title: Segmental Isotopic Labeling of Proteins for NMR Study Using Intein Technology.
abstract: 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.
authors.affiliation: iHuman Institute, ShanghaiTech University, 99 Haike Road, Pudong, Shanghai, 201203, China.
authors.forename: Dongsheng
authors.initials: D
authors.lastname: Liu
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY, 10461, USA. david.cowburn@einstein.yu.edu.
authors.forename: David
authors.initials: D
authors.lastname: Cowburn
mesh_terms: D047668:Inteins; D007553:Isotope Labeling / Q000379:methods*; D019906:Nuclear Magnetic Resonance, Biomolecular; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics
chemical_list: D011993:Recombinant Fusion Proteins
keywords: Expressed protein ligation; Intein; Isotopic labeling; NMR; Structural biology