Phosphate Binding Energy and Catalysis by Small and Large Molecules

Publisher Website
Google Scholar
Abstract
Catalysis is an important process in chemistry and enzymology. The rate acceleration for any catalyzed reaction is the difference between the activation barriers for the uncatalyzed (ΔGHO⧧) and catalyzed (ΔGMe⧧) reactions, which corresponds to the binding energy (ΔGS⧧ = ΔGMe⧧ − ΔGHO⧧) for transfer of the reaction transition state from solution to the catalyst. This transition state binding energy is a fundamental descriptor of catalyzed reactions, and its evaluation is necessary for an understanding of any and all catalytic processes. We have evaluated the transition state binding energies obtained from interactions between low molecular weight metal ion complexes or high molecular weight protein catalysts and the phosphate group of bound substrate. Work on catalysis by small molecules is exemplified by studies on the mechanism of action of Zn2(1)(H2O). A binding energy of ΔGS⧧ = −9.6 kcal/mol was determined for Zn2(1)(H2O)-catalyzed cleavage of the RNA analogue HpPNP. The pH−rate profile for this cleavage reaction showed that there is optimal catalytic activity at high pH, where the catalyst is in the basic form [Zn2(1)(HO−)]. However, it was also shown that the active form of the catalyst is Zn2(1)(H2O) and that this recognizes the C2-oxygen-ionized substrate in the cleavage reaction. The active catalyst Zn2(1)(H2O) shows a high affinity for oxyphosphorane transition state dianions and a stable methyl phosphate transition state analogue, compared with the affinity for phosphate monoanion substrates. The transition state binding energies, ΔGS⧧, for cleavage of HpPNP catalyzed by a variety of Zn2+ and Eu3+ metal ion complexes reflect the increase in the catalytic activity with increasing total positive charge at the catalyst. These values of ΔGS⧧ are affected by interactions between the metal ion and its ligands, but these effects are small in comparison with ΔGS⧧ observed for catalysis by free metal ions, where the ligands are water. Enzymes are unique in having evolved mechanisms to effectively utilize binding interactions with nonreacting fragments of the substrate in stabilization of the reaction transition state. Orotidine 5′-monophosphate decarboxylase, α-glycerol phosphate dehydrogenase, and triosephosphate isomerase catalyze dissimilar decarboxylation, hydride transfer, and proton transfer reactions, respectively. Each enzyme derives ca. 12 kcal/mol of transition state stabilization from protein interactions with the nonreacting phosphate group, which is larger than the highest ∼10 kcal/mol transition state stabilization that we have determined for small-molecule catalysis of phosphate diester cleavage in water. Each of these enzymes catalyze the slow reaction of a truncated substrate that lacks the phosphate group, and in each case, the reaction of the truncated substrate is strongly activated by the allosteric binding of the second substrate “piece” phosphite dianion, HPO32−. We propose a modular design for these enzymes with a classical active site that recognizes the reactive substrate fragment and a separate phosphodianion binding site. The second site is created, in part, by flexible protein loops that wrap around the substrate phosphodianion group and bury the substrate in an environment with an optimal local dielectric constant for the catalyzed reaction and with the most favorable positioning of the catalytic side chains. This design is easily generalized to a wide variety of enzyme-catalyzed reactions.

This publication has 49 references indexed in Scilit: