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A series of bis(ethene)rhodium(I) complexes of β-diketonates and similar ligands were prepared. The complexes were characterized by elemental analysis, ¹H NMR spectroscopy and mass spectrometry. Crystal structures are reported for bis(η²-ethene)-(1,3(1-ferrocenyl)butanedionato-0.0 ')rhodium(I), 13, (1,5-cyclooctadieiie)(2-acetylphen-oxy-0,0')rhodium(I),18, and some related molecules. [FORMULA OMITTED] The complexes are catalyst precursors for the homogeneous hydrogenation of olefins and the hydrosilylation of ketones. The generated hydrogenation catalysts are effective for the reduction of sterically unhindered carbon-carbon double bonds. These catalysts are active in the presence of alcohol, aromatic and carboxylic acid functional groups on the olefin substrate. The catalysts decompose to give a heterogeneously active rhodium precipitate when reducing olefinic substrates which are too sterically hindered. Study of the hydrosilylation reaction revealed that the complexes generate active hydrosilylation catalysts. A series of optically active ligands were tested for their ability to effect asymmetry in the silyl ether products. The bis(ethene)rhodium(I) complexes are fluxional in the NMR time scale due to the motion of the ethylene ligands. A detailed ¹H NMR study was conducted on one of the complexes, bis(η²-ethene)(2-acetylphenoxy-0,0 ')rhodium(I) 15 to explore the nature of the rearrangement. [FORMULA OMITTED] The ¹H NMR study revealed the presence of two different modes of fluxionality. The first type of motion is intramolecular in nature and is due to the rotation of the ethylene ligands about the rhodium-ethylene bond axis. The second type is intermolecular in nature and is due to exchange of the ethylene ligands. This exchange is a measure of the lability of the ethylene ligands. The system allowed the separate study of the ethylene ligands and ΔG‡ values were obtained for each ligand for both of the exchange processes. The results of the study indicate the independance of the ethylene ligands with respect to both fluxional processes. / Science, Faculty of / Chemistry, Department of / Graduate

The pathogenesis of low back pain is complex but likely involves the intervertebral disc (Nachemson, 1976). Direct evidence for its importance is lacking because an accurate in vivo method of imaging the lumbar intervertebral disc has not been established. The objective of this research was to develop classifications of gross morphologic appearance and magnetic resonance image (MRI) of the disc that describe the changes associated with aging and degeneration, thereby permitting interpretation of the MRI in terms of gross morphology and allowing correlation of morphologic, chemical, mechanical, radiologic and epidemiologic data with a standard reference of disc aging and degeneration. The classifications were developed on the basis of literature review, detailed examination of 55 discs and expert advice. Two sets of three observers, one for the morphologic classification and one for the MRI classification evaluated 68 life size randomized duplicates of discs making detailed observations about overall category and 17 regional morphologic parameters and 11 regional MRI parameters. The data was tested to demonstrate the validity of the classifications using established criteria (Tugwell & Bombardier, 1982; Guyatt 4 Kirschner, 1985; Feinstein, 1985). The consistency with which the classifications could be applied was evaluated by calculating weighted kappa, a statistical test of agreement that corrects for agreement by chance; the ability of the classifications to distinguish stages in the process of ageing and degeneration by stepwise discriminant analysis; their conformity with other measures by comparisons within and between classifications and, comparisons with histologic and chemical data. The degree of agreement for all six intra-observer pairs was 'almost perfect' (weighted kappa > 0.80); for 5 interobserver pairs 'substantial' (weighted kappa > 0.60) and for one MRI interobserver pair 'moderate' (weighted kappa > 0.50). This represented a satisfactory level of agreement and indicated the classifications could be applied consistently (Feinstein, 1981). The linear regression model developed by stepwise discriminant analysis clearly demonstrated the ability of the classifications to distinguish distinct stages in disc aging and degeneration. Wilk's lambda, a likelihood ratio statistic reflecting discriminatory function, approached zero in both the morphologic (0.0408) and MRI (0.0H80) classifications. In both models, parameters pertaining to the nucleus pulposus of the disc accounted for the majority of the variance (morphologic partial R² 0.8598 and MRI partial R² 0.8811) suggesting nuclear parameters are the most important in distinguishing overall category. The correlation table generated by principal component analysis demonstrated that the categories assigned to regional parameters correlated significantly (p > 0.0001) with each other and with the overall category. From the linear combinations of parameters (principal components) generated the weighting of the nucleus pulposus behaved independently attesting to its importance. Comparisons of the morphologic and MRI classifications yielded high indices of trend (Pearson correlation coefficient of 0.81) and concordance (kappa of 0.62). Trends in the histologic and chemical data were consistent with the classifications but could not be evaluated statistically because only 15 specimens were studied. This research suggests that the classifications are valid and will form a basis for the interpretation of MRI. Preliminary evidence suggested MRI is sensitive to early changes in extracellular matrix composition not apparent in gross morphology. / Medicine, Faculty of / Graduate

