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<p> We investigated the local electronic properties of the triangular-lattice materials NaxCO2 (x = 0.3, 0.72) and the superconductor Na0.3Co02-1.3H20 by 59Co and 170 Nuclear Magnetic Resonance(NMR). For Na0.72Co02 , 59Co NMR line shape shows clearly that there are two types of Co sites - Co(A) site and Co(B) site. The electronic character of Co(A) site is close to that of the less magnetic Co+3-like ion with spin rv 0, while the electronic character of Co(B) site is close to that of the strongly magnetic Co+4-like ions with spin ~ 1/2. The temperature dependence of the Knight shifts suggests that the Co(A) and Co(B) sites are electronically coupled, which is not consistent with simple phase separation. The local Co electronic environments propagate to the adjacent 0 layers through p-d hybridization. Therefore, there are two types of oxygen sites, O(A) site and O(B) site. We introduced a different route to do K vs x plot analysis for the Co sites and determined that for the Co sites, Karbitat(A) is 1.816 % and Kspin(A) is about rv 0.2 %; Karbitat(B) is 4.0255 % and Kspin(B) is at least 1.5 %. For Na0 .3Co02 , 59Co NMR line shape shows that the Co valence is averaged out in this material. There are two types of oxygen sites, 0( C) site and O(D) site, presumably because of the nearest neighbor Na+ sites. The constant behavior of Knight shifts below 100 K for both the Co and 0 sites suggests the emergence of a low temperature canonical Fermi-liquid behavior. For the superconductor Na0.3CoO2-1.3H20, both 59Co and 170 NMR line shapes show that there is only one type of Co site and oxygen site. The Knight shifts of 59Co and 170 are temperature independent below 100 K down to Tc. Combined with our spin-lattice relaxation 1/T 1 T measurements, we can rule out the possibility of ferromagnetic scenario of spin excitations above Tc. </p> / Thesis / Master of Science (MSc)

<p> The formation of soluble amyloid oligomers by polypeptide chains is the main pathogenic mechanism underlying several neurodegenerative disorders including some of the most common debilitating and aging-related illnesses such as Alzheimer's and Parkinson's diseases. However, the molecular basis of polypeptide oligomerization and amyloid formation is currently not fully understood. In this thesis the focus will be on the early steps of oligomer formation that precede the nucleation of amyloid fibrils, that are still reversible. The reversibility of these initial self-association equilibria makes them an attractive target for therapeutic intervention in the treatment of amyloid diseases. Specifically three general questions will be addressed: (a) What are the residues within a given polypeptide chain that mediate self-recognition? (b) What are the driving forces for self-association? (c) Is self-recognition coupled with conformation changes? </p> <p> The objective of this thesis is to provide initial responses to these key questions using as prototypical system the Ap (12-28) peptide, which has been previously proposed as a model for the initial self-association events that are linked to Alzheimer's disease. Given the flexibility of this peptide the main tool for its investigation will be Nuclear Magnetic Resonance (NMR) spectroscopy. Specifically, both classical (i.e., TOCSY and NOESY) and more novel (i.e. saturation transfer difference and off-resonance relaxation) NMR experiments were used to probe the soluble oligomers through the comparative analysis of samples with different monomer/oligomer distributions. The combined analysis of this integrated set of experiments reveals that while the residues in the central hydrophobic core (CHC) drive self-recognition, stable oligomers require a conformational change towards more folded structures that affects residues well outside the CHC. The conformational change occurring upon self-association thus effectively couples CHC and non-CHC residues. This model may also explain why mutations outside the CHC (i.e. E22, D23) can affect significantly the kinetics of self-association. </p> <p> / Thesis / Master of Science (MSc)

