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Solid-state NMR serves as a powerful method for investigating atomic-level details of insoluble biomolecules, enabling the determination of protein 3D structures and probing molecular motions across a broad range of timescales. In this thesis, I present structural studies on a novel heterotypic and functional amyloid, dynamics studies, and chemical shift anisotropy studies of a microcrystalline protein, ubiquitin. In Chapter 1, I provide a summary of the main interactions in solid-state NMR and discuss relevant pulse sequences employed in this thesis. Chapter 2 briefly explores the characteristic properties of amyloids, highlighting well-studied examples of disease-related and functional amyloids. Special treatments employed in amyloid structure determination using solid-state NMR are also summarized. Chapter 3 presents structural studies on a heterotypic functional amyloid, mcmvM45-hsRIPK3, where M45 is a protein encoded by murine cytomegalovirus (MCMV) and RIPK3 is from humans. Both M45 and RIPK3 belong to a family of RHIM-containing proteins, which are involved in innate immunity and immune response through necroptosis. SSNMR data on various isotopically labeled samples enable the chemical shift assignment for both M45 and RIPK3, providing intra- and inter-molecular contacts. By combining these constraints, we calculate the structure of the hetero-amyloid M45-RIPK3, reporting two structures distinct from RIPK1-RIPK3. In Chapter 4, I measure backbone 15N-13CO order parameters of microcrystalline ubiquitin using DCP-REDOR. Two isotopically labeled samples, 1-13C-glucose and 1,3-13C-glycerol, D₂O labeled, are studied and compared, identifying mobile residues and assessing the effect of isotropic labeling on the measurements of backbone 15N-13Co order parameters. Experimental order parameters are compared with a 1μs MD simulation for insights. Chapter 5 focuses on the chemical shift anisotropy (CSA) of uniformly labeled microcrystalline ubiquitin using a novel pulse sequence allowing the measurement of large CSAs under practical conditions. We explore CSA parameter trends, correlations between isotropic shifts, and hydrogen bond geometries. Comparison with solution-NMR results demonstrates high consistencies with asymmetry parameters (η), providing insights into the motion modes of microcrystalline proteins alongside order parameter measurements. Chapter 6 provides a comprehensive summary of the conclusions drawn from the preceding chapters, while also outlining future directions for each project.

<p> Experimental carbon-13 nuclear magnetic resonance chemical shifts are given for substituted 1,3-dioxanes, tetrahydrofurans, dihydro-2-furanones and succinic anhydrides. The experimental data are used to improve and extend the carbon-13 chemical shift prediction parameters which allow one to estimate with considerable accuracy unknown chemical shift values.</p> / Thesis / Master of Science (MSc)

<p> Nuclear magnetic resonance spectroscopy has been used to study donor-acceptor complexes of boron trifluoride with ketones, water, and methanol. Various NMR techniques have been applied which, in certain favourable cases, have yielded information on the stoichiometry, structure, relative stabilities, and reactions of the complexes in solution. A new method is proposed for the detection of BF3 complexes having low formation constants.</p> <p> The magnitude and sign of the B-F coupling constant in the tetra-fluoroborate anion have been found to be dependent on the solvent. This solvent dependence is interpreted in terms of solvation and association of the ion in solution.</p> <p> 19F spectra of 10BF3 over a range of temperatures agree well with line shapes calculated, using Pople's expressions, for coupling of fluorine-19 with boron-10 (I=3) which is undergoing partial quadrupole relaxation.</p> / Thesis / Doctor of Philosophy (PhD)

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The nuclear magnetic resonance spectrum of the protons in rubidium hydrogen sulfate, and the spin-lattice relaxation time for the protons in lithium hydrazinium sulfate have been examined as a function of temperature, in order to obtain information about the motions of the protons in the different states of these substances. Rubidium hydrogen sulfate becomes ferroelectric below -15°C, and the possibility of a change in the proton resonance linewidth associated with this transition was investigated. The free-precession (spin echo) NMR technique was used to obtain the proton spin-lattice relaxation time in lithium hydrazinium sulfate, over a wide temperature range. The observed changes in relaxation time gave further data on the hydrazinium ion motions that were inferred from proton NMR studies by Cuthbert and Petch in 1963. / Thesis / Master of Science (MSc)

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