Amino acid selective unlabeling in protein NMR spectroscopy.

Three-dimensional structure determination of proteins by NMR requires the acquisition of multidimensional spectra followed by assignment of chemical shifts to the respective nuclei. In order to speed up this process, resonances corresponding to individual amino acid types are often selectively identified and assigned. One of the ways of achieving this is by using the method of "selective unlabeling." In this method, resonances from one or more amino acid types are suppressed selectively in the NMR spectra, which can be achieved using both cell-based and cell-free methods. This helps not only in identifying them but also results in spectral simplification by reducing the number of peaks observed. Further, the assignments are not limited to amino acids that are specifically unlabeled. Using specially designed NMR experiments, assignments of amino acids in the neighborhood of those being selectively unlabeled can also be obtained. In this chapter, we discuss the theoretical and practical aspects of selective unlabeling focusing on how the sample is prepared, which amino acid or a combination of amino acids should be optimally chosen for unlabeling, and how this method can be used for sequential assignments of proteins.