MosA is an enzyme from Sinorhizobium meliloti L5-30, a beneficial soil bacterium. Initial investigation into this enzyme categorized it as a methyltransferase. Further investigation revealed that this was incorrect, and that MosA is actually a dihydrodipicolinate synthase, part of the N-acetylneuraminate lyase superfamily. One of the characteristics of enzyme superfamilies is their low sequence identity, but relatively high structural similarity. The structural investigation reported here confirms the high structural similarity between MosA and other superfamily members. <p>Investigation of MosA was carried out by means of x-ray crystallography. It was believed that detailed structural information may shed light into not only the enzymatic mechanism, but also the inhibition of MosA by lysine, the final product of the enzymatic pathway. Insight into enzyme mechanism and inhibition may ultimately prove useful in herbicide or insecticide development, as other dihydrodipicolinate synthases from harmful fungi, bacteria, or plants, make attractive targets for inhibition. Lysine is an essential amino acid for humans, meaning that there is no endogenous lysine production to block the use of these hypothetical inhibitors. Specific inhibitors based on crystal structures have proven to be effective in the past and hopefully, will continue to be useful in the future. <p>Here we report the structure of MosA, solved to 1.95 Å resolution with lysine 161 forming a Schiff-base adduct with pyruvate. This adduct is consistent with the currently accepted dihydrodipicolinate synthase enzyme mechanism.

MosA is an enzyme from Sinorhizobium meliloti L5-30, a beneficial soil bacterium. Initial investigation into this enzyme categorized it as a methyltransferase. Further investigation revealed that this was incorrect, and that MosA is actually a dihydrodipicolinate synthase, part of the N-acetylneuraminate lyase superfamily. One of the characteristics of enzyme superfamilies is their low sequence identity, but relatively high structural similarity. The structural investigation reported here confirms the high structural similarity between MosA and other superfamily members. <p>Investigation of MosA was carried out by means of x-ray crystallography. It was believed that detailed structural information may shed light into not only the enzymatic mechanism, but also the inhibition of MosA by lysine, the final product of the enzymatic pathway. Insight into enzyme mechanism and inhibition may ultimately prove useful in herbicide or insecticide development, as other dihydrodipicolinate synthases from harmful fungi, bacteria, or plants, make attractive targets for inhibition. Lysine is an essential amino acid for humans, meaning that there is no endogenous lysine production to block the use of these hypothetical inhibitors. Specific inhibitors based on crystal structures have proven to be effective in the past and hopefully, will continue to be useful in the future. <p>Here we report the structure of MosA, solved to 1.95 Å resolution with lysine 161 forming a Schiff-base adduct with pyruvate. This adduct is consistent with the currently accepted dihydrodipicolinate synthase enzyme mechanism.

O uso de veículos aéreos não tripulados (VANTs) tem se tornado cada vez mais comum, principalmente em aplicações de uso civil. No cenário militar, o uso de VANTs tem focado o cumprimento de missões específicas que podem ser divididas em duas grandes categorias: sensoriamento remoto e transporte de material de emprego militar. Este trabalho se concentra na categoria do sensoriamento remoto. O trabalho foca a definição de um modelo e uma arquitetura de referência para o desenvolvimento de sensores inteligentes orientados a missões específicas. O principal objetivo destas missões é a geração de mapas temáticos. Neste trabalho são investigados processos e mecanismos que possibilitem a geração desta categoria de mapas. Neste sentido, o conceito de MOSA (Mission Oriented Sensor Array) é proposto e modelado. Como estudos de caso dos conceitos apresentados são propostos dois sistemas de mapeamento automático de fontes sonoras, um para o caso civil e outro para o caso militar. Essas fontes podem ter origem no ruído gerado por grandes animais (inclusive humanos), por motores de combustão interna de veículos ou por atividade de artilharia (incluindo caçadores). Os MOSAs modelados para esta aplicação são baseados na integração de dados provenientes de um sensor de imageamento termal e uma rede de sensores acústicos em solo. A integração das informações de posicionamento providas pelos sensores utilizados, em uma base cartográfica única, é um dos aspectos importantes tratados neste trabalho. As principais contribuições do trabalho são a proposta de sistemas MOSA, incluindo conceitos, modelos, arquitetura e a implementação de referência representada pelo sistema de mapeamento automático de fontes sonoras. / The use of unmanned aerial vehicles (UAV) has become increasingly common, particularly in civilian applications. In the military scenario, the use of UAVs has focused the accomplishment of specific tasks in two broad categories: remote sensing and transport of military material. This work focuses the remote sensing category. It address the definition of a model and reference architecture for the development of smart sensors oriented to specific tasks. The main objective of these missions is to generate thematic maps. This work investigates processes and mechanisms that enable the automatic generation of thematic maps. In this sense, the concept of MOSA (Mission Oriented Sensor Array) is proposed and modeled. As case studies, we propose two automatic mapping systems for on-the-ground generated sound sources, one for the civilian case and one for the military case. These sounds may come from the noise generated by large animals (including humans), from internal combustion engine vehicles or from artillery activity (including hunters). The MOSAs modeled for this application integrate data from a thermal imaging sensor and an on-the-ground network of acoustic sensors. The fusion of position information, provided by the two sensors, into a single cartographic basis is one of the key aspects addressed in this work. The main contributions are the proposed MOSA systems, including concepts, models and architecture and the reference implementation comprised by the system for automatic mapping of sound sources.

