Aromatic residues contribute a significant part (roughly 25% of the volume on average) of the hydrophobic core of proteins, and are prevalent in protein binding interfaces. Histidine and tyrosine also play prominent roles in enzyme catalysis. For these reasons, we are interested in studying the dynamics of aromatic side chains. We have developed methods to measure aromatic side-chain dynamics on both fast (ps–ns) and slow (μs–ms) time scales.
We have applied these methods to study protein folding, ring-flip dynamics and other types of conformational exchange, as well as the change in fluctuation amplitudes upon ligand binding.
Current knowledge indicate that ring flips can take place on a wide range of time scales, from nanosecond in exposed and loosely packed sites, to hundreds of milliseconds in the densely packed interior of proteins. In the latter case, ring flips occur when the surrounding protein core undergoes transient "breathing" motions in a concerted fashion. Studies of ring flip dynamics can provide us with a rich picture of the conformational fluctuations in terms of rate constants, energy barriers and activation volumes. Actual measurements of ring flip rates have been carried out in only a handful of cases. Our recent work has identified relatively slow flip rates for several rings in BPTI that were previously believed to flip very fast.
Our NMR experiments rely on site-specific isotope enrichment to yield isolated 13C nuclei. We are working on improved approaches to generate site-specific labeling using bacterial expression on defined media.