840 resultados para NA TRANSPORTERS
Resumo:
Le motif imidazole, un htrocycle 5 atomes contenant 2 atomes dazote et trois atomes de carbone, prsente des proprits physico-chimiques intressantes qui en font un compos de choix pour plusieurs applications. Parmi ces proprits, la fonctionnalisation simple des deux atomes dazote pour former un sel dimidazolium est trs intressante. Ces sels sont dexcellents prcurseurs de carbnes N-htrocycliques (NHC) et sont couramment utiliss pour synthtiser des ligands en vue dune utilisation en catalyse organomtallique. Dautre part, cette famille de composs possde des proprits anionophores permettant une utilisation en transport anionique. Le prsent travail contient les rsultats de travaux concernant ces deux domaines, soit la catalyse et le transport anionique. Dans un premier temps, les proprits de drivs de limidazole sont exploites pour former un catalyseur de type palladium-NHC qui est utilis pour catalyser la raction de Suzuki-Miyaura en milieu aqueux. Lefficacit de ce catalyseur a t dmontre en utilisant aussi peu que 0,001 mol% pour un rendement quantitatif. Il sagit de la premire occurrence dun processus htrogne et recyclable dans leau, utilisant un catalyseur de type Pd-NHC et qui ne ncessite aucun additif ou co-solvant. Le recyclage a t prouv jusqu 10 cycles sans diminution apparente de lactivit du catalyseur. Dans un second temps, plusieurs sels dimidazolium ont t tests en tant que transporteurs transmembranaires danions chlorures. Les proprits intrinsques des sels utiliss qui en font des transporteurs efficaces ont t lucides. Ainsi, les paramtres qui semblent affecter le plus le transport anionique sont le changement du contre-anion du sel dimidazolium de mme que la propension de ce dernier sauto-assembler via une succession dempilements-. De plus, les proprits du transport ont t lucides, montrant la formation de canaux transmembranaires qui permettent non-seulement la diffusion dions Cl-, mais aussi le transport de protons et dions Ca2+. Lintrt de cette recherche repose dabord dans le traitement de diverses pathologies voyant leur origine dans le dysfonctionnement du transport anionique. Cependant, les proprits bactricides des sels dimidazolium utiliss ont t identifies lors des dernires expriences.
Resumo:
Le nombre important de vhicules sur le rseau routier peut entraner des problmes d'encombrement et de scurit. Les usagers des rseaux routiers qui nous intressent sont les camionneurs qui transportent des marchandises, pouvant rouler avec des vhicules non conformes ou emprunter des routes interdites pour gagner du temps. Le transport de matires dangereuses est rglement et certains lieux, surtout les ponts et les tunnels, leur sont interdits d'accs. Pour aider faire appliquer les lois en vigueur, il existe un systme de contrles routiers compos de structures fixes et de patrouilles mobiles. Le dploiement stratgique de ces ressources de contrle mise sur la connaissance du comportement des camionneurs que nous allons tudier travers l'analyse de leurs choix de routes. Un problme de choix de routes peut se modliser en utilisant la thorie des choix discrets, elle-mme fonde sur la thorie de l'utilit alatoire. Traiter ce type de problme avec cette thorie est complexe. Les modles que nous utiliserons sont tels, que nous serons amens faire face des problmes de corrlation, puisque plusieurs routes partagent probablement des arcs. De plus, puisque nous travaillons sur le rseau routier du Qubec, le choix de routes peut se faire parmi un ensemble de routes dont le nombre est potentiellement infini si on considre celles ayant des boucles. Enfin, l'tude des choix faits par un humain n'est pas triviale. Avec l'aide du modle de choix de routes retenu, nous pourrons calculer une expression de la probabilit qu'une route soit prise par le camionneur. Nous avons abord cette tude du comportement en commenant par un travail de description des donnes collectes. Le questionnaire utilis par les contrleurs permet de collecter des donnes concernant les camionneurs, leurs vhicules et le lieu du contrle. La description des donnes observes est une tape essentielle, car elle permet de prsenter clairement un analyste potentiel ce qui est accessible pour tudier les comportements des camionneurs. Les donnes observes lors d'un contrle constitueront ce que nous appellerons une observation. Avec les attributs du rseau, il sera possible de modliser le rseau routier du Qubec. Une slection de certains attributs permettra de spcifier la fonction d'utilit et par consquent la fonction permettant de calculer les probabilits de choix de routes par un camionneur. Il devient alors possible d'tudier un comportement en se basant sur des observations. Celles provenant du terrain ne nous donnent pas suffisamment d'information actuellement et mme en spcifiant bien un modle, l'estimation des paramtres n'est pas possible. Cette dernire est base sur la mthode du maximum de vraisemblance. Nous avons l'outil, mais il nous manque la matire premire que sont les observations, pour continuer l'tude. L'ide est de poursuivre avec des observations de synthse. Nous ferons des estimations avec des observations compltes puis, pour se rapprocher des conditions relles, nous continuerons avec des observations partielles. Ceci constitue d'ailleurs un dfi majeur. Nous proposons pour ces dernires, de nous servir des rsultats des travaux de (Bierlaire et Frejinger, 2008) en les combinant avec ceux de (Fosgerau, Frejinger et Karlstrm, 2013). Bien qu'elles soient de nature synthtiques, les observations que nous utilisons nous mneront des rsultats tels, que nous serons en mesure de fournir une proposition concrte qui pourrait aider optimiser les dcisions des responsables des contrles routiers. En effet, nous avons russi estimer, sur le rseau rel du Qubec, avec un seuil de signification de 0,05 les valeurs des paramtres d'un modle de choix de routes discrets, mme lorsque les observations sont partielles. Ces rsultats donneront lieu des recommandations sur les changements faire dans le questionnaire permettant de collecter des donnes.
