Organofluorine compounds find various applications ranging from pharmaceuticals to refrigerants, insecticides, high-value fluoropolymers and reagents in catalysis. However, the synthesis of organofluorine compounds depends on toxic chemicals such as hydrogen fluoride, chlorinated hydrocarbons, reactive F2 gas and environmentally persistent long-chain fluorosurfactants. Recently more sustainable, energy-efficient syntheses have been developed using base metal-catalyzed transformations of fluoroalkenes and the formation and functionalization of d6-8 fluorometallacycles. In this thesis, we use manganese complex precursors to prepare the first examples of d4 fluorometallacycles. Work in Chapter 2 describes the synthesis and one-electron reduction of manganese bis(diphosphine)- and tetrakis(phosphite) dibromide complexes, MnBr2(P-P)2 and MnBr2[P(O-i-Pr)3]4 and reactions of the corresponding reduced Mn(I)Br complexes with tetrafluoroethylene (TFE). Products proposed to be d4 perfluorometallacycles, MnBr[-CF2(CF2)2CF2-](P-P) proved to be unstable, reforming TFE upon application of vacuum. In Chapter 3 we show that photolysis of ligated manganese(I) carbonyl bromide complexes, MnBrLn(CO)5-n, in the presence of TFE, chlorotrifluoroethylene (CTFE) or perfluoro(methyl vinyl ether) (PMVE) in tetrahydrofuran affords the Mn-H insertion products, Mn(CF2CFXH)(L2)(CO)3 (X = F, Cl, OCF3) only for L2 = DPPE [1,2-bis(diphenylphosphino)ethane] as well as a solid by-product proposed to be MnBr2Ln. These reactions are accompanied by THF fluoroalkylation products, O[-(CH2)3CH(CF2CFHX)-]. By switching to methyl t-butyl ether solvent, we showed that exhaustive photolysis of MnBr(CO)5 + 3 equiv. of DPPE gave a new product proposed to be the first stable d4 fluorometallacycle, MnBr[-(CF2)4-](CO)(DPPE). Reactions of the fluoroalkenes with zerovalent Mn2(CO)10 also contributed to our understanding of potential reaction pathways to form these Mn-H-derived products. Previous work in the Baker group compared FeH2(dmpe)2 and [FeH(H2)(dmpe)2]+ as catalysts for the dehydrogenation of amine-boranes [dmpe = 1,2-bis(dimethylphosphino)ethane]. In Chapter 4 the catalytic reactivity and selectivity of MnH(H2)(dmpe)2 are compared with those observed using the Fe analogs and the catalyst resting state, Mn(2-BH4)(dmpe)2, is identified. Finally, in Chapter 5 we summarize the findings of this thesis and suggests future directions based on this work.

No description available.

L'alternance de deux groupes de polarités très différentes, CH2 et CF2, permet au poly fluorure de vinylidène (PVDF) d’être un polymère industriellement très intéressant. Cependant, cette spécificité mène aussi à d’importantes inversions du monomère lors de la polymérisation vinylique. Pendant la polymérisation, en complément de la propagation tête-queue, CH2CF2CH2CF2, les monomères inversés conduisent à l’addition en queue-queue, CF2CH2CH2CF2, et tête-tête, CH2CF2CF2CH2. Le taux de transformation de polymère se trouve expérimentalement entre 3 et 7%. Ce pourcentage élevé entraine sans aucun doute la modification de propriétés macroscopiques. En utilisant la dynamique moléculaire, cette thèse a pour but de montrer l'effet de ces défauts sur la température de transition vitreuse (Tg), la dynamique locale et sur la température de fusion (Tm) du PVDF. En phase amorphe, le PVDF avec différents pourcentages de régio-défauts a été étudié : 3.6, 4.1, 9.3 et 23%. Cette étude permet de prédire le comportement de polymères qui ne sont pas synthétisés. Étant donné que les Tg simulées et expérimentales concordent avec précision, les motifs moléculaires qui donnent lieu à l'effet plastifiant de l'inversion de monomères peuvent être envisagés. En plus d'accentuer leur effet de plastifiant, la conclusion significative est que la relaxation de la chaîne peut être révélée en abordant explicitement des mouvements locaux. Car cette procédure ne peut pas être déduite de la connaissance du Tg, nous avons basé notre analyse sur le fait cela : 1) Nous avons démontré que des relations linéaires directes entre Tg et l'énergie d'activation conformationnelle de transition (Ea) extraite à partir d'un graphe d'Arrhenius, existent. Ce diagramme correspond au logarithme naturel des taux de transition entre les états rotameriques contre l'inverse de la température. La pente de cette courbe rapporte directement à cet Ea efficace. Un tel lien a été seulement spéculé dans la littérature. 