A solid-state NMR tool box for the investigation of ATP-fueled protein engines

[Display omitted] •Solid-state NMR approaches to assign and investigate large proteins and complexes thereof.•ATP-mimics to study ATP hydrolysis.•Conformational and dynamic changes during enzymatic reaction cycles monitored by NMR.•Protein-nucleotide contacts probed by solid-state NMR spectroscopy.•Fast Magic-Angle spinning and proton-detection to identify protons involved in noncovalent interactions. Motor proteins are involved in a variety of cellular processes. Their main purpose is to convert the chemical energy released during adenosine triphosphate (ATP) hydrolysis into mechanical work. In this review, solid-state Nuclear Magnetic Resonance (NMR) approaches are discussed allowing studies of structures, conformational events and dynamic features of motor proteins during a variety of enzymatic reactions. Solid-state NMR benefits from straightforward sample preparation based on sedimentation of the proteins directly into the Magic-Angle Spinning (MAS) rotor. Protein resonance assignment is the crucial and often time-limiting step in interpreting the wealth of information encoded in the NMR spectra. Herein, potentials, challenges and limitations in resonance assignment for large motor proteins are presented, focussing on both biochemical and spectroscopic approaches. This work highlights NMR tools available to study the action of the motor domain and its coupling to functional processes, as well as to identify protein-nucleotide interactions during events such as DNA replication. Arrested protein states of reaction coordinates such as ATP hydrolysis can be trapped for NMR studies by using stable, non-hydrolysable ATP analogues that mimic the physiological relevant states as accurately as possible. Recent advances in solid-state NMR techniques ranging from Dynamic Nuclear Polarization (DNP), 31P-based heteronuclear correlation experiments, 1H-detected spectra at fast MAS frequencies >100 kHz to paramagnetic NMR are summarized and their applications to the bacterial DnaB helicase from Helicobacter pylori are discussed.