Resumo:
Le prsent travail de recherche se propose danalyser les dispositifs de gouvernance nodale de la scurit locale en France, alors que le paradigme a vu le jour et sest dvelopp dans les pays anglo-saxons fortement dcentraliss. Cest quen France les dispositifs de gouvernance nodale sapparentent bien plus un dialogue entre central et local quentre secteur public et priv. La recherche identifie ainsi les caractristiques de la gouvernance nodale au cur des dispositifs partenariaux de la scurit locale, supports par le Contrat Local de Scurit (CLS), le Conseil Local de Scurit et de Prvention de la Dlinquance (CLSPD) ou encore le Groupe Local de Traitement de la Dlinquance (GLTD). La recherche identifie ainsi les stratgies de dcentrage de ltat et de transfert de la production de scurit vers une diversit dacteurs locaux, dont les maires et les services municipaux. Une diversit de politiques publiques locales de scurit de pertinences diffrentes voit alors le jour. Le premier enseignement de cette recherche est limportance du rle jou par le node super-structurel, que nous appelons super-node et qui regroupe le maire ou llu local la scurit, le responsable de la police dtat, celui de la police municipale et le reprsentant de ltat. Il apparat que dans le dispositif de gouvernance nodale, ce groupe informel gnre la dynamique collective qui permet de regrouper, tant les producteurs que les consommateurs de scurit locale gravitant au sein du rseau local de scurit. La quarantaine dentrevues qualitatives permet galement didentifier que la Justice, productrice de scurit comme peut ltre aussi la scurit prive ou la mdiation sociale, apparat plus distante que ce que pouvait laisser penser ltude des textes rglementaires organisant le partenariat. Les bailleurs sociaux, les transporteurs et lducation nationale apparaissent clairement comme des acteurs importants, mais priphriques de scurit, en intgrant cette famille largie de la scurit locale. Le deuxime enseignement est relatif au fonctionnement mme du dispositif nodal ainsi que du super-node, la recherche permettant didentifier les ressources mutualises par lensemble des nodes. Cela permet galement didentifier les mcanismes de rpartition des tches entre les diffrents acteurs et plus particulirement entre les deux organisations policires dtat et municipale, travaillant autant en comptition, quen complmentarit. Cette recherche explore galement le rle jou par linformation dans le fonctionnement du super-node ainsi que limportance de la confiance dans les relations interpersonnelles des reprsentants des nodes au sein du super-node. Enfin, ltude permet galement de mettre en perspective les limites du dispositif actuel de gouvernance nodale : le dfaut avr doutils performants permettant dinformer convenablement le super-node quant aux phnomnes de violence ainsi que dvaluer lefficience du dispositif. Cela permet galement de questionner lautonomie des dispositifs de gouvernance nodale, la confiance pouvant ouvrir la dviance et la collgialit au dfaut de la traabilit de la responsabilit. La fracture avec la socit civile apparat clairement et ne facilite pas le contrle sur un mode de production de scurit qui se dveloppe en parallle des dispositifs traditionnels de dmocratie locale.
