<p> AroA catalyzes a carboxyvinyl transfer reaction, forming enolpyruvyl shikimate 3-phosphate (EPSP) from shikimate 3-phosphate (S3P) and phosphoenolpyruvate (PEP). Upon extended incubation, it forms EPSP ketal by intramolecular nucleophilic attack of O4H on C2' of the enolpyruvyl group. EPSP ketal was previously proposed to form by non-enzymatic breakdown of the tetrahedral intermediate (THI) which had dissociated from AroA. In this study, EPSP ketal formed in the presence of excess AroA, which demonstrated that it was formed in the active site. This eliminated non-enzymatic THI breakdown as its source, and demonstrated that AroA forms either a discrete EPSP cationic intermediate, or cl transition state with high cationic character. The pH dependence of non-enzymatic EPSP hydrolysis was examined in order to understand the intrinsic reactivity of the enolpyruvyl group. Acid catalysis accelerated EPSP hydrolysis> 10^8-fold. These results provide evidence for enolpyruvyl activation through protonation at C3', forming an unstable cationic intermediate, or a highly cation-like transition state. The incorporation of 2H into EPSP from solvent 2H20 during AreA-catalyzed hydrolysis was much slower than the hydrolysis rate, in the absence of inorganic phosphate in the reaction. This demonstrated that KIEs on AroA-catalyzed EPSP hydrolysis, when they are measured in the future, will reflect protonation of EPSP. A method was developed for KIE measurements on acid-catalyzed EPSP hydrolysis, which showed good reproducibility and no buffer dependence. Further experiments need to be completed on the acid-catalyzed KIEs and enzyme-catalyzed KIEs, followed by transition state analysis. This will precisely define the transition state structure of the enzyme-catalyzed EPSP hydrolysis reaction, and provide a good starting point for designing AroA inhibitors.</p> / Thesis / Master of Science (MSc)

<p dir="ltr">While electrospray ionization facilitates the mass determination of smaller analytes, ESI of macromolecular native protein complexes is complicated by narrow charge state distributions and overlapping charge states, hindering mass analysis. This problem is further exacerbated with heterogeneous protein mixtures that yield ions of similar <i>m/z</i> values. Charge-reduction of a selected precursor population via ion/ion reaction provides further <i>m/z</i> separation and utilizes an extended mass range, allowing for mass determination. All experiments were performed on a TripleTOF 5600 quadrupole TOF mass spectrometer (SCIEX), modified for ion/ion reactions. Alternatively pulsed nano-electrospray ionization allowed for sequential injection of reagent and analyte ions. Selected cations were reacted with different anions, and charge-reduced product ions were mass analyzed on a TOF with modifications and tuning for an extended <i>m/z</i> range of 200,000+. Charge reduction via proton transfer reaction (PTR) involves a perfluorinated anion reacting with a multiply charged cation and results in the loss of one proton removal at a time. Through multiple iterations of PTR, the overlapping charge states of protein ions (from an unstained protein standard mixture consisting of 12 recombinant proteins of masses10kDa-200kDa) within the <i>m/z</i> 6500-8500 and <i>m/z</i> 4000-5000 ranges, generated under native conditions by ESI, transforms to a product spectrum with single-digit charge states, thereby deconvoluting the precursor “blob”.</p>

