Transition state analogues for nucleotidyl transfer reactions:: Structure and stability of pentavalent vanadate and phosphate ester dianions

The structures and energy of phosphate dimethyl ester and vanadate dimethyl ester have been calculated using B3LYP/TZVP density functional quantum chemical methods and polarized continuum (PCM) and Langevin dipoles solvation models. These calculations were carried out to obtain fundamental information on the ability of vanadate esters to function as transition state analogues for the nucleotidyl transfer reaction catalyzed by DNA polymerases. Base-catalyzed methanolysis of the phosphate and vanadate dimethyl esters were the model reactions examined in this study. The structures of the phosphate and vanadate dimethyl esters and pentavalent intermediates in aqueous solution were optimized and evaluated at the PCM/B3LYP/TZVP level. The three-dimensional free energy surfaces for the studied reactions were determined at the PCM/B3LYP/TZVP//B3LYP/TZVP level. Comparison with experimental structural data obtained from the Cambridge Structural Database and with the observed kinetics of phosphate diester hydrolysis demonstrated that the level of theory chosen for these studies was appropriate. The results showed that structurally and electrostatically the vanadate dimethylester and a five-coordinate nearly trigonal bipyramidal intermediate were reasonable analogues for the parent phosphorus systems. Despite these similarities in structure, the energetics of the two systems were different, and the transition states of the two model reactions were found on different areas of the potential energy surface. When the binding energy of a transition state-DNA polymerase complex was extrapolated to a transition state analogue-DNA polymerase complex, the formation of a simple dianionic pentavalent vanadate ester adduct in the enzyme active site was not found to be sufficiently favorable. This finding suggests that additional stabilization of this adduct is needed before this type of transition state analogue will be likely to yield stable adducts with this class of enzymes. New possible candidates for such complexes are suggested.