2) Nous avons calculé des relations d'Arrhenius pour différents genres de torsions le long de la chaîne d'épine dorsale. En conséquence, une barrière d'énergie potentielle, ea, est associée à la rotation d'un lien dans un environnement spécifique. L'addition de ces énergies pesées par le pourcentage de chaque lien le long de l'épine dorsale, donne un ea moyen qui est équivalent à l'ea efficace. À l'aide de cette procédure, nous avons maintenant accès au mouvement local de la chaîne entière. 3) Nous avons vérifié cette procédure pour calculer une valeur pour le Tg du copolymère alternatif du l'éthylène-tétrafluoroéthylène (E-TFE), qui possède les segments qui sont présents le PVDF changé. L'ambiguïté concernant la valeur de la Tg du copolymère E_TFE peut être résolue grâce à cette approche, puisque le PVDF avec 50% de défauts régio-isomériques conduit à l'E_TFE. D'ailleurs, nous avons étudié les temps de relaxation pour la fonction d'autocorrélation de torsion au-dessus d'un large éventail de température. La dynamique locale est alors spécifiquement étudiée. L'équation Vogel-Fulcher-Tammann (VFT) est utilisée pour décrire le processus de relaxation associée aux mouvements coopératifs des segments le long de la chaîne. Nous avons également étudié le possibilité d'utiliser le Kohlrausch-Williams-Watts (KWW), fonction exponentielle étirée, afin de décrire la dépendance temporelle du processus de relaxation, ce travail a été effectué à différentes températures. Les résultats concordent bien avec les données expérimentales. L'objectif principal de cette section est d'étudier conjointement la fréquence des transitions conformationnelles et le temps de relaxation obtenu par la fonction d’autocorrelation de torsion, sur une plage importante de température, afin d’établir un entre les fréquences des transitions conformationnelles et le comportement de type VFT. Nous montrons pour la première fois qu’une relation linéaire peut être établie entre la barrière de transition conformationnelle et l’énergie d’activation effective. Nous montrons pour la première fois qu'une relation linéaire peut être établie entre la barrière de transition conformationnelle, Ea et l'énergie d'activation effective, B, responsables de la dynamique locale. Parmi les cinq phases cristallines que présente le PVDF, les cristaux α et ß présentent des propriétés particulières intéressantes et ont fait l'objet d'une attention significative. Ces deux structures cristallines sont celles que l’on rencontre le plus souvent, la phase α est la plus thermodynamiquement stable le cristal β possède des propriétés ferroélectriques. Toutefois, le comportement lors de la fusion de ces deux phases cristallines n’est pas encore totalement compris. Certains chercheurs pensent que la température de fusion de la phase β est supérieure à la phase alpha . D'autres affirment que le pic endothermique vu sur le thermogramme obtenue par calorimétrie différentielle à balayage (DSC) a été attribué par erreur à la phase β, cela à cause d’une confusion dans les références . À cet égard, le comportement de la Tm des cristaux α et β par rapport à leur épaisseur est obtenu par la dynamique moléculaire. Différents types de nanocristaux composés de chaînes de PVDF, sans ou avec 10% de régio-défauts, ayant des longueurs différentes ont ainsi été simulées dans les phases α et β. On applique l'équation de Gibbs-Thomson (G-T) afin de déterminer l'énergie de surface et l’enthalpie de fusion des nanocristaux. Les valeurs déterminées sont en accord avec les données expérimentales. Nous avons montré que le PVDF en phase β pur a une température de fusion inférieure à celle du PVDF en phase α pur. Cependant, en insérant des défauts à l'intérieur du cristal, la phase α modifiée présente une température de fusion inférieure à celle de la phase β modifiée. / Abstract : Alternating two groups, CH2 and CF2, of very different polarities along the backbone chain of polyvinylidene fluoride (PVDF) leads to very interesting properties, such as ferroelectricity. However, these properties are affected by the presence of regioisomerism defects (monomer inversion) that appear during the synthesis. During the polymerization, in addition to the Head-to-Tail (HT) sequences, CH2CF2CH2CF2, the