<div><p>Electrostatic interactions play a pivotal role in
enzymatic catalysis and are increasingly modeled explicitly in computational
enzyme design; nevertheless, they are challenging to measure experimentally.
Using vibrational Stark effect (VSE) spectroscopy, we have measured electric
fields inside the active site of the enzyme ketosteroid isomerase (KSI). These studies have
shown that these fields can be unusually large, but it has been unclear to what
extent they specifically stabilize the transition state (TS) relative to a
ground state (GS). In the following, we use crystallography and computational
modeling to show that KSI’s intrinsic electric field is nearly perfectly
oriented to stabilize the geometry of its reaction’s TS. Moreover, we find that this electric field
adjusts the orientation of its substrate in the ground state so that the
substrate needs to only undergo minimal structural changes upon activation to
its TS. This work provides evidence that
the active site electric field in KSI is preorganized to facilitate catalysis
and provides a template for how electrostatic preorganization can be measured
in enzymatic systems. <br></p></div>
Solvation Response along the Reaction Coordinate in the Active Site of Ketosteroid Isomerase