A novel partial agonist of the exchange protein activated by cAMP isoform 1 (EPAC1), I942, was recently discovered and shown to reduce the guanine exchange factor activity of cAMP-bound EPAC1 to approximately 10% relative to cAMP activation. However, the inhibition mechanism of I942 remains unknown. Here, we utilize NMR spectroscopy to probe the inhibitory I942 - EPAC1 interactions at atomic resolution. The EPAC1 - I942 interface was mapped through intermolecular NOEs measured by 15N and 13C filtered NOESY-HSQC experiment. Intermolecular NOE mapping combined with other protein NMR methods, such as saturation transfer difference, transfer Nuclear Overhauser Effect spectroscopy and chemical shift mapping, we revealed that I942 interacts with the phosphate binding cassette (PBC) and base binding region (BBR) of the EPAC1 cyclic nucleotide binding (CNB) domain, similar to cAMP. The PBC controls the conformation of the hinge region, and subsequently, allosterically shifts the hinge region between its active/inactive states. Molecular dynamics simulation based on the NMR spectroscopy data revealed that EPAC1-CNB adopts an intermediate conformation between its inactive and active states, which explains the partial agonist nature of I942. / Thesis / Master of Science (MSc) / The exchange protein activated by cAMP (EPAC) is a receptor for the classical secondary messenger cAMP. EPAC is present in multiple human systems and plays a pivotal role in the development of a wide range of diseases. In this study, we aim to establish the inhibition mechanism of a novel small molecule EPAC inhibitor/partial agonist I942 using NMR spectroscopy with the goal of achieving a better understanding of EPAC inhibition and paving the way for new small molecule EPAC inhibitors that can potentially treat EPAC-related diseases such as heart failure and diabetes.

Mass spectrometry was used in conjunction with numerous 1-D and 2-D NMR techniques to determine the structures, devoid of stereochemistry, of five different compounds isolated from the extracts of Eunicea succinea, Eunicea tourneforti, and an unidentified species isolated from the Eunicea genus by Professor Meledath Govindan, of the University of the Virgin Islands. Three of the compounds were then identified as the known compounds eunicin, 12,13-bisepieupalmerin, and 7(S),8(S)-epoxy-1(S),11(R)-dolabella-3E,12(18) -dien-13-one by comparison of their spectroscopic data and optical rotations with those published in the literature. Optical rotations could not be measured accurately for the other two compounds because of small sample sizes; therefore, another method had to be found to elucidate the stereochemistry of these two structures. To solve this problem, molecular modeling and NOESY were employed. Comparison of the NOESY interactions to the thermodynamically available conformations of several possible stereoisomers, calculated by molecular modeling, proved to be a useful technique. One of the remaining two structures was identified as the known stereoisomer euniolide. The stereochemistry of the one remaining structure could not be assigned because sample size was not large enough to obtain a clean NOESY spectra. Finally, based on published synthetic work by Corey and Kania, the absolute stereochemistry of the dolabellane was revised to 7(R),8(R)-epoxy-1(R),11(S)- dolabella-3E,12(18)-dien-13-one. / Master of Science

Ion transport of electrolytes determines the performance of many electroactive devices, from fuel cells to batteries to soft mechanical actuators. This dissertation aims to address some fundamental issues regarding ion transport of ion dense electrolytes using electrophoretic NMR and NMR diffusometry. I first describe the design and fabrication of the first instrumentation capable of reliable ENMR on highly ion-dense electrolytes such as ionic liquids and electrolytes for zinc-air batteries. I design a new electrophoretic NMR sample cell using parallel capillaries to investigate the electrophoretic mobilities of pure ionic liquids. It shows the first study of a highly ion-dense electrolyte with electrophoretic NMR. Then I employ NMR diffusometry and electrophoretic NMR to investigate ion association of pure ionic liquids. Then I use electrophoretic NMR technique to investigate the electrophoretic mobilities of electrolytes for zinc-air batteries. For Zn2+ salt added dicyanamide (dca) based ionic liquids, I investigate the effects of Zn2+ salt on chemical shift of dca and ion motion. The combination of mobilities measurements and diffusion measurements provides some new insight of ion aggregation. We explore ion transport of ionic liquids inside the ionic polymer Nafion as a function of hydration level. When ionic liquids diffuse inside ionic polymers, isolated anions diffuse faster (e 4X) than cations at high hydration whereas ion associations result in substantially faster cation diffusion (d 3X) at low hydration inside membranes, revealing prevalent anionic aggregates. Finally, we compare diffusion activation energy measurements in a hydrated perfluorosulfonate ionomer and aqueous solutions of triflic acid, which provides insight into water transport dynamics on sub-nm lengthscales. And we explore the physical meaning of activation energy, characterizing local intermolecular interactions that occur on the pre-diffusional (~ 1 ps) timescale. / Ph. D.

