performance are achieved with the proposed FLC-VSC controller. The effectiveness of the proposed controller is proven in front of disturbances and noise effects.

Variable structure control with chattering reduction of a generalized T-S model

In this paper, a fuzzy logic controller (FLC) based variable structure control (VSC) is presented. The main objective is to obtain an improved performance of highly non-linear unstable systems. New functions for chattering reduction and error convergence without sacrificing invariant properties are proposed. The main feature of the proposed method is that the switching function is added as an additional fuzzy variable and will be introduced in the premise part of the fuzzy rules; together with the state variables. In this work, a tuning of the well known weighting parameters approach is proposed to optimize local and global approximation and modelling capability of the Takagi-Sugeno (T-S) fuzzy model to improve the choice of the performance index and minimize it. The main problem encountered is that the T-S identification method can not be applied when the membership functions are overlapped by pairs. This in turn restricts the application of the T-S method because this type of membership function has been widely used in control applications. The approach developed here can be considered as a generalized version of the T-S method. An inverted pendulum mounted on a cart is chosen to evaluate the robustness, effectiveness, accuracy and remarkable performance of the proposed estimation approach in comparison with the original T-S model. Simulation results indicate the potential, simplicity and generality of the estimation method and the robustness of the chattering reduction algorithm. In this paper, we prove that the proposed estimation algorithm converge the very fast, thereby making it very practical to use. The application of the proposed FLC-VSC shows that both alleviation of chattering and robust performance are achieved.

Flood frequency analysis by a bivariate model based on copulas

La adecuada estimación de avenidas de diseño asociadas a altos periodos de retorno es necesaria para el diseño y gestión de estructuras hidráulicas como presas. En la práctica, la estimación de estos cuantiles se realiza normalmente a través de análisis de frecuencia univariados, basados en su mayoría en el estudio de caudales punta. Sin embargo, la naturaleza de las avenidas es multivariada, siendo esencial tener en cuenta características representativas de las avenidas, tales como caudal punta, volumen y duración del hidrograma, con el fin de llevar a cabo un análisis apropiado; especialmente cuando el caudal de entrada se transforma en un caudal de salida diferente durante el proceso de laminación en un embalse o llanura de inundación. Los análisis de frecuencia de avenidas multivariados han sido tradicionalmente llevados a cabo mediante el uso de distribuciones bivariadas estándar con el fin de modelar variables correlacionadas. Sin embargo, su uso conlleva limitaciones como la necesidad de usar el mismo tipo de distribuciones marginales para todas las variables y la existencia de una relación de dependencia lineal entre ellas. Recientemente, el uso de cópulas se ha extendido en hidrología debido a sus beneficios en relación al contexto multivariado, permitiendo superar los inconvenientes de las técnicas tradicionales. Una copula es una función que representa la estructura de dependencia de las variables de estudio, y permite obtener la distribución de frecuencia multivariada de dichas variables mediante sus distribuciones marginales, sin importar el tipo de distribución marginal utilizada. La estimación de periodos de retorno multivariados, y por lo tanto, de cuantiles multivariados, también se facilita debido a la manera en la que las cópulas están formuladas. La presente tesis doctoral busca proporcionar metodologías que mejoren las técnicas tradicionales usadas por profesionales para estimar cuantiles de avenida más adecuados para el diseño y la gestión de presas, así como para la evaluación del riesgo de avenida, mediante análisis de frecuencia de avenidas bivariados basados en cópulas. Las variables consideradas para ello son el caudal punta y el volumen del hidrograma. Con el objetivo de llevar a cabo un estudio completo, la presente investigación abarca: (i) el análisis de frecuencia de avenidas local bivariado centrado en examinar y comparar los periodos de retorno teóricos basados en la probabilidad natural de ocurrencia de una avenida, con el periodo de retorno asociado al riesgo de sobrevertido de la presa bajo análisis, con el fin de proporcionar cuantiles en una estación de aforo determinada; (ii) la extensión del enfoque local al regional, proporcionando un procedimiento completo para llevar a cabo un análisis de frecuencia de avenidas regional bivariado para proporcionar cuantiles en estaciones sin aforar o para mejorar la estimación de dichos cuantiles en estaciones aforadas; (iii) el uso de cópulas para investigar tendencias bivariadas en avenidas debido al aumento de los niveles de urbanización en una cuenca; y (iv) la extensión de series de avenida observadas mediante la combinación de los beneficios de un modelo basado en cópulas y de un modelo hidrometeorológico. Accurate design flood estimates associated with high return periods are necessary to design and manage hydraulic structures such as dams. In practice, the estimate of such quantiles is usually done via univariate flood frequency analyses, mostly based on the study of peak flows. Nevertheless, the nature of floods is multivariate, being essential to consider representative flood characteristics, such as flood peak, hydrograph volume and hydrograph duration to carry out an appropriate analysis; especially when the inflow peak is transformed into a different outflow peak during the routing process in a reservoir or floodplain. Multivariate flood frequency analyses have been traditionally performed by using standard bivariate distributions to model correlated variables, yet they entail some shortcomings such as the need of using the same kind of marginal distribution for all variables and the assumption of a linear dependence relation between them. Recently, the use of copulas has been extended in hydrology because of their benefits regarding dealing with the multivariate context, as they overcome the drawbacks of the traditional approach. A copula is a function that represents the dependence structure of the studied variables, and allows obtaining the multivariate frequency distribution of them by using their marginal distributions, regardless of the kind of marginal distributions considered. The estimate of multivariate return periods, and therefore multivariate quantiles, is also facilitated by the way in which copulas are formulated. The present doctoral thesis seeks to provide methodologies that improve traditional techniques used by practitioners, in order to estimate more appropriate flood quantiles for dam design, dam management and flood risk assessment, through bivariate flood frequency analyses based on the copula approach. The flood variables considered for that goal are peak flow and hydrograph volume. In order to accomplish a complete study, the present research addresses: (i) a bivariate local flood frequency analysis focused on examining and comparing theoretical return periods based on the natural probability of occurrence of a flood, with the return period associated with the risk of dam overtopping, to estimate quantiles at a given gauged site; (ii) the extension of the local to the regional approach, supplying a complete procedure for performing a bivariate regional flood frequency analysis to either estimate quantiles at ungauged sites or improve at-site estimates at gauged sites; (iii) the use of copulas to investigate bivariate flood trends due to increasing urbanisation levels in a catchment; and (iv) the extension of observed flood series by combining the benefits of a copula-based model and a hydro-meteorological model.

