Segmental isotopic labeling of samples for NMR studies is attractive for large complex biomacromolecular systems, especially for studies of function-related protein-ligand interactions and protein dynamics (Goto and Kay, Curr Opin Struct Biol 10:585-592, 2000; Rosa et al., Molecules (Basel, Switzerland) 18:440, 2013; Hiroaki, Expert Opin Drug Discovery 8:523-536, 2013). Advantages of segmental isotopic labeling include selective examination of specific segment(s) within a protein by NMR, significantly reducing the spectral complexity for large proteins, and allowing for the application of a variety of solution-based NMR strategies. By utilizing intein techniques (Wood and Camarero, J Biol Chem 289:14512-14519, 2014; Paulus, Annu Rev Biochem 69:447-496, 2000), two related approaches can generally be used in the segmental isotopic labeling of proteins: expressed protein ligation (Muir, Annu Rev Biochem 72:249-289, 2003) and protein trans-splicing (Shah et al., J Am Chem Soc 134:11338-11341, 2012). Here, we describe general implementation and latest improvements of expressed protein ligation method for the production of segmental isotopic labeled NMR samples.
Keywords: Expressed protein ligation; Intein; Isotopic labeling; NMR; Structural biology.