Solid-state NMR protocols for unveiling dynamics and (drug) interactions of membrane-bound proteins

Magic angle spinning solid-state NMR (MAS ssNMR) is a versatile tool for studying the structure and dynamics of membrane proteins, as well as their interactions with ligands and drugs. Its power lies in the ability to provide atomic-level information on samples under physiological-like conditions. Moreover, it can illuminate dynamics across a wide range of timescales with great relevance to membrane protein function and dysfunction. In this protocol paper, we highlight key aspects of sample preparation, data acquisition, and interpretation, based on our own experience and the broader literature. We discuss key protocol steps along with important considerations for sample preparation and parameters for ssNMR measurements, with reference to the special requirements of membrane-based samples. Such samples display physiologically relevant dynamics across different motional regimes that can be probed by NMR but also can interfere with certain NMR measurements. We guide the reader through the whole process from sample preparation to complex NMR characterization techniques. Throughout the report, we refer back to examples from our own prior work on the interactions between cytochrome c and cardiolipin-containing membranes, with a discussion of the lipid dependence and interactions with a peroxidase-activity inhibitor. We conclude with a short discussion of alternative and new methods that are further boosting the power and versatility of ssNMR as a tool to study membrane-bound proteins and their ligands or drug interactions.