Energetics of the Catalytic Reaction of Ribonuclease A: A Computational Study of Alternative Mechanisms

Abstract

The energetics of the catalytic reaction of ribonuclease A have been explored with empirical valence bond (EVB) simulations to elucidate the origin of the enormous catalytic power of this enzyme and to examine different mechanistic alternatives. The two mechanisms analyzed were a general acid-general base mechanism with a dianionic transition state and a triester-like mechanism with a monoionic transition state. The first step of the analysis used experimental information to determine the activation energy of each assumed mechanism in a water cage. This has provided an experimentally based reference point for the catalytic effect of the enzyme. The next step of the analysis involved EVB simulations of the reaction in water and calibrated these simulations against the above-mentioned energetics of the reference reaction. The simulations were performed in the protein environment without changing any EVB parameters. The catalysis was measured as the difference in the overall activation free energy of the respective mechanism in water and in protein. In the mechanism with the dianionic transition state a catalytic effect of ∼18 ( 6 kcal/mol was established which is in good agreement with the experimentally derived estimate of ∼21 kcal/mol. In the mechanism with the monoionic transition state, a value of ∼12 ( 6 kcal/mol was calculated which is 5 kcal/mol lower than its estimated value, making it slightly less likely to be the actual mechanism in the enzyme. The origin of the catalytic power is attributed to an electrostatic reduction of the activation barriers. This reduction is associated with the preorganized polar environment of the enzyme.

Molecular mechanism of the site-specific self-cleavage of the RNA phosphodiester backbone by a twister ribozyme

“…The activation free energy of the rate-limiting step in aqueous solution is in agreement with the values reported by Glennon and Warshel [38] that combine experimental data of Wolfenden et al [39] and computational values for the transesterification reactions. The proposal that the process takes place by a stepwise mechanism controlled by the second transition state can be supported by KIEs calculations.…”

Ribonucleases: From Prototypes to Therapeutic Targets? (General Articles)

“…The protons which are transferred from the 2'-OH group to the catalytic base and from the catalytic acid to the leaving 5'-oxygen are shown 'in flight', in line with a proton inventory study on the hydrolysis step catalyzed by RNase A [23]. Whereas the general acid/base reaction scheme was introduced initially four decades ago [24], considerable debate still persists about the details of the catalyzed reactions, notably the anionic character of the transition state [25,26]. The likelihood of various mechanistic proposals has been tested most elaborately for bovine pancreatic RNase A [27] and RNase T 1 (EC 3.1.27.3) secreted by the slime mold Aspergillus oryzae [28,29].…”

The hospitality/tourism academic leader as catalytic agent: a model for sustainable institutional change

“…The term ''catalytic mechanism'' is used in science to describe the initiation of a transition state (catalytic) with a resultant unpredicted outcome (mechanism) (Glennon and Warshel, 1998). Collins (1999) applies the scientific meaning to organizations by describing catalytic mechanisms as critical links between objectives and actual performance.…”

“…When applied to organizations, the intent for change will create a state that facilitates ''change agentry'' in organizations (Blanch, 1999). The literature in chemistry suggests that catalytic mechanisms (caused by catalytic agents) act as stimulants that affect changes in other mechanisms, with the degree of change being measured by the activation of free energy during the reaction, otherwise stated as ''change agentry'' (Glennon and Warshel, 1998).…”