Mechanistic Insight into the Catalytic Activity of ββα-Metallonucleases from Computer Simulations: Vibrio vulnificus Periplasmic Nuclease as a Test Case

Using information from wild‐type and mutant Vibrio vulnificus nuclease (Vvn) and I‐PpoI homing endonuclease co‐crystallized with different oligodeoxynucleotides, we have built the complex of Vvn with a DNA octamer and carried out a series of simulations to dissect the catalytic mechanism of this metallonuclease in a stepwise fashion. The distinct roles played in the reaction by individual active site residues, the metal cation and water molecules have been clarified by using a combination of classical molecular dynamics simulations and quantum mechanical calculations. Our results strongly support the most parsimonious catalytic mechanism, namely one in which a single water molecule from bulk solvent is used to cleave the phosphodiester bond and protonate the 3′‐hydroxylate leaving group. Parsimony in water splitting: Although two water molecules are usually depicted in the catalytic mechanism of ββα‐metallonucleases, one to cleave the phosphodiester bond and another to protonate the leaving hydroxylate, our computational results strongly suggest that it is the same water molecule from bulk solvent that provides both the nucleophilic hydroxide and the proton necessary to complete the reaction (see scheme).