The nuclear structme of 17Ne and 13O has been studied by the 20Ne(3He,6He)17Ne and 16O(3He,6He)13O reactions at 70 MeV and 80 MeV, respectively. Fifteen levels were identified, and angular distributions have been measured for nine of these levels in 17Ne, while for 13O eighteen levels were identified, but angular distributions were obtained for only ten levels. The observed transferred angular momentum dependence of these angular distributions allowed spin-parity assignments. The T= 3/2 quartet analog states in mass A=17 have been completed for six levels. The results of the isobaric multiplet mass equation analysis show a slight linear dependence of the b and c coefficients on the excitation energy. It was found that the coefficients for the positive parity states do not follow the systematics of the negative parity states. The absolute values of the b and c coefficients are larger for the positive parity states. An analysis in terms of Coulomb energy displacement indicates a possible configuration mixing or core polarization effect in these states. The d coefficient also has a large deviation from zero, only for the positive parity states indicating a possible expansion of the radial wavefunction or some isospin symmetry breaking effects. Further detailed theoretical interpretation of these effects may bring valuable information about the configuration and structure of these states. The leveis in 13O were measured with good energy resolution, and thus, it was possible to identify the first excited state unambiguously. However, if one identifies this state as the analog of the known first excited state in the mirror nucleus 13B, this leads to one of the largest level shifts known in literature. / The nuclear structme of 17Ne and 13O has been studied by the 20Ne(3He,6He)17Ne and 16O(3He,6He)13O reactions at 70 MeV and 80 MeV, respectively. Fifteen levels were identified, and angular distributions have been measured for nine of these levels in 17Ne, while for 13O eighteen levels were identified, but angular distributions were obtained for only ten levels. The observed transferred angular momentum dependence of these angular distributions allowed spin-parity assignments. The T= 3/2 quartet analog states in mass A=17 have been completed for six levels. The results of the isobaric multiplet mass equation analysis show a slight linear dependence of the b and c coefficients on the excitation energy. It was found that the coefficients for the positive parity states do not follow the systematics of the negative parity states. The absolute values of the b and c coefficients are larger for the positive parity states. An analysis in terms of Coulomb energy displacement indicates a possible configuration mixing or core polarization effect in these states. The d coefficient also has a large deviation from zero, only for the positive parity states indicating a possible expansion of the radial wavefunction or some isospin symmetry breaking effects. Further detailed theoretical interpretation of these effects may bring valuable information about the configuration and structure of these states. The leveis in 13O were measured with good energy resolution, and thus, it was possible to identify the first excited state unambiguously. However, if one identifies this state as the analog of the known first excited state in the mirror nucleus 13B, this leads to one of the largest level shifts known in literature.

The work described in this thesis is focused on the development of new bidentate iridium complexes and their applications in the asymmetric reduction of olefins, ketones and imines. Three new types of iridium complexes were synthesized, which included pyridine derived chiral N,P-iridium complexes, achiral NHC complexes and chiral NHC-phosphine complexes. A study of their catalytic applications demonstrated a high efficiency of the N,P-iridium complexes for asymmetric hydrogenation of olefins, with good enantioselectivity. The carbene complexes were found to be very efficient hydrogen transfer mediators capable of abstracting hydrogen from alcohols and subsequently transfer it to other unsaturated bonds. This hydrogen transferring property of the carbene complexes was used in the development of C–C and C–N bond formation reactions via the hydrogen borrowing process. The complexes displayed high catalytic reactivity using 0.5–1.0 mol% of the catalyst and mild reaction conditions. Finally chiral carbene complexes were found to be activated by hydrogen gas. Their corresponding iridium hydride species were able to reduce ketones and imines with high efficiency and enantioselectivity without any additives, base or acid. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 5: Submitted. Paper 6: Manuscript.</p><p> </p>

As part of an ongoing program to study fundamental symmetries in nuclear physics, a thorough investigation into shell-model isospin-symmetry-breaking (ISB) calculation theory has been conducted using direct reactions to observe detailed nuclear-structure information. The work presented in this Thesis focuses on the 62Ga superallowed beta-decay system, and consists of two primary experiments; 1) A 64Zn(d,t)63Zn single-neutron transfer reaction, aimed at observing spectroscopic strengths to help guide calculation model-space truncations for the beta-decay wave function radial-overlap component of ISB, and 2) A two-neutron 64Zn(p,t)62Zn transfer to search for excited 0+ states in the daughter nucleus of 62Ga. The experiments were performed at the Maier-Leibnitz-Laboratory, on the joint campuses of the Ludwig-Maximilians Universitat and the Technische Universitat Munchen, in Garching, Germany. In total, 162 states in 63Zn were populated from the 64Zn(d,t) reaction, up to an excitation energy of 4.8 MeV, including the observation of 125 new levels, and unique spin/parity assignments for 92 states. As a result, this work provides the most complete picture for low-spin states in 63Zn to date. A comparison of the extracted S values to the predicted shell-model spectroscopic factors shows an overall over-prediction of strength for the 2p3/2 orbital, and a large disagreement for the 1f7/2 orbital above ~3.5 MeV. No significant 1g9/2 strength was observed, leading to the conclusion that the importance of the 1g9/2 orbital for ISB is small. Additionally, 67 states were observed in 62Zn using the two-neutron pickup mechanism, including the observation of five 0+ states. More than 99% of the total 0+ (p,t) cross-section is observed in the ground-state reaction channel, implying a nearly maximal overlap of the wave functions with the two-nucleon transfer operator. The dominance of the ground-state-to-ground-state (p,t) cross section is strikingly similar to the dominance of the superallowed Fermi beta-decay between isobaric-analogue 0+ states. This suggests that the population of excited 0+ states in the (p,t) reaction may reflect the population in the Fermi decay process, and can be used to guide future experimental and theoretical work. Further discussion of these results as they relate to the ISB correction calculations, and the implications for future theoretical work are presented in this Thesis. / This work was supported in part by the Natural Sciences and Engineering Research Council of Canada, the Ontario Ministry of Economic Development and Innovation, the DFG Cluster of Excellence `Origin and Structure of the Universe', and NSF grant PHY-1068217.

