Methyl-specific isotopic labeling: a molecular tool box for solution NMR studies of large proteins

• Published in: Current opinion in structural biology 2015-06, Vol.32, p.113-122
• Main Authors: Kerfah, Rime, Plevin, Michael J, Sounier, Remy, Gans, Pierre, Boisbouvier, Jerome
• Format: Article
• Language: English
• Subjects: Animals; Biochemistry, Molecular Biology; Carbon Isotopes - chemistry; Deuterium - chemistry; Humans; Isotope Labeling - methods; Life Sciences; Membrane Proteins - chemistry; Methylation; Models, Molecular; Nuclear Magnetic Resonance, Biomolecular - methods; Protein Conformation; Proteins - chemistry; Structural Biology
• ISSN: 0959-440X; 1879-033X
• DOI: 10.1016/j.sbi.2015.03.009

•Methyl-specific labeling for solution NMR studies of proteins above 100kDa.•Specific labeling of any combination of methyl groups in protein overexpressed in E. coli.•Detection of NOEs between remote CH3 probes in membrane proteins and large complexes.•Dynamics, interaction and mechanistic investigations of protein machineries. Nuclear magnetic resonance (NMR) spectroscopy is a uniquely powerful tool for studying the structure, dynamics and interactions of biomolecules at atomic resolution. In the past 15 years, the development of new isotopic labeling strategies has opened the possibility of exploiting NMR spectroscopy in the study of supra-molecular complexes with molecular weights of up to 1MDa. At the core of these isotopic labeling developments is the specific introduction of [1H,13C]-labeled methyl probes into perdeuterated proteins. Here, we describe the evolution of these approaches and discuss their impact on structural and biological studies. The relevant protocols are succinctly reviewed for single and combinatorial isotopic-labeling of methyl-containing residues, and examples of applications on challenging biological systems, including high molecular weight and membrane proteins, are presented.