Global Dynamics and Exchange Kinetics of a Protein on the Surface of Nanoparticles Revealed by Relaxation-Based Solution NMR Spectroscopy

The global motions and exchange kinetics of a model protein, ubiquitin, bound to the surface of negatively charged lipid-based nanoparticles (liposomes) are derived from combined analysis of exchange lifetime broadening arising from binding to nanoparticles of differing size. The relative contributions of residence time and rotational tumbling to the total effective correlation time of the bound protein are modulated by nanoparticle size, thereby permitting the various motional and exchange parameters to be determined. The residence time of ubiquitin bound to the surface of both large and small unilamellar liposomes is similar to 20 mu s. Bound ubiquitin undergoes internal rotation about an axis approximately perpendicular to the lipid surface on a low microsecond time scale (similar to 2 mu s), while simultaneously wobbling in a cone of semiangle 30-55 degrees centered about the internal rotation axis on the nanosecond time scale. The binding, interface of ubiquitin with liposomes is mapped by intermolecular paramagnetic relaxation enhancement using Gd3+-tagged vesicles, to a predominantly positively charged surface orthogonal to the internal rotation axis.