New Methods in Biomolecular NMR Spectroscopy

NMR (Nuclear Magnetic Resonance) is now a routine method in the structure determination of biological macromolecules. NOE (Nuclear Overhauser Effect) and J (scalar) couplings are used in the process of structure determination. NOEs define short distance contacts and J couplings define dihedral angles. However, these constraints are only strictly local in nature and contain no information about long-range order such as relative orientation of protein domains of overall molecular form and shape. Such information is available through dipolar couplings, which unfortunately average to zero in isotropic solution. Magnetic susceptibility alignment or very dilute liquid crystal solution can prevent complete averaging. The resulting residual dipolar couplings can now be readily measured with the advent of higher magnetic fields in NMR. They depend on the orientation of the internuclear vectors relative to the molecular frame and provide the desire long range information. These restraints can be subsequently used in structure calculation to improve the accuracy of protein structures. New techniques to measure dipolar splitting will be applied to the study of structure and dynamics of biomolecules in solution. Two protein Ubiquitin and Calmodulin will be studied in the course of the project.