Abstract
Aromatic ring flips are a hallmark of protein dynamics. They are experimentally studied by NMR spectroscopy, where recent advances have led to improved characterization across a wide range of time scales. Results on different proteins have been interpreted as continuous diffusive ring rotations or jumplike flips, leading to diverging views of the protein interior as being fluidlike or solidlike, respectively. It is challenging to distinguish between these mechanisms and other types of conformational exchange because chemical-shift-mediated line broadening provides only conclusive evidence for ring flips only if the system can be moved from the slow- to intermediate/fast-exchange regime. Moreover, whenever the chemical shift difference between the two symmetry-related sites is close to zero, it is not generally possible to determine the exchange time scale. Here we resolve these issues by measuring residual dipolar coupling (RDC)-mediated exchange contributions using NMR relaxation dispersion experiments on proteins dissolved in dilute liquid crystalline media. Excellent agreement is found between the experimental difference in RDC between the two symmetry-related sites and the value calculated from high-resolution X-ray structures, demonstrating that dynamics measured for F52 in the B1 domain of protein G reports on distinct, jumplike flips rather than other types of conformational exchange.