Role of magnesium ions in DNA recognition by the EcoRV restriction endonuclease

The restriction endonuclease EcoRV binds two magnesium ions. One of these ions, Mg-A(2+), binds to the phosphate group where the cleavage occurs and is required for catalysis, but the role of the other ion, Mg-B(2+) is debated. Here, multiple independent molecular dynamics simulations suggest that Mg-B(2+) is crucial for achieving a tightly bound protein-DNA complex and stabilizing a conformation that allows cleavage. In the absence of Mg-B(2+) in all simulations the protein-DNA hydrogen bond network is significantly disrupted and the sharp kink at the central base pair step of the DNA, which is observed in the two-metal complex, is not present. Also, the active site residues rearrange in such a way that the formation of a nucleophile, required for DNA hydrolysis, is unlikely. (C) 2011 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.