How Much Do Enzymes Really Gain by Restraining Their Reacting Fragments?

Abstract

The steric effect, exerted by enzymes on their reacting substrates, has been considered as a major factor in enzyme catalysis. In particular, it has been proposed that enzymes catalyze their reactions by pushing their reacting fragments to a catalytic configuration which is sometimes called near attack configuration (NAC). This work uses computer simulation approaches to determine the relative importance of the steric contribution to enzyme catalysis. The steric proposal is expressed in terms of well defined thermodynamic cycles that compare the reaction in the enzyme to the corresponding reaction in water. The S N2 reaction of haloalkane dehalogenase from Xanthobacter autotrophicus GJ10, which was used in previous studies to support the strain concept is chosen as a test case for this proposal. The empirical valence bond (EVB) method provides the reaction potential surfaces in our studies. The reliability and efficiency of this method make it possible to obtain stable results for the steric free energy. Two independent strategies are used to evaluate the actual magnitude of the steric effect. The first applies restraints on the substrate coordinates in water in a way that mimics the steric effect of the protein active site. These restraints are then released and the free energy associated with the release process provides the desired estimate of the steric effect. The second approach eliminates the electrostatic interactions between the substrate and the surrounding in the enzyme and in water, and compares the corresponding reaction profiles. The difference between the resulting profiles provides a direct estimate of the nonelectrostatic contribution to catalysis and the corresponding steric effect. It is found that the nonelectrostatic contribution is about -0.7 kcal/mol while the full "apparent steric contribution" is about -2.2 kcal/mol. The apparent steric effect includes about -1.5 kcal/mol electrostatic contribution. The total electrostatic contribution is found to account for almost all the observed catalytic effect (∼-6.1 kcal/mol of the -6.8 calculated total catalytic effect). Thus, it is concluded that the steric effect is not the major source of the catalytic power of haloalkane dehalogenase. Furthermore, it is found that the largest component of the apparent steric effect is associated with the solvent reorganization energy. This solvent-induced effect is quite different from the traditional picture of balance between the repulsive interaction of the reactive fragments and the steric force of the protein.

Transition state theory can be used in studies of enzyme catalysis: lessons from simulations of tunnelling and dynamical effects in lipoxygenase and other systems

“…Such compression will in fact reduce the tunnelling contributions, since the largest contribution to the KIE comes from ðS 2 00 Þ D =ðS 2 00 Þ H , whose value decreases drastically when D H and u H decrease. Furthermore, as we have shown repeatedly, enzymes are flexible and unlikely to be able to change drastically the reaction surface (Warshel 1991;Shurki et al 2002). Thus, it is quite likely that the main difference between reactions with large tunnelling and small tunnelling corrections is the intrinsic shape of the potential surface rather than the effect of the environment on this surface.…”

Perspective on Diabatic Models of Chemical Reactivity as Illustrated by the Gas-Phase SN2 Reaction of Acetate Ion with 1,2-Dichloroethane

“…We conclude that additional termssnot included in our calculationsswhose parameters were not published are needed to reproduce the potential underlying the published EVB calculations of refs 2-4. We therefore retract all quantitative results presented in our paper for the EVB potential of ref 4.…”

A new paradigm for electrostatic catalysis of radical reactions in vitamin B 12 enzymes

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