O uso de veículos aéreos não tripulados (VANTs) tem se tornado cada vez mais comum, principalmente em aplicações de uso civil. No cenário militar, o uso de VANTs tem focado o cumprimento de missões específicas que podem ser divididas em duas grandes categorias: sensoriamento remoto e transporte de material de emprego militar. Este trabalho se concentra na categoria do sensoriamento remoto. O trabalho foca a definição de um modelo e uma arquitetura de referência para o desenvolvimento de sensores inteligentes orientados a missões específicas. O principal objetivo destas missões é a geração de mapas temáticos. Neste trabalho são investigados processos e mecanismos que possibilitem a geração desta categoria de mapas. Neste sentido, o conceito de MOSA (Mission Oriented Sensor Array) é proposto e modelado. Como estudos de caso dos conceitos apresentados são propostos dois sistemas de mapeamento automático de fontes sonoras, um para o caso civil e outro para o caso militar. Essas fontes podem ter origem no ruído gerado por grandes animais (inclusive humanos), por motores de combustão interna de veículos ou por atividade de artilharia (incluindo caçadores). Os MOSAs modelados para esta aplicação são baseados na integração de dados provenientes de um sensor de imageamento termal e uma rede de sensores acústicos em solo. A integração das informações de posicionamento providas pelos sensores utilizados, em uma base cartográfica única, é um dos aspectos importantes tratados neste trabalho. As principais contribuições do trabalho são a proposta de sistemas MOSA, incluindo conceitos, modelos, arquitetura e a implementação de referência representada pelo sistema de mapeamento automático de fontes sonoras. / The use of unmanned aerial vehicles (UAV) has become increasingly common, particularly in civilian applications. In the military scenario, the use of UAVs has focused the accomplishment of specific tasks in two broad categories: remote sensing and transport of military material. This work focuses the remote sensing category. It address the definition of a model and reference architecture for the development of smart sensors oriented to specific tasks. The main objective of these missions is to generate thematic maps. This work investigates processes and mechanisms that enable the automatic generation of thematic maps. In this sense, the concept of MOSA (Mission Oriented Sensor Array) is proposed and modeled. As case studies, we propose two automatic mapping systems for on-the-ground generated sound sources, one for the civilian case and one for the military case. These sounds may come from the noise generated by large animals (including humans), from internal combustion engine vehicles or from artillery activity (including hunters). The MOSAs modeled for this application integrate data from a thermal imaging sensor and an on-the-ground network of acoustic sensors. The fusion of position information, provided by the two sensors, into a single cartographic basis is one of the key aspects addressed in this work. The main contributions are the proposed MOSA systems, including concepts, models and architecture and the reference implementation comprised by the system for automatic mapping of sound sources.

