Chapter 8 Segmental Isotopic Labeling of Proteins for Nuclear Magnetic Resonance

Dongsheng Liu, Rong Xu, David Cowburn

Research output: Chapter in Book/Report/Conference proceedingChapter

32 Scopus citations

Abstract

Nuclear magnetic resonance (NMR) spectroscopy has emerged as one of the principle techniques of structural biology. It is not only a powerful method for elucidating the three-dimensional structures under near physiological conditions but also a convenient method for studying protein-ligand interactions and protein dynamics. A major drawback of macromolecular NMR is its size limitation, caused by slower tumbling rates and greater complexity of the spectra as size increases. Segmental isotopic labeling allows for specific segment(s) within a protein to be selectively examined by NMR, thus significantly reducing the spectral complexity for large proteins and allowing for the application of a variety of solution-based NMR strategies. Two related approaches are generally used in the segmental isotopic labeling of proteins: expressed protein ligation and protein trans-splicing. Here, we describe the methodology and recent application of expressed protein ligation and protein trans-splicing for NMR structural studies of proteins and protein complexes. We also describe the protocol used in our lab for the segmental isotopic labeling of a 50-kDa protein Csk (C-terminal Src kinase) using expressed protein ligation methods.

Original languageEnglish (US)
Title of host publicationMethods in Enzymology
EditorsTom Muir, John Abelson
Pages151-175
Number of pages25
DOIs
StatePublished - 2009

Publication series

NameMethods in Enzymology
Volume462
ISSN (Print)0076-6879

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

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    Liu, D., Xu, R., & Cowburn, D. (2009). Chapter 8 Segmental Isotopic Labeling of Proteins for Nuclear Magnetic Resonance. In T. Muir, & J. Abelson (Eds.), Methods in Enzymology (pp. 151-175). (Methods in Enzymology; Vol. 462). https://doi.org/10.1016/S0076-6879(09)62008-5