Dipolar and quadrupolar couplings measured from NMR experiments, and order parameters obtained from these couplings, are reported for a number of small solutes dissolved in several nematic liquid crystals. These results are discussed in terms of the solute-solvent interactions. It has been shown that the interaction between the solute molecular quadrupole moment and a mean external electric field gradient due to the liquid crystal accounts for most but not all of the ordering of molecular hydrogen. The remaining contribution to ordering is discussed in terms of possible mechanisms. The anisotropic couplings observed for methane and acetylene are discussed in terms of a model which takes into account the interaction between the vibrations and rotations of the solute. Excellent agreement between the observed and calculated dipolar couplings is obtained. Evidence is given that these two solutes experience the same field gradient as molecular hydrogen. In a mixture of 55wt per cent 1132 (Merck ZLI 1132) and 45wt per cent EBBA (N-(4-ethoxybenzylidene)-4'-n-butylaniline at 301.4K the deuterons in D₂ experience no external electric field gradient. The order parameters of a series of solutes in this mixture are calculated in terms of a simple model for the short range interactions which depend on the size and shape of the solute. These calculated order parameters are in very good agreement with the experimental results. In liquid crystals where the field gradient is not zero it is shown that the combination of the short range interaction model and the electric field gradient - molecular quadrupole moment mechanism predicts the order parameters very well. / Science, Faculty of / Chemistry, Department of / Graduate

This thesis discusses in general terms the theory and application of time-differential perturbed angular correlation gamma-ray spectroscopy (TDPAC) to the study of solid state physics. The technique yields valuable information on the electric field gradients experienced by radionuclides which have been substituted for nonradioactive isotopes or inserted as impurities into various inorganic compounds. The indium halides are examined in a series of experiments. The first applications of this technique to the study of graphite intercalation compounds are discussed. / Science, Faculty of / Physics and Astronomy, Department of / Graduate

Magister Scientiae - MSC (Biochemistry) / Proteins play an important role in cells, as the morphology, function and activities of the cell depend on the proteins they express. The key to understanding how different proteins function lies in an understanding of the molecular structure. The overall aim of this thesis was the determination of the structure of DWNN domains. This thesis described the preparation of samples of human DWNN suitable for structural analysis by nuclear magnetic resonance spectroscopy (NMR), as well as NMR analysis. / South Africa

Contrary to a previous report, S-adenosyl-L-methionine (SAM) affords stable solutions in D2O and the 1H NMR spectrum can be determined. Comparison with the spectra of the model compounds adenosine, L-methionine and L-methionine-S-methyl sulfonium iodide allows complete assignment of the proton resonances. Coupling constants were determined by homonuclear decoupling and graphical analysis and were confined by computer simulation. Details of the molecular conformation were determined by application of the Karplus equation and calculation of relative rotational isomer populations. Evidence indicates that the ribose ring is puckered preferentially in the C3'-exo conformation and that the C4'-C5' bond is constrained to a rotamer in which the sulfonium center is gauche to H4'. No conformational constraints were detected for the Cα-Cβ and Cβ-Cα bonds of the methionine side chain. The purine ring was shown to be oriented preferentially anti by intermolecular association studies with adenosine 5'-phosphate in the presence of MN(II). Spectra of samples of (-)S-adenosyl-L-methionine of biological origin, differing in activity, counter ion and commercial source, have consistently revealed the presence of a small amount of the (+) sulfonium diastereomer. Arguments are presented to explain the failure of previous workers to detect (+)S-adenosyl-L-methionine in biological preparations.