reversed monomer units lead to formation of Tail-to-Tail (TT), CF2CH2CH2CF2, and Head-to-Head (HH), CH2CF2CF2CH2, links. The rate of this chain alteration experimentally lies between 3 and 7 %. This percentage undoubtedly brings changes in macroscopic properties. The aim of this thesis is to reveal the impact of these defects on the glass transition temperature (Tg), local dynamics and melting temperature (Tm) of PVDF by using Molecular Dynamics (MD) simulation. In amorphous phase, PVDF chains with different percentages of regiodefects were investigated: 0, 3.6, 4.1, 9.3, and 23 %. This study makes it possible to predict the experimental behavior of polymers which have not yet been synthesized. Once Tg is acquired, the relaxation of the chain can be investigated through the calculation of the activation energy (Ea) of the conformational transition. The significant conclusion is that the relaxation of the chain can be revealed by addressing the local motions. More specifically: a) We demonstrate a direct linear relationship between Tg and Ea extracted from an Arrhenius plot. This diagram corresponds to the natural logarithm of transition rates between rotameric states versus the inverse of the temperature. The slope of this curve yields directly Ea. Such a link was only speculated in the literature. b) A significant finding of this work is that the mobility of the chain can be associated with different types of bonds in PVDF with regiodefects. c) Based on the analysis of Ea for the different bond contributions, we proposed a value for the Tg of ethylene-tetrafluoroethylene (E-TFE), an isomeric polymer of PVDF with 50% regiodefects. Experimentally, the available data for the Tg of E-TFE are limited and highly variable. For example, it has been reported as varying from -108 °C to 145 °C. The ambiguity of Tg for this copolymer can be resolved with this approach. Furthermore, we studied the relaxation time associated with the torsional autocorrelation function (TACF) over a wide temperature range. The Vogel-Fulcher-Tammann (VFT) equation was used to describe the temperature dependence of the relaxation time. The Kohlrausch Williams Watts (KWW) stretched exponential function is then applied to fit the time dependence of the relaxation process at various temperatures. The results obtained from this work were found to be in good agreement with the experimental data. A particular interest in this study is the question of how the non-Arrhenius VFT of relaxation process is related to the Arrhenius behavior of conformational jump rates near the glass transition. In both cases, the energies (the conformational transition energy (Ea) and the effective activation energy (B) in VFT equation), were very close to the value of a single torsional barrier. However, in contrast to the relaxation time associated with TACF, the rates of conformational jumps show the activation energy higher than the single barrier value. We have shown that a linear relationship can be established between the conformational transition energy and the effective activation energy. In crystalline PVDF, among the five typical phases, the α and β crystals are of particular interest. The α phase is the most thermodynamically stable form and the β crystal possesses ferroelectric properties. The melting behaviour of these two crystal phases is not so clear. Some researchers believe that the melting temperature of the β phase is higher than that of the α phase. Others have claimed that the higher melting temperature of the peak in Differential Scanning Calorimetry (DSC) has been mistakenly attributed to β phase melting, due to confusion in the referencing of literature sources. In this regard, the melting temperatures of α and β crystals (with and without regiodefects) with respect to their thickness are captured by MD simulation. We then applied the Gibbs-Thomson (G-T) equation to determine the melting temperature, as well as the surface energy and enthalpy of fusion, for α and β nanocrystals. We have shown that pure β phase PVDF has a lower melting temperature than pure α phase PVDF. However, by inserting regiodefects randomly inside the crystal, the α phase with regiodefects shows a lower melting temperature than that of the β phase with regiodefects. We attributed this behaviour to the different structures of the two phases.

