Methyl Groups as Probes for Proteins and Complexes in In-Cell NMR Experiments

Studying protein components of large intracellular complexes by in-cell NMR has so far been impossible because the backbone resonances are unobservable due to their slow tumbling rates. We describe a methodology that overcomes this difficulty through selective labeling of methyl groups, which possess more favorable relaxation behavior. Comparison of different in-cell labeling schemes with three different proteins, calmodulin, NmerA, and FKBP, shows that selective labeling with [13C]methyl groups on methionine and alanine provides excellent sensitivity with low background levels at very low costs.