Enzyme Catalysis

The active site is the functionally relevant part of the enzyme. The binding of the substrate and the catalytic rate enhancement depends on the non-covalent interactions and dynamics of the active site. It has been hypothesized that a nominally non-polar protein matrix can produce a huge inhomogeneous electric field similar to that of a polar solvation environment. However, to date, experimental validation of this hypothesis has only been proved for a single protein (ketosteroid isomerase). Vibrational Stark spectroscopy allows us to experimentally predict electric fields exerted by the active site residues along the reaction coordinate (bond dipole of the substrate that is involved in the reaction). Indirect estimation of the local electrostatic at the active site can be made by inserting external IR chromophores (either using unnatural amino acids or post-translational modification of amino acids). In our lab, we are calibrating electric fields inside various proteins to underpin the role of electrostatics towards enzyme activity/protein function.

Similarly, the floppiness of the active site, as well as ultrafast dynamics of a directional hydrogen bond can play a mojor role toward enzyme catalysis. Using 2D-IR spectroscopy we can measure the dynamics of the active site and understand the role of dynamics toward function/activity.