MosA is a protein found in <i>Sinorhizobium meliloti</i> L5-30 and has been suggested to be responsible for the biosynthesis of the rhizopine 3-O-methyl-scyllo-inosamine (3-MSI) from scyllo-inosamine (SI). However, we have shown MosA is a dihydrodipicolinate synthase (DHDPS) catalyzing the condensation of pyruvate with aspartate-β-semialdehyde (ASA). Since the DHDPS reaction occurs through a Schiff base aldol-type mechanism it was proposed that MosA could be an O-methyltransferase utilizing 2-oxo-butyrate (2-OB) as a novel methyl donor. This interesting yet unlikely possibility would explain MosA's role in the biosynthesis of 3-MSI without ignoring its similarity to DHDPS. Alternatively, MosA may have two catalytic domains one of which possesses a novel binding motif for S-Adenosyl methionine (SAM) to account for methyltransfer activity. In vitro demonstration of MosAs methyltransferase activity is required to resolve this apparent contradiction.<p>This dissertation describes the chemical synthesis of the rhizopines, investigation into whether MosA has a direct role in rhizopine biosynthesis and the thermodynamic characterization of compounds interacting with MosA as observed by isothermal titration calorimetry. <p>Initial investigation into MosAs methyltransferase activity began with 2-OBs interaction with the enzyme. Inhibition experiments determined 2-OB is a competitive inhibitor with respect to pyruvate of the DHDPS reaction of MosA. Furthermore, protein mass spectrometry of MosA in the presence of 2-OB and sodium borohydride indicated that a Schiff base enzyme intermediate was indeed being formed providing evidence that the proposed mechanism may exist. However, neither of the rhizopines had any effect on the DHDPS activity and HPLC assays determined that no 3-MSI was being produced by MosA in the presence of SI and 2-OB. Furthermore, HPLC assays failed to detect methyl transfer activity by MosA utilizing the SAM as a methyl donor. <p>Isothermal titration calorimetry provided thermodynamic characterization of the pyruvate and 2-OB Schiff base intermediates formed with MosA. In addition, ITC provided insight into the nature and thermodynamics of (S)-lysines inhibition of MosA. ITC failed to detect any interactions between the rhizopines or SAM with MosA. These results indicate that MosA is only a DHDPS and does not catalyze the formation of 3-MSI from SI as hypothesized in the literature.

MosA is a protein found in <i>Sinorhizobium meliloti</i> L5-30 and has been suggested to be responsible for the biosynthesis of the rhizopine 3-O-methyl-scyllo-inosamine (3-MSI) from scyllo-inosamine (SI). However, we have shown MosA is a dihydrodipicolinate synthase (DHDPS) catalyzing the condensation of pyruvate with aspartate-β-semialdehyde (ASA). Since the DHDPS reaction occurs through a Schiff base aldol-type mechanism it was proposed that MosA could be an O-methyltransferase utilizing 2-oxo-butyrate (2-OB) as a novel methyl donor. This interesting yet unlikely possibility would explain MosA's role in the biosynthesis of 3-MSI without ignoring its similarity to DHDPS. Alternatively, MosA may have two catalytic domains one of which possesses a novel binding motif for S-Adenosyl methionine (SAM) to account for methyltransfer activity. In vitro demonstration of MosAs methyltransferase activity is required to resolve this apparent contradiction.<p>This dissertation describes the chemical synthesis of the rhizopines, investigation into whether MosA has a direct role in rhizopine biosynthesis and the thermodynamic characterization of compounds interacting with MosA as observed by isothermal titration calorimetry. <p>Initial investigation into MosAs methyltransferase activity began with 2-OBs interaction with the enzyme. Inhibition experiments determined 2-OB is a competitive inhibitor with respect to pyruvate of the DHDPS reaction of MosA. Furthermore, protein mass spectrometry of MosA in the presence of 2-OB and sodium borohydride indicated that a Schiff base enzyme intermediate was indeed being formed providing evidence that the proposed mechanism may exist. However, neither of the rhizopines had any effect on the DHDPS activity and HPLC assays determined that no 3-MSI was being produced by MosA in the presence of SI and 2-OB. Furthermore, HPLC assays failed to detect methyl transfer activity by MosA utilizing the SAM as a methyl donor. <p>Isothermal titration calorimetry provided thermodynamic characterization of the pyruvate and 2-OB Schiff base intermediates formed with MosA. In addition, ITC provided insight into the nature and thermodynamics of (S)-lysines inhibition of MosA. ITC failed to detect any interactions between the rhizopines or SAM with MosA. These results indicate that MosA is only a DHDPS and does not catalyze the formation of 3-MSI from SI as hypothesized in the literature.

Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal.