O sistema de secreção tipo IV (T4SS) da família de bactérias Xanthomonadaceae transfere efetores (X-Tfes) com a capacidade de matar outras bactérias, conferindo uma vantagem em comunidades bacterianas mistas para colonizar diferentes nichos como o solo ou as superfícies das plantas. Os X-Tfes possuem diferentes domínios putativos com atividades hidrolíticas contra componentes do envelope celular bacteriano do tipo: glicohidrolases, transglicosilases, amidases e lipases. Os X-Tfes por sua atividade biológica inata podem ocasionar dano intracelular para a bactéria que os produz. Para se proteger contra estas atividades, também são produzidas lipoproteínas com função inibitoria (X-Tfis) localizadas no periplasma. Os genes que codificam os X-Tfes e os X-Tfis estão organizados em operons, o que permite gerar os pares efetor/inibidor simultaneamente. Entre os potenciais X-Tfes do fitopatógeno Xanthomonas citri estão Xac1918 e Xac0574. Xac1918 é uma proteína com um domínio da superfamília da lisozima e um domínio conhecido como RTX (Repeats in Toxin) de ligação ao cálcio, enquanto Xac0574 tem um domínio da superfamília da lipase 3. Os seus possíveis inibidores, Xac1917 e Xac0573 respectivamente, apresentam um peptídeo sinal no N-terminal contendo o lipobox representativo das lipoproteínas. As proteínas Xac0574 e Xac0573 são monômeros em solução que formam um complexo estável 1:1, favorecido termodinamicamente (ΔG°= -12 Kcal/mol) com uma constante de dissociação de 2,45 nM, garantindo que a bactéria fique protegida contra os efeitos nocivos de Xac0574 quando é produzida intracelularmente. Xac0574 é uma fosfolipase A1, sem atividade lisofosfolipase, com a capacidade de hidrolisar os três fosfolipídios majoritários que compõem a membrana celular bacteriana, fosfatidilglicerol (PG), cardiolipina e fosfatidiletanolamina (PE), mostrando uma aparente preferência pelo último. A atividade enzimática de Xac0574 explica a forte inibição do crescimento celular em E. coli após da sua indução heteróloga, já que gera uma diminuição de quase 10 vezes da população celular comparada com a cultura não induzida com a mesma construção. Poroutro lado, Xac0573 inibe efetivamente a atividade enzimática de Xac0574 ao formar o complexo, além de não ter atividade fosfolipase nem lisofosfolipase. Foram produzidos cristais da Xac1918 e Xac0573 que difrataram com uma resolução de 3,0 e 2,5 Å, respectivamente. Porém, só foi gerado um modelo de Xac0573. Xac0573 está composta por duas folhas β antiparalelas com uma topologia característica de β sanduíche Com uma pequena hélice e duas voltas. Um alinhamento de homólogos de Xac0573 identificou nas extremidades da proteína as regiões conservadas, constituindo duas possíveis interfaces de interação que podem ser as responsáveis por bloquear o acesso dos fosfolipídios ao sítio catalítico ou impedir os rearranjos estruturais de Xac0574 que são necessários para a sua atividade enzimática. Adicionalmente, a topologia da Xac0573 é semelhante do domínio C2, conhecido em eucariotos como domínio de ligação ao lipídio e ao cálcio, e está envolvido em processos de sinalização de segundos mensageiros lipídicos, proteínas de trafego de membranas e mecanismos de fusão de membranas. Nossos resultados apontam para uma nova função biológica do domínio C2 como um inibidor enzimático intracelular em bactérias. / The type IV secretion system (T4SS) of the bacteria family Xanthomonadaceae transfers effectors (X-Tfes) with that can kill other bacterial cells, conferring an advantage to the bacterial community during colonization of different niches in the soil or on the plant surface. The X-Tfes possess different putative domains with hydrolytic activity against components of the bacterial cellular envelope, including glycohydrolase, transglycolase, amidase and lipase domain. The innate biological activity of X-Tfes can cause intracellular damage. Therefore, the bacteria that produce them also produce lipoproteins with inhibitor function (X-Tfis) located in the periplasm for their protection. The genes that code for X-Tfes and X-Tfis are organized in operons that allow for their simultaneous expression. Among the X-Tfes of the phytopathogen Xanthomonas citri are Xac1918 and Xac0574. Xac1918 is carries a lysozyme superfamily domain, as well as a domain known as RTX (Repeats in Toxic) predict to bind calcium, while, Xac0574 has a domain belonging to the lipase 3 superfamily. Their