Histone deacetylase (HDAC) inhibition is an emerging novel therapeutic strategy in cancer therapy. HDAC inhibitors (HDACi) have shown ability to block angiogenesis and cell cycling, as well as initiate differentiation and apoptosis. In fact, suberoylanilide hydroxamic acid (SAHA) is the first in the class of HDACi approved by the FDA for the treatment of cutaneous T cell lymphoma. On the other hand, there is a sustained interest in the use of gold nanoparticles (AuNPs) for various cancer diagnostic and therapeutic applications - bioimaging, drug delivery, and binary therapy techniques such as photodynamic and photothermal therapies. This interest in AuNPs is facilitated by favorable attributes such as ease of fabrication, bioconjugation and biocompatibility, and unique optical and electronic properties. However, HDACi- and AuNPs- based antitumor agents are plagued with problems common to all chemotherapeutic agents such as lack of selectivity, which often results in systemic toxicity. Therefore, availability of a methodology to selectively deliver AuNPs and HDACi to cancer cells will significantly improve their therapeutic indices and lead to the identification of novel agents for use in diagnostic imaging and targeted cancer therapy applications.

L’intérêt des peptides comme des médicaments potentiels est en constante augmentation. Des stratégies ont été développées pour améliorer la sélectivité, l’activité, et la stabilité des peptides vis-à-vis de la protéolyse. Dans ce mémoire de thèse, deux nouvelles modifications de peptides sont proposées.Dans le premier chapitre, nous présentons une nouvelle application de la réaction de transfert O-N acylique pour la synthèse de peptides agrafés (ou ‘stapled peptides’). L’introduction d’une ‘agrafe’ dans un peptide est un moyen de stabiliser une structure secondaire hélicoïdale en établissant un pont entre les résidus appropriés des chaînes latérales. Dans notre cas, l'agrafe est formée par un O-acyl isodipeptide. La liaison ester peut être convertie en liaison amide par un transfert O-N acylique. Cette stratégie permet une amélioration de la solubilité d’un peptide hydrophobe agrafé, avant son réarrangement à pH neutre.Dans le dernier chapitre, nous avons développé une méthodologie nouvelle pour la dimérisation de peptides non protégés. Cette méthode repose sur la formation de liaisons siloxane entre des peptides hybrides portant chacun un groupement dimethylchlorosilane. Nous avons ainsi dimérisé une séquence dérivée de la protéine p53, impliquée dans l’apoptose. A titre de comparaison, cette même séquence a été dimérisée en utilisant la cycloaddition d’Huisgen entre deux peptides modifiés possédant un azoture ou un alcyne en position N-terminale. Enfin, plusieurs dimères de la séquence du GHRP-6 (growth hormone releasing peptide) ont été synthétisés, avec des bras dimethylhydroxysilane placés à différentes positions. L’homodimérisation a été effectuée dans l'eau à pH neutre. / The interest in peptides as potential drug candidates was revived and is increasing constantly. Strategies have been developed to improve their selectivity and activity, and their stability toward proteolysis. In this thesis, two new peptide modifications are proposed.In the first chapter, we present a new application of the O-N acyl transfer reaction for the synthesis of stapled peptides. Peptide ‘stapling’ is a way of stabilizing secondary helical structure by establishment of a bridge between the side chains of suitable residues. In our case, the staple is formed by an O-acyl isopeptide which can be converted into amide bond by acyl migration. This strategy allows an improved solubility of the stapled hydrophobic peptide prior to rearrangement at neutral pH.In the last chapter, we developed also a new methodology for the dimerization of unprotected peptides. This method is based on siloxane bond formation between hybrid dimethylhydroxysilane peptides. A peptide derived from the tumor suppressor protein p53 was dimerized in water, at neutral pH using this methodology. The method was compared with the homodimerization carried out by Cu(I) azide-alkyne cycloaddition (CuAAC). For that purpose, p53 peptide derivative was synthesized with azide and alkyne linkers at the N-terminus. At last, several homodimers of growth hormone releasing hexapeptide (GHRP-6) were synthesized, with dimethylhydroxy silane linkers placed at various positions.

