Testing Electrostatic Complementarity in Enzyme Catalysis: Hydrogen Bonding in the Ketosteroid Isomerase Oxyanion Hole
Figure 6
Schematic Depiction of an Experimental Method to Isolate and Probe the Role of Electrostatic Complementarity to the Transition State
(A) In the reaction to be probed there is an accumulation of negative charge in the transition state. Two phenolates with different substituents (X and Y) and therefore different p Ka values and different charge densities on the oxygen atom are used to mimic the increase in charge localization going from the ground state to the transition state. As charge localization on the oxygen increases, hydrogen bonds may strengthen in both water and in the enzyme active site (larger dots for hydrogen bonds formed by phenolate Y). The equilibria and represent the exchange of one phenolate for the other in water and on the enzyme, respectively, mimicking the change in charge localization along the reaction coordinate. The equilibria KX and KY are the affinities of the two phenolates, i.e., the equilibrium for their transfer from water to the enzyme.
(B) The log of the affinities of the substituted phenolates from (A) are plotted versus p Ka. If the phenolate with substituent Y binds more strongly than that with substituent X (red; KY > KX), then the enzyme is better than water at stabilizing increased charge localization and there is a favorable contribution to catalysis from electrostatic complementarity. The sign and steepness of the slope, as established using a series of substituted phenolates, can determine the sign and magnitude of the contribution to catalysis.