A continuum damage model characterization based on the soft tissue mesostructure

Material properties of soft tissues are highly conditioned by the hierarchical structure of this kind of composites. These collagen-based tissues present a complex framework of fibres, fibrils, tropocollagen molecules and amino-acids. As the structural mechanisms that control the degradation of soft tissues are related with the behaviour of its fundamental constituents, the relationship between the molecular and intermolecular properties and the tissue behaviour needs to be studied.

Cuticle Structure in Relation to Chemical Composition: Re-assessing the Prevailing Model

The surface of most aerial plant organs is covered with a cuticle that provides protection against multiple stress factors including dehydration. Interest on the nature of this external layer dates back to the beginning of the 19th century and since then, several studies facilitated a better understanding of cuticular chemical composition and structure. The prevailing undertanding of the cuticle as a lipidic, hydrophobic layer which is independent from the epidermal cell wall underneath stems from the concept developed by Brongniart and von Mohl during the first half of the 19th century. Such early investigations on plant cuticles attempted to link chemical composition and structure with the existing technologies, and have not been directly challenged for decades. Beginning with a historical overview about the development of cuticular studies, this review is aimed at critically assessing the information available on cuticle chemical composition and structure, considering studies performed with cuticles and isolated cuticular chemical components. The concept of the cuticle as a lipid layer independent from the cell wall is subsequently challenged, based on the existing literature, and on new findings pointing toward the cell wall nature of this layer, also providing examples of different leaf cuticle structures. Finally, the need for a re-assessment of the chemical and structural nature of the plant cuticle is highlighted, considering its cell wall nature and variability among organs, species, developmental stages, and biotic and abiotic factors during plant growth.