The biological functions of proteins often rely on structural changes and the rates at which these conformational changes occur. Studies show that regions of a protein which are known to be involved in enzyme catalysis or in contact with the substrate are identifiable by NMR spectroscopy to be more flexible, evidenced through measuring order parameters of specific bond vectors. While generalized NMR can allow for detailed characterization of the extent and time scales of these conformational fluctuations, NMR cannot easily produce the structures of sparsely populated intermediates nor can it produce explicit complex atomistic-level mechanisms needed for the full understanding of such processes. Practically, preparing a protein with appropriate isotropic enrichment to study a set of specific bond vectors experimentally is challenging as well. Oftentimes, measuring the dynamics of neighboring bond vectors are necessitated. Detailed studies of the coupling interactions among specific residues and protein regions can be fulfilled by the use of molecular dynamics (MD) simulations. However, MD simulations rely on the ergodic hypothesis to mimic experimental conditions, requiring long simulation times. Simulations are additionally limited by the availability of accurate and reliable molecular mechanics force fields, which continue to be improved to better match experimental data. Much can also be learned from chemical theory and simulations to improve the methods in which experimental data is processed and analyzed. The overarching goals of this thesis are to improve upon the results generated by existing methods in NMR spin relaxation spectroscopy, whether that be through: (i) improving analytical techniques of raw NMR data or through (ii) supporting experimental results with atomistically-detailed MD simulations. The majority of this work is exemplified through the protein Escherichia coli ribonuclease HI (ecRNH). Ribonuclease HI (RNase H) is a conserved endonuclease responsible for cleaving the RNA strand of DNA/RNA hybrids in many biological processes, including reverse transcription of the viral genome in retroviral reverse transcriptases and Okazaki fragment processing during DNA replication of the lagging strand. RNase H belongs to a broader superfamily of nucleotidyl-transferases with conserved structure and mechanism, including retroviral integrases, Holliday junction resolvases, and transposases. RNase H has historically been the subject of many investigations in folding, structure, and dynamics. In support of the first aim, we discuss new methods of obtaining more precise experimental results for order parameters and time constants for the ILV methyl groups. Deuterium relaxation rate constants are determined by the spectral density function for reorientation of the C-D bond vector at zero, single-quantum, and double-quantum 2H frequencies. We interpolate relaxation rates measured at available NMR spectrometer frequencies in order to perform a joint single/double-quantum analysis. This yields approximately 10-15% more precise estimates of model-free parameters and consequently provides a general strategy for further interpolation and extrapolation of data gathered from existing NMR spectrometers for analysis of 2H spin relaxation data in biological macromolecules. In support of the second aim, we calculate autocorrelation functions and generalized order parameters for the ILV methyl side chain groups from MD simulation trajectories to assess the orientational motions of the side chain bond vectors. We demonstrate that motions of the side chain bond vectors can be separated into: (i) fluctuations within a given dihedral angle rotamer, (ii) jumps among the different rotamers, and (iii) motions from the protein backbone itself, through the C-alpha carbon. We are able to match order parameters of constitutive motions to conventionally calculated order parameters with an R2= 0.9962, 0.9708, and 0.9905 for Valine, Leucine, and Isoleucine residues, respectively. Some longer side chain residues such as Leucine and Isoleucine have correlated χ1 and χ2 dihedral angle rotational motions. This provides a method of evaluating the relative contributions of each constitutive motion towards the overall flexibility of a side chain. Multiple contributors of motion are possible for intermediate and low order parameters, signifying more flexible residues. While developing protocols for MD simulations, we evaluate the effects of running 1-microsecond long simulations and compare them to solution state NMR spectroscopy. If the overall tumbling time is removed from the simulation, then analysis blocks of 5-10 times the tumbling time is optimal to eliminate contributions from slower dynamics, which would not normally be measured in solution state NMR spectroscopy. We also assess the quality of the TIP4P(-EW) water model over TIP3P; although TIP4P simulates the isotropic tumbling time well for ecRNH, internal motions are equally not affected by either water model due to well-segregated motions. Additionally, the TIP4P water model does not appear to be able to replicate an axially symmetric shape for ecRNH (ecRNH is mostly spherical and only slightly axially symmetric). The final work of this thesis returns to the first overarching aim; we develop a specialized method that utilizes probability distribution functions to model spectral density functions. We derive the inverse Gaussian probability distribution function from general properties of spectral density functions at low and high frequencies for macromolecules in solution, using the principle of maximum entropy. The resulting model-free spectral density functions are finite at a frequency of zero and can be used to describe distributions of either overall or internal correlation times using the model-free ansatz. The approach is validated using 15N backbone relaxation data for the intrinsically disordered, DNA-binding region of the bZip transcription factor domain of the Saccharomyces cerevisiae protein GCN4, in the absence of cognate DNA.

<p>The history and philosophy underlying the design of the experimental component of an intensive introductory course in Nuclear Magnetic Resonance Spectroscopy (NMR) is presented. The concept of a " hands on " experience is used to transform the complete NMR novice into a competent operator. In addition to exposing the student to classical elucidation tools, advanced techniques, such as two dimensional and solid state NMR, are introduced. Through the use of unknowns, participants are encouraged to develop practical problem solving strategies. The detailed contents of the twenty-six experiments developed for the laboratory sessions are reviewed. The five day course schedule, and a coversheet, containing pertinent references for each experiment, is included. An examination of the unknowns utilized in the course, and a glossary of NMR terms is also presented. / Master of Science