Theoretical Study of Phosphodiester Hydrolysis in Nucleotide Pyrophosphatase/Phosphodiesterase. Environmental Effects on the Reaction Mechanism

摘要

Abstract: We here present a theoretical study of the alkaline hydrolysis of methyl p-nitrophenyl phosphaten(MpNPP ) in aqueous solution and in the active site of nucleotide pyrophosphatase/phosphodiesterasen(NPP). The analysis of our simulations, carried out by means of hybrid quantum mechanics/molecularnmechanics (QM/MM) methods, shows that the reaction takes place through different reaction mechanismsndepending on the environment. Thus, while in aqueous solution the reaction occurs by means of an ANDNnmechanism, the enzymatic process takes place through a DNAN mechanism. In the first case, we foundnassociative transition-state (TS) structures, while in the enzyme TS structures have dissociative character.nThe reason for this change is rationalized in terms of the very different nature of the electrostatic interactionsnestablished in each of the environments: while the aqueous solution reduces the repulsion between thennegatively charged reacting fragments, assisting their approach, the NPP active site stabilizes the chargendistribution of dissociative TS structures, allowing the reaction to proceed with a significantly reduced freenenergy cost. Interestingly, the NPP active site is able to accommodate different substrates, and it seemsnthat the nature of the TSs depends on their electronic characteristics. So, in the case of the MpNPPnsubstrate, the nitro group establishes hydrogen-bond interactions with water molecules and residues foundnin the outer part of the catalytic site, while the leaving group oxygen atom does not coordinate directly withnany of the zinc atoms of the active site. If methyl phenyl phosphate is used as substrate, then the chargenon the leaving group is supported to larger extent by the oxygen atom and the phenolate anion can benthen coordinated to one of the two zinc atoms present in the active site.