Research in the laboratory is centered around enhancing our understanding of protein function from the perspective of physical structural biology.  In past decades, proteins were essentially viewed as static structures.  Today, they are widely appreciated to be dynamic ensembles of interconverting structures.  Such behavior can be clearly seen in proteins that undergo dramatic shape changes in different functional states.  However, the effects of dynamics can also be important when the motions are “less coordinated” structurally and occur within the original protein shape.  The ensemble nature of proteins has far-reaching implications for understanding basic natural protein functions such as ligand binding, enzyme catalysis, and allostery.  By extension, an understanding of protein dynamics should lead to improvements in protein engineering and rational drug design.

We are investigating the role of structural dynamics in protein function for a variety of proteins important in metabolism and signal transduction.  This includes enzymes such as dihydrofolate reductase (DHFR), which is essential in all organisms for nucleotide biosynthesis.  It is also the target for drugs used to treat human cancers and antibiotics and is therefore an excellent system for studying protein-drug interactions.  Another class of proteins in which we are interested is the PDZ (PSD95 - Discs Large - ZO-1) domain.  These are found throughout nature in a variety of contexts and are widespread as signal transduction modules in higher eukaryotes.  PDZ domains specialize in binding C-terminal sequences of signaling enzymes, transmembrane receptors, proteases, etc.  They are also recognized as models for intramolecular communication.  Our NMR studies on PDZ domains implicate dynamics as a viable mechanism for long-range communication and allostery.  Other ongoing projects in the lab are directed at the role of protein dynamics in allostery and their distinguishing features from non-allosteric proteins.

How do we characterize protein dynamics?  Our preferred method is heteronuclear NMR spectroscopy, which is uniquely suited to study both structure and dynamics in proteins and other biological macromolecules. A major advantage of NMR is that spectroscopic probes are distributed uniformly throughout the biomolecule, such as NH or CH atom pairs, providing large amounts of molecular information.  To gain information on protein dynamics, NMR spin relaxation is highly sensitive to molecular motion over a range of timescales.  We look at the relaxation properties of ¹⁵N, ¹³C, and ²H spins located throughout the protein scaffold, and interpret these in terms of amplitudes and timescales of individual bond vectors.  Slower motions on the microsecond-millisecond timescale can be detected to yield site-specific kinetic, thermodynamic, and structural information on the switching between discrete conformational states.  In many cases, these NMR-relaxation dynamics are used to complement other structural data from X-ray crystallography, or thermodynamic and kinetic biophysical measurements using methods such as fluorescence spectroscopy, calorimetry, amide hydrogen exchange, and molecular dynamics simulations.