Resonance assignments in proteins

The assignment of resonances in the complex NMR spectrum of a protein is the first step in any study of protein structure, function or dynamics. Prior to 1980, assignment was achieved using 1D NMR, and based, for the most part, on the assumption that the structure of the protein in solution was the same as in the X-ray structure. The introduction of 2D NMR techniques such as COSY and NOESY in the early 1980's dramatically increased the resolution of protein NMR spectra. This led to the development of a systematic method for the assignment of the 2D NMR spectra of proteins that relied only on information about the amino acid sequence of the protein; this method is the sequential assignment procedure. The availability of uniform N labelling and the development of 3D NMR methods in the late 1980’s led to increased resolution in NMR spectra and increased the size of proteins that could be assigned using the sequential assignment methodology. Double labelling with N and C led, in the early 1990’s to the development of an alternative assignment approach based on scalar couplings; this again increased the molecular weight limit for complete assignment. Recently, this triple-resonance approach in conjunction with deuteration and new TROSY methods has increased the molecular weight limit to beyond 30kD. In this chapter the elements of the sequential assignment and the triple-resonance assignment strategies are outlined.