The nuclear structme of 17Ne and 13O has been studied by the 20Ne(3He,6He)17Ne and 16O(3He,6He)13O reactions at 70 MeV and 80 MeV, respectively. Fifteen levels were identified, and angular distributions have been measured for nine of these levels in 17Ne, while for 13O eighteen levels were identified, but angular distributions were obtained for only ten levels. The observed transferred angular momentum dependence of these angular distributions allowed spin-parity assignments. The T= 3/2 quartet analog states in mass A=17 have been completed for six levels. The results of the isobaric multiplet mass equation analysis show a slight linear dependence of the b and c coefficients on the excitation energy. It was found that the coefficients for the positive parity states do not follow the systematics of the negative parity states. The absolute values of the b and c coefficients are larger for the positive parity states. An analysis in terms of Coulomb energy displacement indicates a possible configuration mixing or core polarization effect in these states. The d coefficient also has a large deviation from zero, only for the positive parity states indicating a possible expansion of the radial wavefunction or some isospin symmetry breaking effects. Further detailed theoretical interpretation of these effects may bring valuable information about the configuration and structure of these states. The leveis in 13O were measured with good energy resolution, and thus, it was possible to identify the first excited state unambiguously. However, if one identifies this state as the analog of the known first excited state in the mirror nucleus 13B, this leads to one of the largest level shifts known in literature. / The nuclear structme of 17Ne and 13O has been studied by the 20Ne(3He,6He)17Ne and 16O(3He,6He)13O reactions at 70 MeV and 80 MeV, respectively. Fifteen levels were identified, and angular distributions have been measured for nine of these levels in 17Ne, while for 13O eighteen levels were identified, but angular distributions were obtained for only ten levels. The observed transferred angular momentum dependence of these angular distributions allowed spin-parity assignments. The T= 3/2 quartet analog states in mass A=17 have been completed for six levels. The results of the isobaric multiplet mass equation analysis show a slight linear dependence of the b and c coefficients on the excitation energy. It was found that the coefficients for the positive parity states do not follow the systematics of the negative parity states. The absolute values of the b and c coefficients are larger for the positive parity states. An analysis in terms of Coulomb energy displacement indicates a possible configuration mixing or core polarization effect in these states. The d coefficient also has a large deviation from zero, only for the positive parity states indicating a possible expansion of the radial wavefunction or some isospin symmetry breaking effects. Further detailed theoretical interpretation of these effects may bring valuable information about the configuration and structure of these states. The leveis in 13O were measured with good energy resolution, and thus, it was possible to identify the first excited state unambiguously. However, if one identifies this state as the analog of the known first excited state in the mirror nucleus 13B, this leads to one of the largest level shifts known in literature.

La physique à la drip-line offre l'opportunité d'étudier les noyaux à la limite du paysage couvert par la physique nucléaire, où l'addition de nouveaux protons et neutrons ne peut plus être supportée. A l'approche de la drip-line, la structure même des noyaux se trouve modifiée par rapport aux plus stables. Les fermetures de couches classiques disparaissent et de nouveaux phénomènes surgissent, tels que la clusterisation et les halos. Dans ce travail, nous avons revisité la spectroscopie du noyau non-lié 10He par l'intermédiaire d'une réaction de transfert d'un proton 11Li(d,3He) à 50 AMeV étudiée par la méthode de masse manquante. Pour ce faire, nous avons utilisé le détecteur MUST2 sur la ligne RIPS du RIKEN Nishina center. La partie expérimentale de ce travail repose sur un dispositif novateur utilisant des détecteurs de silicium très fins. Pour la première fois, la mise en oeuvre de ces derniers en conjonction avec des détecteurs sensibles en position de haute granularité a été démontrée. L'approche par réaction de transfert conduit à des résultats très clairs sur la position du premier état résonant du 10He, trouvé dans ce travail à 1.4(3) MeV, en accord avec la plupart des précédents travaux expérimentaux. L'obtention pour la première fois d'un facteur spectroscopique permet de nous renseigner sur la nature du recouvrement <10He|11Li>, trouvé égale à 0.13(6). / Physics at the drip-line offer the opportunity to study nuclei at the limits of the nuclear landscape, where additional protons or neutrons can no longer be kept in the nucleus. In the vicinity of the drip-lines, the structural features of the nuclei change with respect to the more stable nuclei, the normal shell closure of particles disappear and new phenomena appear, such as clusterization and halo. In this work we have revisited the spectroscopy of the unbound 10He by means of the 11 Li(d,3 He) reaction at 50 AMeV studied in missing mass method with the MUST2 array at the RIPS beam line of RIKEN. The experimental part of this study rely on new detection set-up using very thin silicon detectors. For the first time, the effective operability of these detectors, in conjunction with high granularity position sensitive detectors, was proved. The transfer reaction approach led to clear results on the 10He first resonant state position, found in this work at 1.4(3) MeV, in agreement with most of the previous experimental works. The extraction of a spectroscopic factor for the first time shed light on the <10He|11Li> overlap, found to be 0.13(6).