Design of a generalized tool for the performance assessment under sail based on analytical, numerical and empirical results : a global sailing yacht meta-model

La evaluación de las prestaciones de las embarcaciones a vela ha constituido un objetivo para ingenieros navales y marinos desde los principios de la historia de la navegación. El conocimiento acerca de estas prestaciones, ha crecido desde la identificación de los factores clave relacionados con ellas(eslora, estabilidad, desplazamiento y superficie vélica), a una comprensión más completa de las complejas fuerzas y acoplamientos involucrados en el equilibrio. Junto con este conocimiento, la aparición de los ordenadores ha hecho posible llevar a cabo estas tareas de una forma sistemática. Esto incluye el cálculo detallado de fuerzas, pero también, el uso de estas fuerzas junto con la descripción de una embarcación a vela para la predicción de su comportamiento y, finalmente, sus prestaciones. Esta investigación tiene como objetivo proporcionar una definición global y abierta de un conjunto de modelos y reglas para describir y analizar este comportamiento. Esto se lleva a cabo sin aplicar restricciones en cuanto al tipo de barco o cálculo, sino de una forma generalizada, de modo que sea posible resolver cualquier situación, tanto estacionaria como en el dominio del tiempo. Para ello se comienza con una definición básica de los factores que condicionan el comportamiento de una embarcación a vela. A continuación se proporciona una metodología para gestionar el uso de datos de diferentes orígenes para el cálculo de fuerzas, siempre con el la solución del problema como objetivo. Esta última parte se plasma en un programa de ordenador, PASim, cuyo propósito es evaluar las prestaciones de diferentes ti pos de embarcaciones a vela en un amplio rango de condiciones. Varios ejemplos presentan diferentes usos de PASim con el objetivo de ilustrar algunos de los aspectos discutidos a lo largo de la definición del problema y su solución . Finalmente, se presenta una estructura global de cara a proporcionar una representación virtual de la embarcación real, en la cual, no solo e l comportamiento sino también su manejo, son cercanos a la experiencia de los navegantes en el mundo real. Esta estructura global se propone como el núcleo (un motor de software) de un simulador físico para el que se proporciona una especificación básica. ABSTRACT The assessment of the performance of sailing yachts, and ships in general, has been an objective for naval architects and sailors since the beginning of the history of navigation. The knowledge has grown from identifying the key factors that influence performance(length, stability, displacement and sail area), to a much more complete understanding of the complex forces and couplings involved in the equilibrium. Along with this knowledge, the advent of computers has made it possible to perform the associated tasks in a systematic way. This includes the detailed calculation of forces, but also the use of those forces, along with the description of a sailing yacht, to predict its behavior, and ultimately, its performance. The aim of this investigation is to provide a global and open definition of a set of models and rules to describe and analyze the behavior of a sailing yacht. This is done without applying any restriction to the type of yacht or calculation, but rather in a generalized way, capable of solving any possible situation, whether it is in a steady state or in the time domain. First, the basic definition of the factors that condition the behavior of a sailing yacht is given. Then, a methodology is provided to assist with the use of data from different origins for the calculation of forces, always aiming towards the solution of the problem. This last part is implemented as a computational tool, PASim, intended to assess the performance of different types of sailing yachts in a wide range of conditions. Several examples then present different uses of PASim, as a way to illustrate some of the aspects discussed throughout the definition of the problem and its solution. Finally, a global structure is presented to provide a general virtual representation of the real yacht, in which not only the behavior, but also its handling is close to the experience of the sailors in the real world. This global structure is proposed as the core (a software engine) of a physical yacht simulator, for which a basic specification is provided.

A regularised continuum damage model based on the mesoscopic scale for soft tissue

Material properties of soft fibrous tissues are highly conditioned by the hierarchical structure of this kind of composites. Collagen based tissues present, at decreasing length scales, a complex framework of fibres, fibrils, tropocollagen molecules and amino-acids. Understanding the mechanical behaviour at nano-scale level is critical to accurately incorporate this structural information in phenomenological damage models. In this work we derive a relationship between the mechanical and geometrical properties of the fibril constituents and the soft tissue material parameters at macroscopic scale. A Hodge–Petruska two-dimensional model has been used to describe the fibrils as staggered arrays of tropocollagen molecules. After a mechanical characterisation of each of the fibril components, two fibril failures modes have been defined related with two planes of weakness. A phenomenological continuous damage model with regularised softening was presented along with meso-structurally based definitions for its material parameters. Finally, numerical analysis at fibril, fibre and tissue levels are presented to show the capabilities of the model