Polymeric methylenebis(phenylisocyanate) (pMDI) has increasingly been used in the wood particulate composite industry. Wood composites, especially oriented strand board (OSB) are made with many variations of wood species. Little research has been done to investigate how pMDI adhesion has been affected by species. The present is divided into two parts. First, mode I fracture mechanics and surface free energy analysis was performed to investigate differences in adhesion between southern yellow pine and yellow-poplar bonded with pMDI. Secondly, an improvement in the synthesis of 13C, 15N labeled pMDI is discussed. Mode I fracture results show that pMDI adhesion was affected differently by southern yellow pine compared yellow poplar. The shear energy release rate was significantly higher in pine/pMDI composites than in yellow poplar/pMDI composites. The total surface energy of southern yellow pine was shown to be significantly greater than yellow poplar. The free energy of adhesion (DG) of the pine/pMDI and the poplar/pMDI was investigated. The DG indicated that the pine/pMDI system would take more energy to separate compared to the poplar/pMDI system. Lastly, a double-labeled 13C, 15N pMDI adhesive was successfully synthesized to produce Solid-State NMR composites. / Master of Science

Tesis por compendio / Sus1 is a conserved protein involved in chromatin remodeling and mRNA biogenesis. The SUS1 gene of Saccharomyces cerevisiae is unusual, as it contains two introns and is alternatively spliced, retaining one or both introns in response to changes in environmental conditions. SUS1 splicing may allow the cell to control Sus1 expression, but the mechanisms that regulate this process remain unknown. In this thesis project, we have investigated whether the structure adopted by SUS1 RNA sequences contributes to regulate the splicing of this gene. Using in silico analyses together with NMR spectroscopy, gel electrophoresis and UV thermal denaturation experiments, we first show that the downstream intron (I2) of SUS1 forms a weakly-stable, 37-nucleotide stem-loop structure containing the branch site near its apical loop and the 3' splice site after the stem terminus. A cellular assay revealed that two of four mutants containing altered I2 structures had significantly impaired SUS1 expression. Semi-quantitative RT-PCR experiments indicated that all mutants accumulated unspliced SUS1 pre-mRNA and/or induced distorted levels of fully spliced mRNA relative to wild-type. Concomitantly, Sus1 cellular functions in histone H2B deubiquitination and mRNA export were affected in I2 hairpin mutants that inhibited splicing. The second part of the thesis project focuses on the exon located between the two introns of the SUS1 gene. This middle exon (E2) can be skipped during splicing, is generated in circular form, and has been found to influence the splicing of the flanking introns, an unusual situation in budding yeast where splicing mainly relies on intron recognition. Using NMR spectroscopy, gel electrophoresis, UV thermal denaturation and ribose 2'-OH modification experiments combined with computational predictions, we show that E2 of SUS1 comprises a conserved double-helical stem topped by a three-way junction. One of the hairpins emerging from the junction exhibited significant thermal stability and was closed by an unusually structured purine-rich loop. This loop contained two consecutive sheared G:A base pairs and was structurally related to the substrate loop of the VS ribozyme. Cellular assays revealed that three mutants containing altered E2 structures had impaired SUS1 expression and that a compensatory mutation restoring the conserved stem recovered expression to wild-type levels. Semi-quantitative RT-PCR experiments indicated that all mutants were capable of altering the quantities of unspliced and/or fully-spliced SUS1 RNA transcripts relative to wildtype. Overall, the results gathered in this thesis project indicate that RNA structures formed by the middle exon and the second intron of the S. cerevisiae SUS1 gene are relevant for splicing