EU:s institutioner bereder för närvarande en förordning innehållandes internationellt privat- och processrättsliga regler i mål om makars förmögenhetsförhållanden. Det senast presenterade textförslaget (kompromissförslaget) lades fram av Italien den 10 november 2014.   Uppsatsen har två syften. För det första syftar den till att synliggöra vissa av de gränsdragnings- och tillämpningsproblem som skulle uppstå om kompromissförslaget skulle antas i dess nuvarande utformning. För det andra syftar uppsatsen till att erbjuda lösningar på de problem som identifieras. Uppsatsens fokus ligger på följande aspekter av förslaget:   Materiellt tillämpningsområde. Domsrättsreglering i mål med anledning av en makes död. Lagvalsreglering. Förbehållen för internationellt tvingande regler och ordre public.   För det första kan förordningens gränsdragning gentemot inomstatliga tvister och underhålls-frågor vålla problem. För att minska dessa problem bör det i en artikel anges att förordningen endast är tillämplig på tvister som har internationell anknytning och att en tillräcklig internationell anknytning föreligger om en av makarna innehar tillgångar som är belägna utomlands. För att avgränsa förordningens tillämpningsområde gentemot underhållsfrågor bör nationella domstolar följa den metod som anvisades i van den Boogard. Denna metod är bland annat adekvat för att kvalificera avtal mahr (islamisk brudpenning). Det är däremot inte möjligt att använda metoden för att kvalificera en dom i vilken 12 kap. 1 § äktenskapsbalken (skevdelningsregeln) har tillämpats. Enligt min mening bör i så fall den nationella domstolen göra en helhetsbedömning utifrån svaren på följande frågeställningar:   Vilket är originaldomens huvudsakliga syfte? I vilken utsträckning beaktas makarnas behov och förmåga i originaldomen?   För det andra är domsrättsreglering av mål med anledning av makes död problematiska. Art. 3 i kompromissförslaget hänvisar nämligen till arvsförordningen, vilket i praktiken kan leda till att den avlidnes barn gynnas på bekostnad av den efterlevande maken. Enligt min mening bör denna problematik minskas genom att den efterlevande maken ges en exklusiv rätt att avgöra om lagvalsreglerna i arvsförordningen eller i art. 5 kompromissförslaget ska tillämpas i ett mål om makarnas förmögenhetsförhållanden.   För det tredje kan kompromissförslagets lagvalsregler leda till problematik. Eftersom deras utformning skiljer från lagvalsreglerna i arvsförordningen och 2007 års Haagprotokoll finns det en risk att en make över- eller underkompenseras ekonomiskt. Risken kan minskas genom att makarna ingår ett lagvalsavtal eller genom att en domare tillämpar en nationell jämkningsregel eller en EU-rättslig flyktklausul (t.ex. art. 5 i 2007 års Haagprotokoll). För att ytterligare minska risken för att över- och underkompensation bör en flyktklausul införas i förslaget till förordning om makars förmögenhetsförhållanden.    En viss lagvalsregel är särskilt problematisk, nämligen art. 20a.3 i kompromissförslaget (hemvistbytesregeln). Artikeln har en bristande flexibilitet och bör därför förändras på följande vis:   Rekvisitet ”undantagsvis” bör avlägsnas. Istället bör den i punkt 2 föreslagna tidsgränsen spegla den restriktivitet som önskas. Istället för att lagen i makarnas nya hemvistland ska bli tillämplig efter en ”avsevärt mycket längre tid” bör en fast tidsgräns införas. Det bör förtydligas vad som avses med kravet på att makarna ska ha åberopat lagen i den andra staten för att ordna eller planera sina förmögenhetsförhållanden.   Enligt min mening är det lämpligt att hämta vägledning till hur tidsgränsen (se punkt 2) bör utformas i 4 § andra stycket lagen (1990:272) om internationella frågor rörande makars och sambors förmögenhetsförhållanden.   För det fjärde finns problematik med anknytning till kompromissförslagets förbehåll för internationellt tvingande regler och ordre public. Av resonemanget framgår att lagstiftaren måste föra ett detaljerat resonemang (a detailed assessment) för att 7 kap. 4–9 §§, 11 kap. 8 § och 12 kap. 1–3 §§ äktenskapsbalken (1987:230) ska kunna betecknas som internationellt tvingande även efter att förordningen har trätt i kraft. Slutligen dras slutsatsen att det är önskvärt att ramarna är snäva när det gäller en nationell domstols möjligheter att underlåta att tillämpa utländsk rätt som ger rättskraft åt avtal om mahr.