possible inhibitors, Xac1917 e Xac0573 respectively, carry an N-terminal signal peptide containing a lipobox found in bacterial lipoproteins. The Xac0574 and Xac0573 proteins are both monomers in solution, They can form a stable 1:1 complex, that is thermodynamically favored (ΔG°= -12 Kcal/mol) with a dissociation constant of 2,45 nM. This affinity ensure that the bacterium is protected against the harmful effects of Xac0574 when it is produced intracellularly. We show that Xac0574 is a phospholipase A1, without lisophospholipase activity, and is able to hydrolyze the three most common phospholipids found in the membranes of Gram negative bacteria, namely phosphatidylglycerol (PG), cardiolipin and phosphatidylethanolamine (PE), presenting an apparent preference for PE. The enzymatic activity of Xac0574 explains the strong inhibition of growth of E. coli cells after its heterologous induction: a nearly 10-fold decrease in the cell population is observed when compared to the non-induced culture with the same construct. On the other hand, Xac0573 effectively inhibits the enzymatic activity of Xac0574. Furthermore, Xac0573 does not possess when forming the complex, besides not having phospholipase nor lysophospholipase activity.Crystals of Xac1918 and Xac0573 were produced which diffracted with to resolution of 3.0 and 2.5 Å, respectively. However, we were able to resolve the structure of only Xac0573. Xac0573 is composed of two anti-parallel sheet that form a β-sandwich with three small helices. An alignment to Xac0573 homologs identified conserved regions at the ends of the protein that constitute two possible interfaces of interaction that may be responsible for blocking the access of the phospholipids to the catalytic site or impede the structural rearrangements of Xac0574 that are necessary for its enzymatic activity. Additionally, the topology of Xac0573 is similar to that to C2 domains, known in eukaryotes to bind lipids and calcium and to be involved in signaling processes mediated by lipid second messengers, membrane trafficking and membrane fusion mechanisms. Our results point to a new biological function of the C2 domain as an intracellular enzyme inhibitor in bacteria.

O sistema de secreção tipo IV (T4SS) da família de bactérias Xanthomonadaceae transfere efetores (X-Tfes) com a capacidade de matar outras bactérias, conferindo uma vantagem em comunidades bacterianas mistas para colonizar diferentes nichos como o solo ou as superfícies das plantas. Os X-Tfes possuem diferentes domínios putativos com atividades hidrolíticas contra componentes do envelope celular bacteriano do tipo: glicohidrolases, transglicosilases, amidases e lipases. Os X-Tfes por sua atividade biológica inata podem ocasionar dano intracelular para a bactéria que os produz. Para se proteger contra estas atividades, também são produzidas lipoproteínas com função inibitoria (X-Tfis) localizadas no periplasma. Os genes que codificam os X-Tfes e os X-Tfis estão organizados em operons, o que permite gerar os pares efetor/inibidor simultaneamente. Entre os potenciais X-Tfes do fitopatógeno Xanthomonas citri estão Xac1918 e Xac0574. Xac1918 é uma proteína com um domínio da superfamília da lisozima e um domínio conhecido como RTX (Repeats in Toxin) de ligação ao cálcio, enquanto Xac0574 tem um domínio da superfamília da lipase 3. Os seus possíveis inibidores, Xac1917 e Xac0573 respectivamente, apresentam um peptídeo sinal no N-terminal contendo o lipobox representativo das lipoproteínas. As proteínas Xac0574 e Xac0573 são monômeros em solução que formam um complexo estável 1:1, favorecido termodinamicamente (ΔG°= -12 Kcal/mol) com uma constante de dissociação de 2,45 nM, garantindo que a bactéria fique protegida contra os efeitos nocivos de Xac0574 quando é produzida intracelularmente. Xac0574 é uma fosfolipase A1, sem atividade lisofosfolipase, com a capacidade de hidrolisar os três fosfolipídios majoritários que compõem a membrana celular bacteriana, fosfatidilglicerol (PG), cardiolipina e fosfatidiletanolamina (PE), mostrando uma aparente preferência pelo último. A atividade enzimática de Xac0574 explica a forte inibição do crescimento celular em E. coli após da sua indução heteróloga, já que gera uma diminuição de quase 10 vezes da população celular comparada com a cultura não induzida com a mesma construção. Poroutro lado, Xac0573 inibe efetivamente a atividade enzimática de Xac0574 