Resumo:
La prsence dEscherichia coli pathognes en levages porcins entraine des retards de croissance et la mortalit. La transmission des E. coli pathognes entre les levages et l'abattoir dun mme rseau de production n'est pas bien dcrite. La dtection des gnes de virulence des E. coli pathognes pourrait permettre didentifier un marqueur de contamination dans le rseau. Lobjectif de cette tude a t didentifier un marqueur de contamination E. coli dans un rseau de production porcine dfini afin de dcrire certains modes de transmission des E. coli pathognes. Pour ce faire, une rgion gographique comprenant 10 fermes dengraissement, un abattoir et un rseau de transport a t slectionne. Trois lots de production conscutifs par ferme ont t suivis pendant 12 mois. Des chantillons environnementaux ont t prlevs lintrieur et lextrieur des fermes (3 visites dlevage), dans la cour de labattoir (2 visites lors de sorties de lot) et sur le camion de transport. La dtection des gnes de virulence (eltB, estA, estB, faeG, stxA, stx2A, eae, cnf, papC, iucD, tsh, fedA) dans les chantillons a t ralise par PCR multiplexe conventionnelle. La distribution temporelle et spatiale des gnes de virulence a permis didentifier le marqueur de contamination ETEC/F4 dfini par la dtection dau moins un gne dentrotoxine ETEC (estB, estA et eltB) en combinaison avec le gne de ladhsine fimbriaire (faeG). La distribution des chantillons positifs ETEC/F4 qualifie la cour de labattoir comme un rservoir de contamination frquent par les transporteurs, vecteurs de contamination entre les levages. Ceci suggre le lien microbiologique entre llevage, les transporteurs et labattoir jouant chacun un rle dans la dissmination des microorganismes pathognes et potentiellement zoonotiques en production porcine.
Resumo:
We aim at mapping out a detailed framework that reveals the proportionate flow of cassava and its products along the value chain (VC). Furthermore, we aim at establishing the role of institutions and the linkages between institutions and other VC actors that influence the cassava VC in Uganda. We use both primary and secondary data obtained from four regions in Uganda. Results show that farmers, processors, transporters, traders, consumers and institutions are the major actors. There are four categories of institutions, viz, government, non-government, community based organisations and international agencies. Roles performed by institutions include: development and enforcement of policies, Research and Development (R&D), capacity building, and creation of market access linkages for cassava and its products. Findings reveal that there is no clear nexus and no coordination among farmers/producers, processors, traders, transporters and consumers. However, institutions are well coordinated and play various roles along the VC to influence the dynamics of actors. Policy-wise it is important to establish strong private-public partnerships to bridge the impaired linkages between the actors (farmers/producers, processors, traders, transporters, and consumers) and institutions. Strong partnerships are envisaged to reduce the associated transaction costs amongst the actors.
Resumo:
Most glyco-engineering approaches used to improve quality of recombinant glycoproteins involve the manipulation of glycosyltransferase and/or glycosidase expression. We investigated whether the over expression of nucleotide sugar transporters, particularly the CMP-sialic acid transporter (CMP-SAT), would be a means to improve the sialylation process in CHO cells. We hypothesized that increasing the expression of the CMP-SAT in the cells would increase the transport of the CMP-sialic acid in the Golgi lumen, hence increasing the intra-lumenal CMP-sialic acid pool, and resulting in a possible increase in sialylation extent of proteins being produced. We report the construction of a CMP-SAT expression vector which was used for transfection into CHO-IFN, a CHO cell line producing human IFN. This resulted in approximately 2 to 5 times increase in total CMP-SAT expression in some of the positive clones as compared to untransfected CHO-IFN, as determined using real-time PCR analysis. This in turn concurred with a 9.6% to 16.3% percent increase in site sialylation. This engineering approach has thus been identified as a novel means of improving sialylation in recombinant glycoprotein therapeutics. This strategy can be utilized feasibly on its own, or in combination with existing sialylation improvement strategies. It is believed that such multi-prong approaches are required to effectively manipulate the complex sialylation process, so as to bring us closer to the goal of producing recombinant glycoproteins of high and consistent sialylation from mammalian cells.
