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The thesis extends the concept of Irreducible Spherical Tensor Operators to spin dynamics problems in Nuclear Magnetic and Quadrupole Resonance Spectroscopy. NQR and solid state NMR deal with spin systems which behave as effective single spins in many, pulsed experiments, or as multispin systems when the dipolar interactions among spins cannot be neglected. The thesis considers problems in both areas. New analytic results are derived in the thesis for pure NQR spectroscopy and for the effect of radiofrequency fields in the presence of electric quadrupolar interaction. Three-pulse experiments are proposed for a spin-5/2 system and quantitative calculations are done for the indirect detection of new observables. Attention is focussed on the usefulness of the full-spin density matrix in all these calculations. In addition the concept of multispin tensors is applied to a system of spin-1/2 nuclei with strong dipolar interactions. This leads to coupled differential equations for correlations among spins for which an iterative solution can be obtained. The solutions are discussed in relation to Free Induction Decay experiments available for $ sp{19}$F nuclei in CaF$ sb2$ crystal.

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No description available.

No description available.

No description available.

New techniques in Fourier transform nuclear magnetic resonance spectroscopy are introduced with chemical applications to the study of molecules in the liquid state. Recently a theoretical description of magnetic resonance in terms of single transition operators has been introduced which provides a geometric interpretation of the behaviour of a spin system. This formalism is developed further and extended to the general description of a system of nonequivalent spin-1/2 nuclei. Operator combinations are introduced which allow extension of the convenient geometric representation to the concerted behaviour of coupled spins. The operator formalism is applied to the excitation and detection of multiple quantum transitions, providing a description of the processes in terms of rotating vectors. The process of coherence transfer in two-dimensional Fourier transform experiments is studied in detail and single transition operators are used to derive a general expression for the tip angle dependence of the detection process. A method of discriminating the sense of precession of double quantum coherence is presented and applied to the correlation of chemical shifts of carbon-13 spins in natural abundance. A new technique is presented for the correlation of chemical shift information in coupled proton spin systems in which excitation and indirect detection of double quantum transitions is used to assign coupling patterns in complex spectra. The tip angle dependence of the detection process is used to suppress all but direct correlation of spins. Methods for the measurement of heteronuclear coupling constants in proton spectra are introduced which discriminate the heteronuclear satellites from the parent proton resonance. The technique provides a sensitivity advantage over direct measurement of coupling in the heteronuclear spectrum. Ambiguities are encountered when the protonproton and proton-heteronuclear coupling constants are of the same order of magnitude. This problem is overcome by extension of the basic experiment to its two-dimensional analogue.

The kringle is a name given to a sequence of seventy nine residues linked by three disulphide bridges, multiple copies of which have been found in a number of proteins involved in haemostasis. Kringles from plasminogen and prothrombin have been studied using high resolution nuclear magnetic resonance (NMR) spectroscopy. Preliminary experiments demonstrated the kringles to be globular proteins in aqueous solution having well defined tertiary folds. Resonances have been assigned in the NMR spectrum to all but two of the aromatic residues of kringle 4 of human plasminogen and in addition a number of aliphatic residues have also been assigned. The majority of these assignments have been made to specific protons in the protein with the aid of chemically modified kringle 4 derivatives and comparison with other kringle protein spectra. Nuclear Overhauser enhancement studies have demonstrated a number of structurally significant side chain interactions. The lysine binding site of kringle 4 has been explored using two ω-aminocarboxylic acids as ligands. Further structural probes have included copper and lanthanide ions as well as chemically modified kringle 4 species. Combining all the data has allowed a preliminary molecular model to be built of part of kringle 4 incorporating all the residues between the second and third disulphide bridges. Preliminary studies demonstrate the possibility of being able to refine parts of this structure to an accuracy approaching that of X-ray crystallographic studies. A similar number of assignments have been made in the spectrum of bovine prothrombin fragment 2. Comparison of the data on the kringles of both fragment 2 and kringle 4 shows the two proteins to have preserved structural elements, and in particular one of these involves the conserved residues Trp 25, Leu 45 and Trp 61. Furthermore, the two tryptophan residues have been shown to have quite unique spectral charateristics. Brief studies of human prothrombin fragment 2 reveals similar findings. The results discussed indicate that the kringle sequence gives rise to a well defined fold. In kringle 4 of human plasminogen this is seen to provide a highly specific binding site for certain ω-aminocarboxylic acids. The data is all in accord with kringles being autonomous units within the non protease portion of zymogen molecules and which are ideally suited to include the mediation of protein-protein interactions amongst their biological functions.

This thesis describes an investigation of the folding and stability of a series of derivatives of the proteins lysozyme and α-lactalbumin which lack one or more of their four native disulphide bridges. Removal of the disulphide bridge which links the N- and C-termini from hen lysozyme results in a three-disulphide derivative (CM^6,127-lysozyme). This has a profound effect on its stability against thermal denaturation, the T_m for unfolding being reduced by 25°C at pH 3.8. Calorimetric measurements performed on this three-disulphide derivative indicate that this reduction in stability may be attributed entirely to an increase in the entropy difference between the native and denatured states. Kinetic refolding studies of CM^6,127-lysozyme using stopped flow optical methods and hydrogen exchange pulse labelling in conjunction with NMR and electrospray ionisation mass spectrometry (ESI-MS) suggest that this reduced stability manifests itself primarily in the α-domain of the protein. A transient intermediate populated during refolding of the unmodified protein can no longer be detected during folding of the derivative resulting in highly cooperative folding under the conditions investigated. The structure and stability of a three- and two-disulphide derivative of the homologous protein, α-lactalbumin have been investigated by NMR spectroscopy. The three-disulphide species, like its lysozyme counterpart, can adopt native structure but this is much more unstable than the intact protein. Removal of a second disulphide bridge, however, destabilises α-lactalbumin to the extent that the native state is no longer formed. Instead, in the presence of Ca²⁺ and high concentrations of salt, a partially structured state is induced which has some elements of tertiary structure present. Novel techniques of ESI-MS have been developed to study protein folding and stability using hydrogen exchange techniques. Applications to the investigation of cooperativity in protein folding, stability in native, partially folded and unfolded states, and the interactions of a partially folded protein with the chaperone GroEL are described.