and also influence other processes of SUS1 mRNA biogenesis. / Sus1 es una proteína conservada implicada en remodelación de cromatina y biogénesis de moléculas de ARNm. El gen SUS1 de Saccharomyces cerevisiae es peculiar, ya que contiene dos intrones y sufre un proceso de ayuste (corte y empalme) alternativo, reteniendo uno o ambos intrones en respuesta a cambios en las condiciones ambientales. El ayuste del ARNpre-m de SUS1 puede permitir a la célula controlar la expresión de la proteína Sus1, pero los mecanismos que regulan este proceso son poco conocidos. En este proyecto de tesis hemos investigado si la estructura adoptada por secuencias de ARN de SUS1 contribuye a regular el proceso de ayuste de este gen. Utilizando análisis in silico junto con espectroscopia de RMN, electroforesis en gel y experimentos de desnaturalización térmica monitorizados por UV, primero demostramos que el ARN del segundo intrón (I2) del gen SUS1 forma una horquilla débilmente estable de 37 nucleótidos. Esta horquilla contiene nucleótidos del sitio de ramificación (branch site) en su bucle apical y nucleótidos del sitio 3' de empalme adyacentes al extremo inferior del tallo. A través de ensayos funcionales descubrimos que dos de cuatro mutantes que alteran la estructura de la horquilla I2 exhibían peor expresión de SUS1. Experimentos de RT-PCR semicuantitativos indicaron que todos los mutantes acumularon ARNpre-m SUS1 no ayustado y/o indujeron cambios en los niveles de ARNm maduro con respecto a la secuencia silvestre. Además, las funciones celulares de Sus1 relativas a desubicuitinación de histona H2B y transporte de ARNm se vieron afectadas en los mutantes de la horquilla I2 que inhibían el proceso de ayuste. La segunda parte de la memoria de tesis se centra en el análisis del exón central (E2) situado entre los dos intrones del gen SUS1. Este exón puede eliminarse durante el proceso de ayuste, se genera en forma circular, e influye en el procesamiento de los intrones adyacentes, una situación inusual para las regiones exónicas de S. cerevisiae, donde el ayuste se basa principalmente en el reconocimiento de intrones. Utilizando experimentos de espectroscopía de RMN, electroforesis en gel, desnaturalización térmica y modificación química combinados con predicciones computacionales, demostramos que el ARN del exón E2 de SUS1 forma un tallo conservado de doble hélice coronado por una intersección de tres hélices. Una de las horquillas que emergen de esta intersección presentó una estabilidad térmica significativa, así como un bucle apical rico en purinas inusualmente estructurado. Este bucle contiene dos pares de bases G:A consecutivos y está estructuralmente relacionado con el bucle de substrato de la ribozima VS. Ensayos celulares revelaron que tres mutantes con estructuras modificadas de E2 exhibían peor expresión de SUS1, y que una mutación compensatoria que restauraba el tallo conservado recuperaba la expresión a los niveles de la secuencia silvestre. Experimentos de RT-PCR semicuantitativos indicaron que todos los mutantes de E2 eran capaces de alterar las cantidades de transcritos ayustados y no ayustados de SUS1 con respecto a la secuencia silvestre. En general, los resultados obtenidos en este proyecto de tesis indican que las estructuras de ARN formadas por el exón central y el segundo intrón del gen SUS1 de S. cerevisiae son relevantes para el ayuste y otros procesos implicados en la biogénesis del ARNm del gen SUS1. / Sus1 és una proteïna conservada implicada a la remodelació de la cromatina i la biogènesi de l'ARNm. El gen SUS1 de Saccharomyces cerevisiae és inusual, ja que conté dos introns i s'empalma de manera alternativa, retenint un o ambdós introns en resposta a canvis en les condicions ambientals. L'empalmament de SUS1 pot permetre a la cèl·lula controlar l'expressió de Sus1, però els mecanismes que regulen aquest procés són segueixen sent desconeguts. En aquest projecte de tesi investiguem si l'estructura adoptada per seqüències d'ARN de SUS1 contribueix a regular l'empalmament d'aquest gen. Emprant anàlisi in silico juntament amb