Ce travail présente la synthèse de ligands mixtes germylène-sulfoxyde qui ont été utilisés en chimie de coordination avec plusieurs métaux de transitions et finalement appliqués en catalyse de transfert d'hydrogène. Une seconde partie de ce travail est axée sur la synthèse de nouveaux métallylènes stabilisés par un sulfonimidamide qui ont été appliqués en catalyse de polymérisation. Le premier chapitre présente l'état de l'art des germylènes avec leurs synthèses, modes de stabilisations et finalement leur chimie de coordination et leurs applications en catalyse. Les sulfoxydes seront aussi décrits pour leur application en tant que ligands hémilabiles. Le second chapitre décrit la synthèse de ligands germylène-α-sulfoxide avec une diversité structurale (cyclohexyle, tBu, tolyle..) et avec différents degrés d'oxydation du souffre (thioéther, sulfoxyde et sulfone). La chimie de coordination a permis la synthèse de complexes bis-germylène du groupe IV, bidentes du ruthénium ainsi que monodentes du ruthénium. Le troisième chapitre présente l'extension de la méthodologie à la formation de ligands germylène-β-sulfoxide. L'influence du groupement espaçant les deux entités sur la chimie de coordination a été étudiée et montre la formation de complexes bidentates du groupe IV. La coordination du ruthénium a mené à la surprenante synthèse d'un complexe bis-ruthénium caractérisé par diffraction des rayons X. Finalement, la nouvelle architecture a permis d'obtenir des complexes de nickel (0) dont une structure avec des ligands carbonyles permettant une comparaison TEP. Le chapitre quatre présente la synthèse de nouveaux ligands, analogues des amidinates, pour la stabilisation des métallylènes avec un atome central soufré. L'effet apporté sur les métallylènes par ces nouveaux ligands sulfonimidamides a été étudié par calcul DFT. Le dernier chapitre est centré sur l'application de plusieurs complexes de ruthénium en catalyse de transfert d'hydrogène avec plusieurs substrats carbonylés. L'utilisation de stannylènes stabilisés par des ligands sulfonimidamides a été étudiée en catalyse de polymérisation de la ε-caprolactone. / This work concerns the synthesis of mixed germylene-sulfoxide ligands that were involved in coordination chemistry with several transition metals and finally applied in hydrogen transfer catalysis. In a second part, the synthesis of new metallylenes stabilized by a sulfonimidamide group was develop, which were applied in polymerization catalysis. The first chapter is centered on the state of the art of germylenes with their synthesis, their stabilization and finally their coordination chemistry and applications in catalysis. Sulfoxides will also be described for their potential as hemilabile ligands. The second chapter describes the synthesis of germylene-α-sulfoxide ligands as structurally tunable structures (cyclohexyl, tBu, tolyl..) and with different oxidation states of the sulfur atom (thioether, sulfoxide and sulfone). The coordination chemistry led to the formation of bis-germylene group IV complexes, bidentate-ruthenium and monodentate ruthenium complexes. The third chapter shows the extension of the methodology to the formation of germylene-β-sulfoxide ligands. The influence of the spacer between the two entities on the coordination chemistry was studied and shows the formation of bidentate group IV complexes. Coordination to ruthenium led to obtain a surprising bis-ruthenium complex characterized by X-ray diffraction analysis. Finally, the novel architecture allowed to obtain two nickel (0) complexes, one of them including carbonyl ligands permitting a TEP comparison. The fourth chapter is centered on the synthesis of new ligands, analogue of amidinate substituents for metallylene stabilization with a central sulfur atom. The effect brought to the metallylene center by the sulfonimidamide was studied by DFT calculations. The last chapter concerns the application of several ruthenium complexes in hydrogen transfer reaction with various carbonyl compounds. The use of sulfonimidamide stannylenes was also investigated in polymerization catalysis of ε-caprolactone.