ao formar o complexo, além de não ter atividade fosfolipase nem lisofosfolipase. Foram produzidos cristais da Xac1918 e Xac0573 que difrataram com uma resolução de 3,0 e 2,5 Å, respectivamente. Porém, só foi gerado um modelo de Xac0573. Xac0573 está composta por duas folhas β antiparalelas com uma topologia característica de β sanduíche Com uma pequena hélice e duas voltas. Um alinhamento de homólogos de Xac0573 identificou nas extremidades da proteína as regiões conservadas, constituindo duas possíveis interfaces de interação que podem ser as responsáveis por bloquear o acesso dos fosfolipídios ao sítio catalítico ou impedir os rearranjos estruturais de Xac0574 que são necessários para a sua atividade enzimática. Adicionalmente, a topologia da Xac0573 é semelhante do domínio C2, conhecido em eucariotos como domínio de ligação ao lipídio e ao cálcio, e está envolvido em processos de sinalização de segundos mensageiros lipídicos, proteínas de trafego de membranas e mecanismos de fusão de membranas. Nossos resultados apontam para uma nova função biológica do domínio C2 como um inibidor enzimático intracelular em bactérias. / The type IV secretion system (T4SS) of the bacteria family Xanthomonadaceae transfers effectors (X-Tfes) with that can kill other bacterial cells, conferring an advantage to the bacterial community during colonization of different niches in the soil or on the plant surface. The X-Tfes possess different putative domains with hydrolytic activity against components of the bacterial cellular envelope, including glycohydrolase, transglycolase, amidase and lipase domain. The innate biological activity of X-Tfes can cause intracellular damage. Therefore, the bacteria that produce them also produce lipoproteins with inhibitor function (X-Tfis) located in the periplasm for their protection. The genes that code for X-Tfes and X-Tfis are organized in operons that allow for their simultaneous expression. Among the X-Tfes of the phytopathogen Xanthomonas citri are Xac1918 and Xac0574. Xac1918 is carries a lysozyme superfamily domain, as well as a domain known as RTX (Repeats in Toxic) predict to bind calcium, while, Xac0574 has a domain belonging to the lipase 3 superfamily. Their possible inhibitors, Xac1917 e Xac0573 respectively, carry an N-terminal signal peptide containing a lipobox found in bacterial lipoproteins. The Xac0574 and Xac0573 proteins are both monomers in solution, They can form a stable 1:1 complex, that is thermodynamically favored (ΔG°= -12 Kcal/mol) with a dissociation constant of 2,45 nM. This affinity ensure that the bacterium is protected against the harmful effects of Xac0574 when it is produced intracellularly. We show that Xac0574 is a phospholipase A1, without lisophospholipase activity, and is able to hydrolyze the three most common phospholipids found in the membranes of Gram negative bacteria, namely phosphatidylglycerol (PG), cardiolipin and phosphatidylethanolamine (PE), presenting an apparent preference for PE. The enzymatic activity of Xac0574 explains the strong inhibition of growth of E. coli cells after its heterologous induction: a nearly 10-fold decrease in the cell population is observed when compared to the non-induced culture with the same construct. On the other hand, Xac0573 effectively inhibits the enzymatic activity of Xac0574. Furthermore, Xac0573 does not possess when forming the complex, besides not having phospholipase nor lysophospholipase activity.Crystals of Xac1918 and Xac0573 were produced which diffracted with to resolution of 3.0 and 2.5 Å, respectively. However, we were able to resolve the structure of only Xac0573. Xac0573 is composed of two anti-parallel sheet that form a β-sandwich with three small helices. An alignment to Xac0573 homologs identified conserved regions at the ends of the protein that constitute two possible interfaces of interaction that may be responsible for blocking the access of the phospholipids to the catalytic site or impede the structural rearrangements of Xac0574 that are necessary for its enzymatic activity. Additionally, the topology of Xac0573 is similar to that to C2 domains, known in eukaryotes to bind lipids and calcium and to be involved in signaling processes mediated by lipid second messengers, membrane trafficking and membrane fusion mechanisms. Our results point to a new biological function of the C2 domain as an intracellular enzyme inhibitor in bacteria.

