| Xenon-binding sites in proteins have led to several applications of xenon in biochemical and structural studies. The sensitivity of the 129Xe chemical shift and the intense signals attainable by optical pumping make xenon a useful nuclear magnetic resonance (NMR) probe of local chemical environments. The influence of xenon-protein interactions on 129Xe magnetic resonance properties, particularly the chemical shift, is studied here, and several applications to biomolecular assays are discussed. In this dissertation, 129Xe NMR data of xenon dissolved in aqueous solution with a number of amino acids, peptides, and proteins under both native and denaturing conditions are presented. It is shown that both the 129Xe chemical shift and spin-lattice relaxation rate are affected by weak, nonspecific interactions between xenon and proteins in solution. Nonspecific interactions generally induce shifts that are downfield relative to the frequency of xenon in water; the magnitude of these effects depends on properties of the protein such as size, charge, and chemical functionality. In native proteins, contributions to the observed 129Xe chemical shift are demonstrated to arise from the presence of specific binding interactions at cavities within the protein structure. The effect of these interactions on the shift depends on binding affinity and the size of the cavity. Comparison of chemical shift trends under native and denaturing conditions are used to predict the presence of specific binding sites. The chemical shift of xenon in solution with the maltose binding protein is shown to be sensitive to the conformation of the protein, and structural analysis indicates that this sensitivity results from the presence of a xenon binding site near the maltose-binding cleft. The sensitivity of the 129Xe chemical shift to protein conformation indicates that xenon can be used to assay protein functional states and ligand-binding interactions. In addition, a method for using encapsulated xenon in a ligand-binding assay is presented. |