Resumo:
TransRed.com es un servicio de bolsa de carga terrestre mediante el cual los usuarios actualizan la oferta y demanda del servicio en tiempo real (va internet o telfono), donde se consulta la disponibilidad de carga o de camiones de los afiliados a la asistencia remota. Este servicio se divide en dos: TransRed Empresaral (TRE) y TransRed Independiente (TRI) los usuarios de estos servicios sern empresas de transporte de carga terrestre y transportadores independientes. Este servicio actualmente no es prestado por ninguna empresa en Colombia y existe una necesidad latente del mismo, lo cual es una oportunidad que TransRed.com quiere aprovechar innovando en el sector. El tamao del mercado potencial es de 1.849 empresas de transporte de carga terrestre y 47.000 transportadores independientes lo cual podra representar una facturacin total de ms de 55.000 millones a un precio por cuenta con alta aceptacin de mercado de 80.000 mensuales para las TRE y 60.000 mensuales para las TRI. El objetivo de ventas planteado en este proyecto es de 504 cuentas TRE y 2.020 TRI para el tercer ao, que representa el 27% del mercado TRE y el 4% del mercado TRI. Teniendo en cuenta que el nivel de aceptacin del servicio est entre 70% y 80%, se pretende alcanzar el objetivo de ventas con una fuerza comercial distribuida en las cuatro principales zonas de concentracin de nuestro mercado objetivo (Bogot, Antioquia, Valle y Atlantico). Los resultados financieros del proyecto nos demuestran que cumpliendo los objetivos planteados y con la estructura de costos diseada se obtiene un ROI de 54,33% para el tercer ao, un periodo de recuperacin de la inversin de 1 ao y dos meses y un margen de utilidad neta de 24,47% en el tercer ao. Este servicio permitir a sus usuarios optimizar sus operaciones mediante el uso de tecnologa de vanguardia, mejorando la competitividad del sector e integrando los diferentes actores que participan en l.
Resumo:
Colombia siempre ha velado por tener una mejor infraestructura del pas, haciendo que se mantenga preocupado por su posicin competitiva frente a su desarrollo como hub logstico de Latinoamrica. Esto se ve fundamentado a travs de la poltica nacional logstica escrita en el COMPES 3547. Sin embargo, hay un desconocimiento por los empresarios grandes y pequeos del pas acerca de las pretensiones que el gobierno quiere llevar a cabo sobre los distintos sectores econmicos. La simulacin de estructuras como sistemas es de vital importancia para el desarrollo y mejoramiento de cadenas de suministro. La administracin de la cadena como sistema que integra procesos permite producir constantemente, mantener niveles adecuados de inventario y cumplir con los requerimientos del cliente final. Lo anterior teniendo en cuenta que los principales actores de la cadena son proveedores, fabricantes, clientes, detallistas, transportadores y distribuidores, En un entorno en el que la globalizacin constituye quiz el motor ms importante para el desempeo de la cadena de suministros, pues rompe barreras geogrficas. En sntesis la simulacin es un aporte importante para la correcta planeacin y operacin de la cadena de suministros y esto a la vez permite prestar un buen servicio al cliente mientras se reducen costos y tiempos.
Resumo:
Los gliomas malignos representan una de las formas ms agresivas de los tumores del sistema nervioso central (SNC). De acuerdo con la clasificacin de los tumores cerebrales de la Organizacin Mundial de la Salud (OMS), los astrocitomas han sido categorizados en cuatro grados, determinados por la patologa subyacente. Es as como los gliomas malignos (o de alto grado) incluyen el glioma anaplsico (grado III) as como el glioblastoma multiforme (GBM, grado IV),estos ltimos los ms agresivos con el peor pronstico (1). El manejo teraputico de los tumores del SNC se basa en la ciruga, la radioterapia y la quimioterapia, dependiendo de las caractersticas del tumor, el estadio clnico y la edad (2),(3), sin embargo ninguno de los tratamientos estndar es completamente seguro y compatible con una calidad de vida aceptable (3), (4). En general, la quimioterapia es la primera opcin en los tumores diseminados, como el glioblastoma invasivo y el meduloblastoma de alto riesgo o con metstasis mltiple, pero el pronstico en estos pacientes es muy pobre (2),(3). Solamente nuevas terapias dirigidas (2) como las terapias anti-angiognicas (4); o terapias gnicas muestran un beneficio real en grupos limitados de pacientes con defectos moleculares especficos conocidos (4). De este modo, se hace necesario el desarrollo de nuevas terapias farmacolgicas para atacar los tumores cerebrales. Frente a las terapias los gliomas malignos son con frecuencia quimioresistentes, y esta resistencia parece depender de al menos dos mecanismos: en primer lugar, la pobre penetracin de muchas drogas anticncer a travs de la barrera hematoenceflica (BBB: Blood Brain Barrier), la barrera del fluido sangre-cerebroespinal (BCSFB: Blood-cerebrospinal fluid barrier) y la barrera sangre-tumor (BTB: blood-tumor barrier). Dicha resistencia se debe a la interaccin de la droga con varios transportadores o bombas de eflujo de droga ABC (ABC: ATP-binding cassette) que se sobre expresan en las clulas endoteliales o epiteliales de estas barreras. En segundo lugar, estos transportadores de eflujo de drogas ABC propios de las clulas tumorales confieren un fenotipo conocido como resistencia a