In the first part of the thesis, protein-ligand interactions were investigated using the chaperone LcrH, from Yersinia as target protein. The structure of a peptide encompassing the amphipathic domain (residue 278-300) of the protein YopD from Yersinia was determined by NMR in 40% TFE. The structure of YopD278-300 is a well defined α-helix with a β-turn at the C-terminus of the helix capping the structure. This turn is crucial for the structure as peptides lacking the residues involved in the turn are unstructured. NMR relaxation indicates that the peptide is not monomeric. This is supported by intermolecular NOEs found from residue Phe280 to Ile288 and Val292 indicative of a multimeric structure with the helical structures oriented in an antiparallel manner with hydrophobic residues forming the oligomer. The interaction with the chaperone LcrH was confirmed by 1H relaxation experiments and induced chemical shift changes in the peptide Protein-ligand interactions were investigated further in the second paper using a different approach. A wide range of substances were used in screening for affinity against the chaperones PapD and FimC from uropathogenic Escherichia coli using 1H relaxation NMR experiments, surface plasmon resonance and 19F NMR. Fluorine NMR proved to be advantageous as compared to proton NMR as it is straight forward to identify binding ligands due to the well resolved 19F NMR spectra. Several compounds were found to interact with PapD and FimC through induced line-broadening and chemical shift changes for the ligands. Data corroborate well with surface plasmon resonance and proton NMR experiments. However, our results indicate the substances used in this study to have poor specificity for PapD and FimC as the induced chemical shift is minor and hardly no competitive binding is observed. Paper III and IV is an investigation of the structural features of the allergenic 2S albumin Ber e 1 from Brazil nut. Ber e 1 is a 2S albumin previously identified as the major allergen of Brazil nut. Recent studies have demonstrated that endogenous Brazil nut lipids are required for an immune response to occur in vivo. The structure was obtained from 3D heteronuclear NMR experiments followed by simulated annealing using the software ARIA. Interestingly, the common fold of the 2S albumin family, described as a right-handed super helix with the core composed of a helix bundle, is not found in Ber e 1. Instead the C-terminal region is participating in the formation of the core between helix 3, 4 and 5. The dynamic properties of Ber e 1 were investigated using 15N relaxation experiments and data was analyzed using the model-free approach. The analysis showed that a few residues in the loop between helix 2 and 3 experience decreased mobility, compared to the rest of the loop. This is consistent with NOE data as long range NOEs were found from the loop to the core region of the protein. The anchoring of this loop is a unique feature of Ber e 1, as it is not found in any other structures of 2S albumins. Chemical shift mapping of Ber e 1 upon the addition of lipid extract from Brazil nut identified 4 regions in the protein where chemical shift perturbations were detected. Interestingly, all four structural clusters align along a cleft in the structure formed by helix 1-3 on one side and helix 4-5 on the other. This cleft is big enough to encompass a lipid molecule. It is therefore tempting to speculate whether this cleft is the lipid binding epitope in Ber e 1.

No description available.