A model for the structure of the P domains in the subtilisin-like prohormone convertases

The proprotein convertases are a family of at least seven calcium-dependent endoproteases that process a wide variety of precursor proteins in the secretory pathway. All members of this family possess an N-terminal proregion, a subtilisin-like catalytic module, and an additional downstream well-conserved region of ≈150 amino acid residues, the P domain, which is not found in any other subtilase. The pro and catalytic domains cannot be expressed in the absence of the P domains; their thermodynamic instability may be attributable to the presence of large numbers of negatively charged Glu and Asp side chains in the substrate binding region for recognition of multibasic residue cleavage sites. Based on secondary structure predictions, we here propose that the P domains consist of 8-stranded β-barrels with well-organized inner hydrophobic cores, and therefore are independently folded components of the proprotein convertases. We hypothesize further that the P domains are integrated through strong hydrophobic interactions with the catalytic domains, conferring structural stability and regulating the properties and activity of the convertases. A molecular model of these interdomain interactions is proposed in this report.

Toward optimized carbohydrate-based anticancer vaccines: Epitope clustering, carrier structure, and adjuvant all influence antibody responses to Lewisy conjugates in mice

The feasibility of using carbohydrate-based vaccines for the immunotherapy of cancer is being actively explored at the present time. Although a number of clinical trials have already been conducted with glycoconjugate vaccines, the optimal design and composition of the vaccines has yet to be determined. Among the candidate antigens being examined is Lewisy (Ley), a blood group-related antigen that is overexpressed on the majority of human carcinomas. Using Ley as a model for specificity, we have examined the role of epitope clustering, carrier structure, and adjuvant on the immunogenicity of Ley conjugates in mice. A glycolipopeptide containing a cluster of three contiguous Ley-serine epitopes and the Pam3Cys immunostimulating moiety was found to be superior to a similar construct containing only one Ley-serine epitope in eliciting antitumor cell antibodies. Because only IgM antibodies were produced by this vaccine, the effect on immunogenicity of coupling the glycopeptide to keyhole limpet hemocyanin was examined; although both IgM and IgG antibodies were formed, the antibodies reacted only with the immunizing structure. Reexamination of the clustered Ley-serine Pam3Cys conjugate with the adjuvant QS-21 resulted in the identification of both IgG and IgM antibodies reacting with tumor cells, thus demonstrating the feasibility of an entirely synthetic carbohydrate-based anticancer vaccine in an animal model.

The binding conformation of Taxol in β-tubulin: A model based on electron crystallographic density

The chemotherapeutic drug Taxol is known to interact within a specific site on β-tubulin. Although the general location of the site has been defined by photoaffinity labeling and electron crystallography, the original data were insufficient to make an absolute determination of the bound conformation. We have now correlated the crystallographic density with analysis of Taxol conformations and have found the unique solution to be a T-shaped Taxol structure. This T-shaped or butterfly structure is optimized within the β-tubulin site and exhibits functional similarity to a portion of the B9-B10 loop in the α-tubulin subunit. The model provides structural rationalization for a sizeable body of Taxol structure–activity relationship data, including binding affinity, photoaffinity labeling, and acquired mutation in human cancer cells.

A quantitative model for membrane fusion based on low-energy intermediates

The energetics of a fusion pathway is considered, starting from the contact site where two apposed membranes each locally protrude (as “nipples”) toward each other. The equilibrium distance between the tips of the two nipples is determined by a balance of physical forces: repulsion caused by hydration and attraction generated by fusion proteins. The energy to create the initial stalk, caused by bending of cis monolayer leaflets, is much less when the stalk forms between nipples rather than parallel flat membranes. The stalk cannot, however, expand by bending deformations alone, because this would necessitate the creation of a hydrophobic void of prohibitively high energy. But small movements of the lipids out of the plane of their monolayers allow transformation of the stalk into a modified stalk. This intermediate, not previously considered, is a low-energy structure that can reconfigure into a fusion pore via an additional intermediate, the prepore. The lipids of this latter structure are oriented as in a fusion pore, but the bilayer is locally compressed. All membrane rearrangements occur in a discrete local region without creation of an extended hemifusion diaphragm. Importantly, all steps of the proposed pathway are energetically feasible.