multidrogas (MDR: multidrug resistance), el cual es caracterstico de varios tumores slidos. Este fenotipo tambin est presente en los tumores del SNC y su papel en gliomas es objeto de investigacin (5). Por consiguiente el suministro de medicamentos a travs de la BBB es uno de los problemas vitales en los tratamientos de terapia dirigida. Estudios recientes han demostrado que algunas molculas pequeas utilizadas en estas terapias son sustratos de la glicoprotena P (Pgp: P-gycoprotein), as como tambin de otras bombas de eflujo como las protenas relacionadas con la resistencia a multidrogas (MRPs: multidrug resistance-related proteins (MRPs) o la protena relacionada con cncer de seno (BCRP: breast-cancer resistance related protein)) que no permiten que las drogas de este tipo alcancen el tumor (1). Un sustrato de Pgp y BCRP es la DOXOrubicina (DOXO), un frmaco utilizado en la terapia anti cncer, el cual es muy eficaz para atacar las clulas del tumor cerebral in vitro, pero con un uso clnico limitado por la poca entrega a travs de la barrera hematoenceflica (BBB) y por la resistencia propia de los tumores. Por otra parte las clulas de BBB y las clulas del tumor cerebral tienen tambin protenas superficiales, como el receptor de la lipoprotena de baja densidad (LDLR), que podra utilizarse como blanco teraputico en BBB y tumores cerebrales. Es asi como la importancia de este estudio se basa en la generacin de estrategias teraputicas que promuevan el paso de las drogas a travs de la barrera hematoencefalica y tumoral, y a su vez, se reconozcan mecanismos celulares que induzcan el incremento en la expresin de los transportadores ABC, de manera que puedan ser utilizados como blancos teraputicos.Este estudio demostr que el uso de una nueva estrategia basada en el Caballo de Troya, donde se combina la droga DOXOrubicina, la cual es introducida dentro de un liposoma, salvaguarda la droga de manera que se evita su reconocimiento por parte de los transportadores ABC tanto de la BBB como de las clulas del tumor. La construccin del liposoma permiti utilizar el receptor LDLR de las clulas asegurando la entrada a travs de la BBB y hacia las clulas tumorales a travs de un proceso de endocitosis. Este mecanismo fue asociado al uso de estatinas o drogas anticolesterol las cuales favorecieron la expresin de LDLR y disminuyeron la actividad de los transportadores ABC por nitracin de los mismos, incrementando la eficiencia de nuestro Caballo de Troya. Por consiguiente demostramos que el uso de una nueva estrategia o formulacin denominada ApolipoDOXO ms el uso de estatinas favorece la administracin de frmacos a travs de la BBB, venciendo la resistencia del tumor y reduciendo los efectos colaterales dosis dependiente de la DOXOrubicina. Adems esta estrategia del "Caballo de Troya", es un nuevo enfoque teraputico que puede ser considerado como una nueva estrategia para aumentar la eficacia de diferentes frmacos en varios tumores cerebrales y garantiza una alta eficiencia incluso en un medio hipxico,caracterstico de las clulas cancerosas, donde la expresin del transportador Pgp se vi aumentada. Teniendo en cuenta la relacin entre algunas vas de sealizacin reconocidas como moduladores de la actividad de Pgp, este estudio presenta no solo la estrategia del Caballo de Troya, sino tambin otra propuesta teraputica relacionada con el uso de Temozolomide ms DOXOrubicina. Esta estrategia demostr que el temozolomide logra penetrar la BBB por que interviene en la via de sealizacin de la Wnt/GSK3/-catenina, la cual modula la expresin del transportador Pgp. Se demostr que el TMZ disminuye la protena y el mRNA de Wnt3 permitiendo plantear la hiptesis de que la droga al disminuir la transcripcin del gen Wnt3 en clulas de BBB, incrementa la activacin de la va fosforilando la -catenina y conduciendo a disminuir la -catenina nuclear y por tanto su unin al promotor del gen mdr1. Con base en los resultados este estudio permiti el reconocimiento de tres mecanismos bsicos relacionados con la expresin de los transportadores ABC y asociados a las estrategias empleadas: el primero fue el uso de las estatinas, el cual condujo a la nitracin de los transportadores disminuyendo su actividad por la via del factor de transcripcin NFB; el segundo a partir del uso del temozolomide, el cual metila el gen de Wnt3 reduciendo la actividad de la via de sealizacin de la la -catenina, disminuyendo la expresin del transportador Pgp. El tercero consisti en la determinacin de la relacin entre el eje RhoA/RhoA quinasa como un modulador de la via (no cannica) GSK3/-catenina. Se demostr que la protena quinasa RhoA promovi la activacin de la protena PTB1, la cual al fosforilar a GSK3 indujo la fosforilacin de la -catenina, lo cual dio lugar a su destruccin por el proteosoma, evitando su unin al promotor del gen mdr1 y por tanto reduciendo su expresin. En conclusin las estrategias propuestas en este trabajo incrementaron la citotoxicidad de las clulas tumorales al aumentar la permeabilidad no solo de la barrera hematoenceflica, sino tambin de la propia barrera tumoral. Igualmente, la estrategia del Caballo de Troya podra ser til para la terapia de otras enfermedades asociadas al sistema nervioso central. Por otra parte estos estudios indican que el reconocimiento de mecanismos asociados a la expresin de los transportadores ABC podra constituir una herramienta clave en el desarrollo de nuevas terapias anticncer.
