This proposal represents a multinuclear, multifield, and multiphase nmr investigation of systems of biological interest. The principal nucleus to be employed is cadmium-113. These experiments are designed to exploit the ability of cadmium to replace calcium and zinc in biological systems. Further, by utilizing the favorable magnetic resonance parameters of 113Cd, we plan to monitor the local environment of the metal binding site with respect to its contiguous ligands and its local dynamics. By virtue of its own inorganic chemistry, cadmium's nmr parameters are often averages over several chemically exchanging environments. To circumvent this """"""""problem"""""""" we propose to study cadmium's nmr parameters in the liquid state, supercooled aqueous emulsions, and in the solid state. With such an approach we will develop a firm understanding of the details of these dynamic processes and how these processes modulate 113Cd nmr parameters (coupling constants, relaxation times, isotropic chemical shift and the shielding tensor). This methodology will be applied to two systems: i) model systems and ii) metalloproteins. The model systems include single crystals of cadmium coordination compounds, prosthetic groups such as protoporphyrin IX, variously substituted tetraphenyl porphyrins, and a feasibility study of utilizing 67Zn, 63Cu, and 95Mo in the solid state. The metalloproteins of interest are Concanavalin A (ConA), Carboxypeptidase-A (CPA) and Hemoglobin (Hb). Experiments on ConA and CPA will help us understand the nature of the disorder (static or dynamic) we observe in the 113Cd resonances of these systems in the solid state. Proposed experiments on Cd2+ substituted Hb will demonstrate the feasibility of utilizing 113Cd nmr to study subunit interactions and cooperativity in Fe3Cd Hb. The present application also demonstrates that certain solid state nmr experiments can succeed where liquid state experiments fail, e.g. in the observation of 113Cd resonances in free 113Cd-CPA and with quadrupolar nuclides with I=3/2, 5/2, 7/2 and 9/2. We will exploit this observation and investigate the possibility of employing cross-polarization with enrichment to observe 67Zn nmr in CPA. Analogous experiments are planned for 63Cu and 59Mo nmr. These experiments should provide important cross checks on the 113Cd experiments and open another vista with regard to metal nuclide nmr in general.
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