The intrinsic conformational plasticity of membrane proteins directly influences the magnitude of the orientational-dependent NMR interactions such as dipolar couplings (DC) and chemical shift anisotropy (CSA). As a result, the conventional cross-polarization (CP)-based techniques mainly capture the more rigid regions of membrane proteins, while the most dynamic regions are essentially invisible. Nonetheless, dynamic regions can be detected using experiments in which polarization transfer takes place via J-coupling interactions. Here, we review our recent efforts to develop single and dual acquisition pulse sequences with either 1 H or 13 C detection that utilize both DC and J-coupling mediated transfer to detect both rigid and mobile regions of membrane proteins in native-like lipid environments. We show the application of these new methods for studying the conformational equilibrium of a single-pass membrane protein, phospholamban, which regulates the calcium transport across the sarcoplasmic reticulum (SR) membrane by interacting with the SR Ca 2+ -ATPase. We anticipate that these methods will be ideal to portray the complex dynamics of membrane proteins in their native environments.
Bibliographical noteFunding Information:
This work is supported by the National Institutes of Health (GM 64742). The NMR experiments were carried out at the Minnesota NMR Center.
- Conformationally excited states
- H detection
- Magic angle spinning
- Membrane proteins
- Solid-State NMR