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Preface. Iron is considered to be a minor element employed, in a variety of forms, by nearly all living organisms. In some cases, it is utilised in large quantities, for instance for the formation of magnetosomes within magnetotactic bacteria or during use of iron as a respiratory donor or acceptor by iron oxidising or reducing bacteria. However, in most cases the role of iron is restricted to its use as a cofactor or prosthetic group assisting the biological activity of many different types of protein. The key metabolic processes that are dependent on iron as a cofactor are numerous; they include respiration, light harvesting, nitrogen fixation, the Krebs cycle, redox stress resistance, amino acid synthesis and oxygen transport. Indeed, it is clear that Life in its current form would be impossible in the absence of iron. One of the main reasons for the reliance of Life upon this metal is the ability of iron to exist in multiple redox states, in particular the relatively stable ferrous (Fe2+) and ferric (Fe3+) forms. The availability of these stable oxidation states allows iron to engage in redox reactions over a wide range of midpoint potentials, depending on the coordination environment, making it an extremely adaptable mediator of electron exchange processes. Iron is also one of the most common elements within the Earths crust (5% abundance) and thus is considered to have been readily available when Life evolved on our early, anaerobic planet. However, as oxygen accumulated (the Great oxidation event) within the atmosphere some 2.4 billion years ago, and as the oceans became less acidic, the iron within primordial oceans was converted from its soluble reduced form to its weakly-soluble oxidised ferric form, which precipitated (~1.8 billion years ago) to form the banded iron formations (BIFs) observed today in Precambrian sedimentary rocks around the world. These BIFs provide a geological record marking a transition point away from the ancient anaerobic world towards modern aerobic Earth. They also indicate a period over which the bio-availability of iron shifted from abundance to limitation, a condition that extends to the modern day. Thus, it is considered likely that the vast majority of extant organisms face the common problem of securing sufficient iron from their environment a problem that Life on Earth has had to cope with for some 2 billion years. This struggle for iron is exemplified by the competition for this metal amongst co-habiting microorganisms who resort to stealing (pirating) each others iron supplies! The reliance of micro-organisms upon iron can be disadvantageous to them, and to our innate immune system it represents a chink in the microbial armour, offering an opportunity that can be exploited to ward off pathogenic invaders. In order to infect body tissues and cause disease, pathogens must secure all their iron from the host. To fight such infections, the host specifically withdraws available iron through the action of various iron depleting processes (e.g. the release of lactoferrin and lipocalin-2) this represents an important strategy in our defence against disease. However, pathogens are frequently able to deploy iron acquisition systems that target host iron sources such as transferrin, lactoferrin and hemoproteins, and thus counteract the iron-withdrawal approaches of the host. Inactivation of such host-targeting iron-uptake systems often attenuates the pathogenicity of the invading microbe, illustrating the importance of the battle for iron in the infection process. The role of iron sequestration systems in facilitating microbial infections has been a major driving force in research aimed at unravelling the complexities of microbial iron transport processes. But also, the intricacy of such systems offers a challenge that stimulates the curiosity. One such challenge is to understand how balanced levels of free iron within the cytosol are achieved in a way that avoids toxicity whilst providing sufficient levels for metabolic purposes this is a requirement that all organisms have to meet. Although the systems involved in achieving this balance can be highly variable amongst different microorganisms, the overall strategy is common. On a coarse level, the homeostatic control of cellular iron is maintained through strict control of the uptake, storage and utilisation of available iron, and is co-ordinated by integrated iron-regulatory networks. However, much yet remains to be discovered concerning the fine details of these different iron regulatory processes. As already indicated, perhaps the most difficult task in maintaining iron homeostasis is simply the procurement of sufficient iron from external sources. The importance of this problem is demonstrated by the plethora of