QM/MM Study of a Nucleophilic Substitution Reaction Catalyzed by Uridine Phosphorylase from Vibrio cholerae

Abstract

Uridine phosphorylases are used for biotechnological synthesis of pyrimidine derivatives and, moreover, their substrates and inhibitors are used in medicine. Therefore, studies of the mechanisms of the chemical reaction catalyzed by the enzyme and its specificity for various substrates are relevant. The research into the enzymatic reaction main stage—nucleophilic substitution of the nitrogenous base in uridine with an orthophosphate or orthovanadate group by hybrid QM/MM methods—was carried out. A comparison of various levels of theory and calculation schemes showed that preliminary optimization of the reactants’s geometry, as well as calculation of the initial trajectory of the minimum energy path, can be achieved by semi-empirical methods. At the same time, for the minimum energy path clarification, transition state geometry optimization, and calculation of the thermochemical parameters, it is preferable to use density functional theory in combination with modern ab initio methods. In comparison with the calculations of the activation barrier carried out in a solvent without an enzyme, differences in the kinetics of the enzymatic reaction due to the orientation and concentration actions of amino acid residues of the enzyme were revealed. This led to lowering the activation barrier by 20 kcal/mol and contributed to the reaction under physiologically acceptable conditions. It was shown that the free activation energy during the nucleophilic attack for uridine with hydrovanadate ion is 2 kcal/mol lower than for the hydrophosphate ion and this is consistent with the literature data.