espectrometria de RMN, electroforesi en gel i experiments de desnaturalització tèrmica d'UV, es mostra primer que l'intró aigües a baix (I2) de SUS1 forma una estructura de forqueta de 37 nucleòtids feblement estable que conté el lloc de la branca a prop del seu bucle apical; i el lloc d'empalmamnet 3¿ després de l'extrem de la forqueta. Un assaig cel·lular va revelar que dos de quatre mutants que contenien estructures alterades de l'I2 havien modificat significativament l'expressió de SUS1. Els experiments semi-quantitatius de RT-PCR van indicar que tots els mutants acumulaven el pre-ARNm madur respecte al tipus salvatge. Concomitantment, les funcions cel·lulars de Sus1 a la desubiqüitinació de la histona H2B i l'exportació d'ARNm es van veure afectats als mutants de la forqueta d'I2 que inhibeixen l'empalmament. La segona part del projecte de tesi se centra a l'exó situat entre els dos introns del gen SUS1. Aquest exó (E2) es pot ometre durant l'empalmament, es genera amb forma circular, i s'ha trobat que influeix a l'empalmamet dels introns que flanquegen, una situació inusual al llevat on l'empalmament està basat principalment al reconeixement d'introns. Emprant espectroscòpia de RMN, electroforesi en gel, desnaturalització tèrmica d'UV i experiments de modificació de ribosa 2¿-OH combinats amb prediccions computacionals, mostrem que E2 de SUS1 comprén un tall conservat de doble hèlix corornat per una unió de tres vies. Una de les forquetes que emergeixen de la unió, va mostrar una estabilitat tèrmica significativa i va ser tapada per un bucle ric en purina inusualment estructurat. Aquest bucle contenia dos pars de bases G:A tallats consecutivament i estava estructuralment relacionat amb el bucle de substrat del ribozim VS. Els assajos cel·lulars van revelar que tres mutants que contenien estructures alterades de E2 havien alterat l'expressió de SUS1 i que una mutació compensatòria que restaurava el tall conservat recuperava l'expressió a nivells del tipus salvatge. Els assajos cel·lulars van revelat que tres mutants que contenien estructures alterades d'E2 havien alterat l'expressió de SUS1 i que una mutació compensatòria que restaurava el tall conservat recuperava l'expressió a nivell d'un tipus salvatge. Els experiments semi-quantitatius de RT-PCR van indicar que tots els mutants eren capaços d'alterar les quantitats de transcrits d'ARN de SUS1 no empalmats i/o empalmats en relació amb el tipus salvatge. En general, els resultats obtinguts en aquesta investigació indiquen que les estructures d'ARN formades per l'exó mitjà i el segon intró de SUS1 de S. cerevisiae són rellevants per l'empalmament i també influeixen a altres processos de biogènesi de l'ARN de SUS1 / Abuqattam, AN. (2017). IDENTIFICATION OF RNA STRUCTURES MODULATING THE EXPRESSION OF THE mRNA BIOGENESIS FACTOR SUS1 [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/90524 / Compendio

No / The structure of 5-N-hydroxyethanequindoline hydrochloride methanolate, C17H15ON2 Cl·½CH3OH, M r = 314.78, has been determined from X-ray diffraction data. The crystals are monoclinic, space group C2/c, with Z = 8 molecules per unit cell and a = 18.179(11), b = 7.317(5), c = 24.125(15) Å, β = 110.155(10)°, V c = 3012(3) Å3, crystal density D c = 1.388 Mg m−3. The structure was solved by direct methods, and the asymmetric unit comprises the 5-N-hydroxyethanequindoline hydrochloride and ½CH3OH moiety. The methanol is unusually disordered over a twofold axis with the C atom slightly removed from the twofold axis. Restraints were applied to the bond lengths of the two components of the disordered CH3OH, and to the anisotropic thermal displacement parameters of the disordered CH3OH carbon atom. The heterocyclic quindoline ring system and the first C atom of the hydroxyethane side chain are planar within 0.02 Å, with the terminal C–OH atoms of the side chain significantly out of the plane. The crystal structure is maintained via three hydrogen bonds all involving the chlorine atom an oxygen in the hydroxyethane side chain, a nitrogen in the quindoline moiety and the methanol oxygen.