A structure-based catalytic mechanism for the xanthine oxidase family of molybdenum enzymes.

The crystal structure of the xanthine oxidase-related molybdenum-iron protein aldehyde oxido-reductase from the sulfate reducing anaerobic Gram-negative bacterium Desulfovibrio gigas (Mop) was analyzed in its desulfo-, sulfo-, oxidized, reduced, and alcohol-bound forms at 1.8-A resolution. In the sulfo-form the molybdenum molybdopterin cytosine dinucleotide cofactor has a dithiolene-bound fac-[Mo, = O, = S, ---(OH2)] substructure. Bound inhibitory isopropanol in the inner compartment of the substrate binding tunnel is a model for the Michaelis complex of the reaction with aldehydes (H-C = O,-R). The reaction is proposed to proceed by transfer of the molybdenum-bound water molecule as OH- after proton transfer to Glu-869 to the carbonyl carbon of the substrate in concert with hydride transfer to the sulfido group to generate [MoIV, = O, -SH, ---(O-C = O, -R)). Dissociation of the carboxylic acid product may be facilitated by transient binding of Glu-869 to the molybdenum. The metal-bound water is replenished from a chain of internal water molecules. A second alcohol binding site in the spacious outer compartment may cause the strong substrate inhibition observed. This compartment is the putative binding site of large inhibitors of xanthine oxidase.

A cooperative model for receptor recognition and cell adhesion: evidence from the molecular packing in the 1.6-A crystal structure of the pheromone Er-1 from the ciliated protozoan Euplotes raikovi.

The crystal structure of the pheromone Er-1 from the unicellular eukaryotic organism Euplotes raikovi was determined at 1.6 A resolution and refined to a crystallographic R factor of 19.9%. In the tightly packed crystal, two extensive intermolecular helix-helix interactions arrange the Er-1 molecules into layers. Since the putative receptor of the pheromone is a membrane-bound protein, whose extracellular C-terminal domain is identical in amino acid sequence to the soluble pheromone, the interactions found in the crystal may mimic the pheromone-receptor interactions as they occur on a cell surface. Based on this, we propose a model for the interaction between soluble pheromone molecules and their receptors. In this model, strong pheromone-receptor binding emerges as a consequence of the cooperative utilization of several weak interactions. The model offers an explanation for the results of binding studies and may also explain the adhesion between cells that occurs during mating.

Tertiary structure of an amyloid immunoglobulin light chain protein: a proposed model for amyloid fibril formation.

An immunoglobulin light chain protein was isolated from the urine of an individual (BRE) with systemic amyloidosis. Complete amino acid sequence of the variable region of the light chain (VL) protein established it as a kappa I, which when compared with other kappa I amyloid associated proteins had unique residues, including Ile-34, Leu-40, and Tyr-71. To study the tertiary structure, BRE VL was expressed in Escherichia coli by using a PCR product amplified from the patient BRE's bone marrow DNA. The PCR product was ligated into pCZ11, a thermal-inducible replication vector. Recombinant BRE VL was isolated, purified to homogeneity, and crystallized by using ammonium sulfate as the precipitant. Two crystal forms were obtained. In crystal form I the BRE VL kappa domain crystallizes as a dimer with unit cell constants isomorphous to previously published kappa protein structures. Comparison with a nonamyloid VL kappa domain from patient REI, identified significant differences in position of residues in the hypervariable segments plus variations in framework region (FR) segments 40-46 (FR2) and 66-67 (FR3). In addition, positional differences can be seen along the two types of local diads, corresponding to the monomer-monomer and dimer-dimer interfaces. From the packing diagram, a model for the amyloid light chain (AL) fibril is proposed based on a pseudohexagonal spiral structure with a rise of approximately the width of two dimers per 360 degree turn. This spiral structure could be consistent with the dimensions of amyloid fibrils as determined by electron microscopy.