distinct iron transporters often found within a single bacterium, each targeting different forms (complex or redox state) of iron or a different environmental condition. Thus, microbes devote considerable cellular resource to securing iron from their surroundings, reflecting how successful acquisition of iron can be crucial in the competition for survival. The aim of this book is provide the reader with an overview of iron transport processes within a range of microorganisms and to provide an indication of how microbial iron levels are controlled. This aim is promoted through the inclusion of expert reviews on several well studied examples that illustrate the current state of play concerning our comprehension of how iron is translocated into the bacterial (or fungal) cell and how iron homeostasis is controlled within microbes. The first two chapters (1-2) consider the general properties of microbial iron-chelating compounds (known as siderophores), and the mechanisms used by bacteria to acquire haem and utilise it as an iron source. The following twelve chapters (3-14) focus on specific types of microorganism that are of key interest, covering both an array of pathogens for humans, animals and plants (e.g. species of Bordetella, Shigella, , Erwinia, Vibrio, Aeromonas, Francisella, Campylobacter and Staphylococci, and EHEC) as well as a number of prominent non-pathogens (e.g. the rhizobia, E. coli K-12, Bacteroides spp., cyanobacteria, Bacillus spp. and yeasts). The chapters relay the common themes in microbial iron uptake approaches (e.g. the use of siderophores, TonB-dependent transporters, and ABC transport systems), but also highlight many distinctions (such as use of different types iron regulator and the impact of the presence/absence of a cell wall) in the strategies employed. We hope that those both within and outside the field will find this book useful, stimulating and interesting. We intend that it will provide a source for reference that will assist relevant researchers and provide an entry point for those initiating their studies within this subject. Finally, it is important that we acknowledge and thank wholeheartedly the many contributors who have provided the 14 excellent chapters from which this book is composed. Without their considerable efforts, this book, and the understanding that it relays, would not have been possible. Simon C Andrews and Pierre Cornelis
Resumo:
The extensive development of the ruminant forestomach sets apart their N economy from that of nonruminants in a number of respects. Extensive pregastric fermentation alters the profile of protein reaching the small intestine, largely through the transformation of nitrogenous compounds into microbial protein. This process is fueled primarily by carbohydrate fermentation and includes extensive recycling of N between the body and gut lumen pools. Nitrogen recycling occurs via blood and gut lumen exchanges of urea and NH3, as well as endogenous gut and secretory N entry into the gut lumen, and the subsequent digestion and absorption of microbial and endogenous protein. Factors controlling urea transfer to the gut from blood, including the contributions of urea transporters, remain equivocal. Ammonia produced by microbial degradation of urea and dietary and endogenous AA is utilized by microbial fermentation or absorbed and primarily converted to urea. Therefore, microbial growth and carbohydrate fermentation affect the extent of NH3 absorption and urea N recycling and excretion. The extensive recycling of N to the rumen represents an evolutionary advantage of the ruminant in terms of absorbable protein supply during periods of dietary protein deficiency, or asynchronous carbohydrate and protein supply, but incurs a cost of greater N intakes, especially in terms of excess N excretion. Efforts to improve the efficiency of N utilization in ruminants by synchronizing fermentable energy and N availability have generally met with limited success with regards to production responses. In contrast, imposing asynchrony through oscillating dietary protein concentration, or infrequent supplementation, surprisingly has not negatively affected production responses unless the frequency of supplementation is less than once every 3 d. In some cases, oscillation of dietary protein concentration has improved N retention compared with animals fed an equal amount of dietary protein on a daily basis. This may reflect benefits of Orn cycle adaptations and sustained recycling of urea to the gut. The microbial symbiosis of the ruminant is inherently adaptable to asynchronous N and energy supply. Recycling of urea to the gut buffers the effect of irregular dietary N supply such that intuitive benefits of rumen synchrony in terms of the efficiency of N utilization are typically not observed in practice.
Resumo:
One of the largest contributions to biologically available nitrogen comes from the reduction of N-2 to ammonia by rhizobia in symbiosis with legumes. Plants supply dicarboxylic acids as a carbon source to bacteroids, and in return they receive ammonia. However, metabolic exchange must be more complex, because effective N-2 fixation by Rhizobium leguminosarum bv viciae bacteroids requires either one of two broad-specificity amino acid ABC transporters (Aap and Bra). It was proposed that amino acids cycle between plant and bacteroids, but the model was unconstrained because of the broad solute specificity of Aap and Bra. Here, we constrain the specificity of Bra and ectopically express heterologous transporters to demonstrate that branched-chain amino acid (LIV) transport is essential for effective N-2 fixation. This dependence of bacteroids on the plant for LIV is not due to their known down-regulation of glutamate synthesis, because ectopic expression of glutamate dehydrogenase did not rescue effective N-2 fixation. Instead, the effect is specific to LIV and is accompanied by a major reduction in transcription and activity of LIV biosynthetic enzymes. Bacteroids become symbiotic auxotrophs for LIV and depend on the plant for their supply. Bacteroids with aap bra null mutations are reduced in number, smaller, and have a lower DNA content than wild type. Plants control LIV supply to bacteroids, regulating their development and persistence. This makes it a critical control point for regulation of symbiosis. MICROBIOLOGY
Resumo:
Escherichia coli possesses iron transporters specific for either Fe2+ or Fe3+. Although Fe2+ is far more soluble than Fe3+, it rapidly oxidizes aerobically at pH >= 7. Thus, FeoAB, the major Fe2+ transporter of E. coli, operates anaerobically. However, Fe2+ remains stable aerobically under acidic conditions, although a low-pH Fe2+ importer has not been previously identified. Here we show that ycdNOB (efeUOB) specifies the first such transporter. efeUOB is repressed at high pH by CpxAR, and is Fe2+-Fur repressed. EfeU is homologous to the high-affinity iron permease, Ftr1p, of Saccharomyces cerevisiae and other fungi. EfeO is periplasmic with a cupredoxin N-terminal domain; EfeB is also periplasmic and is haem peroxidase-like. All three Efe proteins are required for Efe function. The efeU gene of E. coli K-12 is cryptic due to a frameshift mutation - repair of the single-base-pair deletion generates a functional EfeUOB system. In contrast, the efeUOB operon of the enterohaemorrhagic strain, O157:1147, lacks any frameshift and is functional. A 'wild-type' K-12 strain bearing a functional EfeUOB displays a major growth advantage under aerobic, low-pH, low-iron conditions when a competing metal is provided. Fe-55 transport assays confirm the ferrous iron specificity of EfeUOB.
Resumo:
The number of solute-binding protein-dependent transporters in rhizobia is dramatically increased compared with the majority of other bacteria so far sequenced. This increase may be due to the high affinity of solute-binding proteins for solutes, permitting the acquisition of a broad range of growth-limiting nutrients from soil and the rhizosphere. The transcriptional induction of these transporters was studied by creating a suite of plasmid and integrated fusions to nearly all ATP-binding cassette (ABC) and tripartite ATP-independent periplasmic (TRAP) transporters of Sinorhizobium meliloti. In total, specific inducers were identified for 76 transport systems, amounting to approximate to 47% of the ABC uptake systems and 53% of the TRAP transporters in S. meliloti. Of these transport systems, 64 are previously uncharacterized in Rhizobia and 24 were induced by solutes not known to be transported by ABC- or TRAP-uptake systems in any organism. This study provides a global expression map of one of the largest transporter families (transportome) and an invaluable tool to both understand their solute specificity and the relationships between members of large paralogous families.
Resumo:
Glutamate excitotoxicity is implicated in the aetiology of amyotrophic lateral sclerosis (ALS) with impairment of glutamate transport into astrocytes a possible cause of glutamate-induced injury to motor neurons. It is possible that mutations of Cu/Zn superoxide dismutase (SOD1), responsible for about 20% of familial ALS, down-regulates glutamate transporters via oxidative stress. We transfected primary mouse astrocytes to investigate the effect of the FALS-linked mutant hSOD1(G93A) and wild-type SOD1 (hSOD1(wt)) on the glutamate uptake system. Using western blotting, immunocytochemistry and RT-PCR it was shown that expression of either hSOD1(G93A) or hSOD1(wt) in astrocytes produced down-regulation of the levels of a glutamate transporter GLT-1, without alterations in its mRNA level. hSOD1(G93A) or hSOD1(wt) expression caused a decrease of the monomeric form of GLT-1 without increasing oxidative multimers of GLT-1. The effects were selective to GLT-1, since another glutamate transporter GLAST protein and mRNA levels were not altered. Reflecting the decrease in GLT-1 protein, [H-3]D-aspartate uptake was reduced in cultures expressing hSOD1(G93A) or hSOD1(wt). The hSOD1-induced decline in GLT-1 protein and [H-3]D-aspartate uptake was not blocked by the antioxidant Trolox nor potentiated by antioxidant depletion using catalase and glutathione peroxidase inhibitors. Measurement of 2',7'-dichlorofluorescein (DCF)-induced fluorescence revealed that expression of hSOD1(G93A) or hSOD1(wt) in astrocytes does not lead to detectable increase of intracellular reactive oxygen species. This study suggests that levels of GLT-1 protein in astrocytes are reduced rapidly by overexpression of hSOD1, and is due to a property shared between the wild-type and G93A mutant form, but does not involve the production of intracellular oxidative stress.
